title: Dodder_Pumpkin_Rep2

city: Blacksburg

first: James

middle: H

last: Westwood

state: VA

address: 401 Latham Hall

department: Plant Pathol. Physiol. Weed Sci.

institute: Virginia Tech

description: The objective of this experiment was to assay for movement of mRNA from tomato (host) into dodder (parasitic plant).  Dodder was grown on pumpkin as one of several control hosts to facilitate identification of endogenous dodder mRNA from those potentially translocated from the .

hyb_protocol: Hybridization was conducted according to Affymetrix standard procedure and performed by the Virginia Bioinformatics Institute Core Laboratory.

scan_protocol: Scanning was conducted according to Affymetrix standard procedure and performed by the Virginia Bioinformatics Institute Core Laboratory.

data_processing: Data was not processed beyond the values provided by the Affymetrix intensity values.

label: Biotin

organism: Cuscuta pentagona

source_name: Dodder grown on pumpkin replicate 2

label_protocol: A linear amplification of total RNA was performed using Ovation Biotin RNA Amplification and Labeling System (NuGEN, San Carlos, CA).

growth_protocol: Pumpkin (Curcurbita maxima, Blue Hubbard Winter Squash, Heirloom Seeds, West Elizabeth, PA) plants were grown in Metro Mix 360 potting medium (Griffin Greenhouse and Nursery Supplies, Richmond, VA) under greenhouse conditions.

extract_protocol: RNA (1-1.5 g) was extracted from dodder by grinding in liquid nitrogen, and extracted using TRIzol reagent (Invitrogen, Carlsbad, CA).  The RNA was precipitated with 100% isopropanol followed by a second precipitation with 100% ethanol and 3M sodium acetate (pH 5.2).  The RNA pellet was dissolved in DEPC-treated water and treated with DNase using a DNA-free Kit (Ambion, Austin, TX).

treatment_protocol: Lespedeza dodder (Cuscuta pentagona) seeds were scarified by soaking in concentrated sulfuric acid for one hour, rinsed thoroughly in water, and planted around the base of two-week old tomato plants. Dodder stems were trained from tomato to four-week old pumpkin plants and dodder connections between tomato and pumpkin were severed two weeks after the formation of new haustorial connections with pumpkin. Dodder stems at least 4 cm from the host plant were harvested after 9-12 weeks of growth.  To obtain sufficient quantity of dodder tissue, many stems were harvested, representing multiple individuals for each harvest time.  All tissues were immediately frozen in liquid nitrogen and stored at -80ºC.

tag: Cuscuta pentagona


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, miR_16_mm19,20, 100 nM, 24 hr

status: Public on Feb 01 2007

type: RNA

city: Boston

first: Julja

last: Burchard

state: MA

address: 33 Avenue Louis Pasteur

department: Computational & Systems Biology

country: USA

institute: Merck & Co., Inc.

description: Linsley PS, Schelter J, Burchard J, Kibukawa M, Martin MM, Bartz SR, Johnson JM, Cummins JM, Raymond CK, Dai H, Chau N, Cleary M, Jackson AL, Carleton M, and Lim L.  MCB (2007).

hyb_protocol: Microarrays are incubated at 40°C for 48 hours in a rotating carousel.  Hybridizations to custom Agilent microarrays are completed as previously described (Hughes et al.Nat Biotech (2001), 19(4):342-7).  Microarrays are washed to remove non-specific hybridized sample. Afterwards, microarrays are dried in an ozone-free nitrogen chamber.

scan_protocol: Microarrays are scanned using the Agilent LP2 laser scanner. The scanner output is a Tiff file, which contains the quantitative hybridization data from each individual microarray. The Tiff files are then processed using Rosetta custom feature extraction software.

data_processing: Data were processed using the Rosetta Resolver® system. Rosetta's custom feature extraction software performs error modeling before data are loaded into the Resolver system. The Resolver system performs a squeeze operation that combines replicates of the same sequence in an array while applying error weighting. The error weighting consists of adjusting for additive and multiplicative noise. A P-value is generated and propagated throughout the system. The P-value represents the probability that a gene is expressed. The Resolver system allows users to set thresholds, below which genes of a P-value are considered to be significantly expressed. The Resolver system also combines multiple arrays using a squeezing process. If multiple spots reference one sequence, summarization is performed using an error-weighted average as described in Roland Stoughton and Hongyue Dai, Statistical Combining of Cell Expression Profiles. US Patent #6,351,712, February 26, 2002.

molecule: total RNA

organism: Homo sapiens

source_name: HCT116 Dicer -/- #2

label_protocol: custom automated version of the aminoallyl MessageAmp II kit from Ambion.

extract_protocol: Qiagen RNeasy spin columns with DNAse treatment

tag: HCT116 Dicer -/- #2, mock, 24 hr


title: HCT116 Dicer -/- #2, mock vs. HCT116 Dicer -/- #2, miR_195, 100 nM, 24 hr

tag: HCT116 Dicer -/- #2, mock


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, miR_16_mm18,19, 100 nM, 24 hr


title: DLD-1 Dicer -/- #6, mock, 24 hr vs. DLD-1 Dicer -/- #6, hsa-miR-20, 25 nM, 24 hr

source_name: DLD-1 Dicer -/- #6

tag: DLD-1 Dicer -/- #6, mock, 24 hr


title: Dodder_Tobacco

description: The objective of this experiment was to assay for movement of mRNA from tomato (host) into dodder (parasitic plant).  Dodder was grown on tobacco as one of several control hosts to facilitate identification of endogenous dodder mRNA from those potentially translocated from the .

source_name: Dodder grown on tobacco

growth_protocol: Tobacco (Nicotiana tabacum) plants were grown in Metro Mix 360 potting medium (Griffin Greenhouse and Nursery Supplies, Richmond, VA) under greenhouse conditions.

treatment_protocol: Lespedeza dodder (Cuscuta pentagona) seeds were scarified by soaking in concentrated sulfuric acid for one hour, rinsed thoroughly in water, and planted around the base of seven-week old tobacco plants.   Dodder stems at least 4 cm from the host plant were harvested after 9-12 weeks of growth.  To obtain sufficient quantity of dodder tissue, many stems were harvested, representing multiple individuals for each harvest time.  All tissues were immediately frozen in liquid nitrogen and stored at -80ºC.


title: DLD-1 Dicer -/- #6, mock, 24 hr vs. DLD-1 Dicer -/- #6, hsa-let-7c, 25 nM, 24 hr


title: Dodder_Tomato_Rep2

description: The objective of this experiment was to assay for movement of mRNA from tomato (host) into dodder (parasitic plant).  RNA from dodder growing on tomato was hybridized to the tomato array to dected tomato-specific transcripts in the parasite.

source_name: Dodder grown on tomato replicate 2

growth_protocol: Tomato plants supporting the dodder were grown in Metro Mix 360 potting medium (Griffin Greenhouse and Nursery Supplies, Richmond, VA) under greenhouse conditions.

extract_protocol: RNA (1-1.5 g) was extracted from tomato by grinding in liquid nitrogen, and extracted using TRIzol reagent (Invitrogen, Carlsbad, CA).  The RNA was precipitated with 100% isopropanol followed by a second precipitation with 100% ethanol and 3M sodium acetate (pH 5.2).  The RNA pellet was dissolved in DEPC-treated water and treated with DNase using a DNA-free Kit (Ambion, Austin, TX).

treatment_protocol: Lespedeza dodder (Cuscuta pentagona) seeds were scarified by soaking in concentrated sulfuric acid for one hour, rinsed thoroughly in water, and planted around the base of two-week old tomato seedlings.   Dodder stems at least 4 cm from the host plant were harvested after 9-12 weeks of growth.  To obtain sufficient quantity of dodder tissue, many stems were harvested, representing multiple individuals for each harvest time.  All tissues were immediately frozen in liquid nitrogen and stored at -80ºC.


title: Dodder_Tomato_Rep1

source_name: Dodder grown on tomato


title: Dodder_Pumpkin_Rep1

source_name: Dodder grown on pumpkin replicate 1


title: Dodder_Tomato_Rep3

source_name: Dodder grown on tomato replicate 3


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-let-7c, 100 nM, 24 hr


title: DLD-1 Dicer -/- #6, mock, 24 hr vs. DLD-1 Dicer -/- #6, hsa-miR-17-5p, 25 nM, 24 hr


title: Tomato_Rep1

description: The objective of this experiment was to assay for movement of mRNA from tomato (host) into dodder (parasitic plant).  This tomato sample served as a control to establish a profile of expressed genes in the host.

organism: Solanum lycopersicum

source_name: Tomato host tissue

growth_protocol: Tomato plants were grown in Metro Mix 360 potting medium (Griffin Greenhouse and Nursery Supplies, Richmond, VA) under greenhouse conditions.

treatment_protocol: Lespedeza dodder (Cuscuta pentagona) seeds were scarified by soaking in concentrated sulfuric acid for one hour, rinsed thoroughly in water, and planted around the base of two-week old tomato seedlings.   Host leaf tissue from parasitized plants, but not including dodder tissue, was harvested. All tissues were immediately frozen in liquid nitrogen and stored at -80ºC.

tag: Lycopersicon esculentum (cv. Ailsa Craig)


title: Tomato_Rep2


title: Tomato_Rep3


title: Dodder_Arabidopsis

description: The objective of this experiment was to assay for movement of mRNA from tomato (host) into dodder (parasitic plant).  Dodder was grown on arabidopsis as one of several control hosts to facilitate identification of endogenous dodder mRNA from those potentially translocated from the .

source_name: Dodder grown on Arabidopsis

growth_protocol: Arabidopsis plants supporting the dodder were grown in Metro Mix 360 potting medium (Griffin Greenhouse and Nursery Supplies, Richmond, VA)in a growth room at 20ºC and 8 hours light.

treatment_protocol: Lespedeza dodder (Cuscuta pentagona) seeds were scarified by soaking in concentrated sulfuric acid for one hour, rinsed thoroughly in water, and planted around the base of four-week old Arabidopsis plants.   Dodder stems at least 4 cm from the host plant were harvested after 9-12 weeks of growth.  To obtain sufficient quantity of dodder tissue, many stems were harvested, representing multiple individuals for each harvest time.  All tissues were immediately frozen in liquid nitrogen and stored at -80ºC.


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-miR-20, 100 nM, 24 hr


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa-miR-215, 25 nM, 24 hr

tag: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-miR-17-5p, 100 nM, 24 hr


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa-miR-192, 25 nM, 24 hr


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-miR-215, 100 nM, 24 hr


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa-miR-15a, 25 nM, 24 hr


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa-miR-16, 25 nM, 24 hr


title: Col-0, leaf tissue, inoculated with Pseudomonas syringae pv. tomato (DC3000hrcC), 1 hr p.i.

type: other

city: Saint Louis

middle: C

last: Carrington

state: MO

address: 975 North Warson Road

institute: Donald Danforth Plant Science Center

description: Amplicons were prepared by 5’ and 3’ adaptor ligation and RT-PCR using small RNA fractions. Samples were sequenced in a multiplexed format in which the 5’ adaptor contained a “barcode” consisting of a four-nucleotide identifier sequence.  Amplicons from three to six samples containing unique barcodes were combined and subjected to picoliter-scale pyrosequencing.

data_processing: Small RNA sequences were parsed from 454 sequencing data files and assigned to specific samples through identification of adaptor boundaries and barcode analysis, respectively.  The value is the number of times each small RNA sequence was identified in each sample.

organism: Arabidopsis thaliana

extract_protocol: Plants were grown under standard greenhouse conditions with supplemental light on a 16-hour photoperiod.  Plants were inoculated with Pseudomonas syringae pv. tomato (DC3000hcrC) for one or three hours before collection of leaf tissue.  Total RNA was extracted using Trizol reagent and small RNA was fractionated on polyacrylamide gels.

tag: Wild-type Col-0, leaf tissue, inoculated with Pseudomonas syringae pv. tomato (DC3000hrcC), samples collect 1 hour post infection


title: Col-0, leaf tissue

source_name: Wild-type Col-0, leaf tissue

extract_protocol: Plants were grown under standard greenhouse conditions with supplemental light on a 16-hour photoperiod.  Lead tissue was collected 4-6 hours into the light cycle.  Total RNA was extracted using Trizol reagent and small RNA was fractionated on polyacrylamide gels.

tag: Wild-type Col-0, leaf tissue


title: rdr6-15, whole seedlings

source_name: rdr6-15 in Col-0 background, whole seedlings

extract_protocol: Seeds were washed with 80% ethanol/0.1% Triton X-100 for ten minutes, then 30% Clorox bleach/0.1% Triton X-100 for 10 minutes, and were rinsed three times with sterile water.  Seeds were imbibed and cold-treated for 72 hours in 0.1% agar prior to plating on MS agar medium overlaid with nylon mesh (Nitex 03-100/47; Sefar America).  Agar plates were oriented vertically in a growth chamber with a 16-hour photoperiod.  3-day-old seedlings were collected 4-6 hours into a light cycle.  Total RNA was extracted using Trizol reagent and small RNA was fractionated on polyacrylamide gels.


title: Composite small RNA sequences

source_name: Various tissues from wild-type and mutant Arabidopsis plants

extract_protocol: Plants were grown under standard greenhouse conditions with supplemental light on a 16-hour photoperiod.  Inflorescence tissue was collected 4-6 hours into the light cycle.  For whole seedlings, seeds were washed with 80% ethanol/0.1% Triton X-100 for ten minutes, then 30% Clorox bleach/0.1% Triton X-100 for 10 minutes, and were rinsed three times with sterile water.  Seeds were imbibed and cold-treated for 72 hours in 0.1% agar prior to plating on MS agar medium overlaid with nylon mesh (Nitex 03-100/47; Sefar America).  Agar plates were oriented vertically in a growth chamber with a 16-hour photoperiod.  3-day-old seedlings were collected 4-6 hours into a light cycle.  Total RNA was extracted using Trizol reagent and small RNA was fractionated on polyacrylamide gels.

tag: The sample contains a composite of small RNAs from 454 sequencing reads with unreadable barcodes in the 5’ adapter and from a 454 sequencing run containing RNA from a composite of Arabidopsis tissue and plant samples. Sequences are from inflorescence (stage 1-12 flowers), leaf, and seedlings tissue of wild-type Col-0 plants, dcl (dcl1-4, dcl2-1, dcl3-1, and dcl4-2) mutants, and rdr (rdr1-1, rdr2-1, and rdr6-15) mutants.


title: Col-0, whole seedlings

source_name: Wild-type Col-0, whole seedlings

tag: Wild-type Col-0, whole seedlings


title: rdr2-1, inflorescence tissue (stage 1-12 flowers)

source_name: rdr2-1 in Col-0 background, inflorescence tissue (stage 1-12 flowers)

extract_protocol: Plants were grown under standard greenhouse conditions with supplemental light on a 16-hour photoperiod.  Inflorescence tissue was collected 4-6 hours into the light cycle.  Total RNA was extracted using Trizol reagent and small RNA was fractionated on polyacrylamide gels.


title: Col-0, leaf tissue, inoculated with Pseudomonas syringae pv. tomato (DC3000hrcC), 3 hr p.i

tag: Wild-type Col-0, leaf tissue, inoculated with Pseudomonas syringae pv. tomato (DC3000hrcC), samples were collected 3 hour post infection


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-miR-16, 100 nM, 24 hr


title: dcl2-1, inflorescence tissue (stage 1-12 flowers)

source_name: dcl2-1 in Col-0 background, inflorescence tissue (stage 1-12 flowers)


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa_miR_106b, 25 nM, 24 hr


title: dcl1-7, inflorescence tissue (stage 1-12 flowers)

source_name: dcl1-7 in Col-0 background, inflorescence tissue (stage 1-12 flowers)


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-miR-15a, 100 nM, 24 hr


title: rdr1-1, inflorescence tissue (stage 1-12 flowers)

source_name: rdr1-1 in Col-0 background, inflorescence tissue (stage 1-12 flowers)


title: rdr6-15, inflorescence tissue (stage 1-12 flowers)

source_name: rdr6-15 in Col-0 background, inflorescence tissue (stage 1-12 flowers)


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa_miR_103, 25 nM, 24 hr

source_name: DLD-1 Dicer -/- #6


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, hsa-miR-192, 100 nM, 24 hr

source_name: HCT116 Dicer -/- #2


title: dcl3-1, inflorescence tissue (stage 1-12 flowers)

source_name: dcl3-1 in Col-0 background, inflorescence tissue (stage 1-12 flowers)


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa_miR_200a, 25 nM, 24 hr


title: HCT116 Dicer -/- #2, mock, 22 hr vs. HCT116 Dicer -/- #2, hsa_miR_103, 22 hr

tag: HCT116 Dicer -/- #2, mock, 22 hr


title: dcl4-2, inflorescence tissue (stage 1-12 flowers)

source_name: dcl4-2 in Col-0 background, inflorescence tissue (stage 1-12 flowers)


title: HCT116 Dicer -/- #2, mock, 22 hr vs. HCT116 Dicer -/- #2, hsa_miR_106b, 22 hr


title: HCT116, mock, 22 hr vs. HCT116, hsa_miR_106b, 100nM, 22 hr

tag: HCT116, mock, 22 hr


title: HCT116, mock, 22 hr vs. HCT116, Luc, 100nM, 22 hr


title: HCT116 Dicer -/- #2, mock, 16 hr vs. HCT116 Dicer -/- #2, KIF11_212 siRNA, 100 nM, 16 hr

tag: HCT116 Dicer -/- #2, mock, 16 hr


title: DLD-1 Dicer -/- #6, mock, 25 nM, 24 hr vs. DLD-1 Dicer -/- #6, hsa_miR_141, 25 nM, 24 hr


title: Hot Spots_9_20_03_WT_1_REV_5ml

type: genomic

city: Chapel Hill

first: Jason

middle: D.

last: Lieb

state: NC

department: Biology

institute: Univeristy of North Carolina at Chapel Hill

description: In eukaryotes, meiotic recombination events are distributed non-randomly in the genome with certain regions having high levels of recombination (hotspots) and others having low levels (coldspots).  Species with similar DNA sequences (for example, chimps and humans) can have strikingly different patterns of hotspots and coldspots and it has been suggested that the sequences that locally regulate recombination activity evolve very rapidly.  Below, using a microarray analysis that allows us to measure the recombination activities of all 6000 yeast genes, we show that elimination of a histone deacetylase encoded by the SIR2 gene significantly changes the recombination activity of 12% of yeast genes, elevating the recombination activity of 5% and reducing the recombination activity of 7%. We also show that many of the genes with suppressed recombination in the sir2 strain are located in large (50-100 kb) sub-telomeric regions and some of the genes with elevated recombination (for example, the ribosomal RNA gene cluster) are targets of Sir2p deacetylation in the wild-type strain. We suggest that changes in the level of chromatin-modifying proteins may be responsible for altered patterns of recombination in closely-related species.

hyb_protocol: Hybridization was performed in 4XSSC, 0.2% SDS and 15 ug of poly A DNA.

scan_protocol: Images were acquired using a GenePix 4000B scanner and Genepix software (Axon Instruments)

data_processing: The normalized median log2 ratios (ch2/ch1) were download from UNC microarray database. Dye swap values were inverted to (ch1/ch2).

molecule: genomic DNA

organism: Saccharomyces cerevisiae

label_protocol: Both the reference genomic DNA and the experimental samples (Spo11p-associated) were used as templates for three rounds of PCR amplification, and the last round utilized Cy3-dUTP as described in the Website for the Cold Spring Harbor Microarray Course (http://www.bio.unc.edu/faculty/lieb/labpages/PDFs/2003_CSH_Array_Course_Protocols_FINAL.pdf).

extract_protocol: The diploids JG169 and MD311 strains are homozygous for a gene encoding an epitope-tagged version of Spo11p (SPO11-ZZ::K.1.URA3) and rad50S.  In rad50S strains, Spo11p is covalently attached to the DNA ends produced by the DSBs that initiate recombination (Keeney et al., 1997).  As described above, strains were sporulated for 24 hours.  The protocol for immunoprecipitation was that described by the Koshland lab (http://www.ciwemb.edu/labs/koshland/Protocols/Yeast/chipmod.html) with the following alterations: 1) formaldehyde was omitted, 2) the SDS lysis buffer contained 1M NaCl, 3) DNA fragments resulting from sonication were about 1.5 kb in size, 4) 5 ml of protein extract derived from 100 ml of sporulating cells were used for the immunoprecipitation, and 5) the Spo11-ZZp-DNA complexes were precipitated with IgG Sepharose beads.  Precipitates were washed two times with TSE-1000 buffer (TSE buffer with 1M NaCl), followed by one wash with TSE-500 buffer.  The resulting precipitate was then washed once with Li/Detergent buffer and once with TE.  All washing steps were done at room temperature.  About half of the cell extract from the 200 ml culture was used as input for the immunoprecipitation.

tag: Precipitated Spo11-ZZ with covalently-attached DNA

source_name: reference DNA

label_protocol: Both the reference genomic DNA and the experimental samples (Spo11p-associated) were used as templates for three rounds of PCR amplification, and the last round utilized Cy5-dUTP as described in the Website for the Cold Spring Harbor Microarray Course (http://www.bio.unc.edu/faculty/lieb/labpages/PDFs/2003_CSH_Array_Course_Protocols_FINAL.pdf).

extract_protocol: Total genomic DNA was isolated from the S. cerevisiae JG169 strain grown in rich growth medium (YPD).  This DNA was sheared by sonication to a size of about 1-2 kb.


title: Hot Spots_9_11_03_WT_3_5ml

data_processing: The normalized median log2 ratios (ch2/ch1) were download from UNC microarray database.

extract_protocol: DNA was extracted from diploid cells of the strain JG169 using the Qiagen Genomic Extraction Kit according to the provided protocol.

tag: Total genomic DNA was isolated from the S. cerevisiae JG169 strain grown in rich growth medium (YPD).  This DNA was sheared by sonication to a size of about 1-2 kb.


title: HCT116/Dicer -/- #2, mock, 24 hr vs. HCT116/Dicer -/- #2, siRNA_141, 100 nM, 24 hr

tag: HCT116/Dicer -/- #2, mock, 24 hr


title: Hot Spots_10_25_03_WT_2_REV_5ml


title: Hot Spots_9_20_03_WT_2_5ml


title: Hot Spots_9_20_03_WT_3_5ml

hyb_protocol: Hybridization was performed in 4XSSC, 0.2% SDS and 15 ?g of poly A DNA.


title: HCT116/Dicer -/- #2, mock, 24 hr vs. HCT116/Dicer -/- #2, siRNA_200b, 100 nM, 24 hr


title: HCT116/Dicer -/- #2, mock, 24 hr vs. HCT116/Dicer -/- #2, siRNA_200a, 100 nM, 24 hr


title: Hot Spots_10_15_03_WT_1_5ml


title: HCT116/Dicer -/- #2, mock, 24 hr vs. HCT116/Dicer -/- #2, Luc, 100 nM, 24 hr


title: Hot Spots_10_03_03_WT_2_5ml


title: Hot Spots_10_03_03_WT_1_5ml


title: Hot Spots_10_03_03_WT_3_5ml


source_name: Inflorescence tissue (stage 1-12 flowers)

tag: Wild-type Col-0, inflorescence tissue (stage 1-12 flowers)


title: HCT116 Dicer -/- #2, mock, 14 hr vs. HCT116 Dicer -/- #2, miR_15a, 100 nM, 14 hr

tag: HCT116 Dicer -/- #2, mock, 14 hr


title: Hot Spots_9_20_03_WT_1


title: HCT116, mock, 16 hr vs. HCT116, KIF11_212 siRNA, 100 nM, 16 hr

tag: HCT116, mock, 16 hr


title: Hot Spots_5_20_04_sir2_312_2

growth_protocol: In most of our experiments, the strain was grown in 200 ml of pre-sporulation medium (SPS; Nag and Petes, 1993) in a two liter flask to a cell concentration measured as OD600= 1.2.  The cells were harvested by centrifugation, washed once with 1% potassium acetate, and then incubated at 220  C. for 24 hours in 200 ml of 1% potassium acetate.


title: HCT116, mock, 24 hr vs. HCT116, siRNA_200a, 100 nM, 24 hr

tag: HCT116, mock, 24 hr


title: Hot Spots_10_25_03_WT_1_5ml


title: Hot Spots_5_20_04_sir2_311_2


title: HCT116, mock, 24 hr vs. HCT116, siRNA_200b, 100 nM, 24 hr


title: HCT116, mock, 24 hr vs. HCT116, siRNA_141, 100 nM, 24 hr


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, miR_106b, 100 nM, 24 hr

tag: HCT116 Dicer -/- #2, mock, 24 hr


title: HCT116 Dicer -/- #2, mock, 14 hr vs. HCT116 Dicer -/- #2, Luc, 100 nM, 14 hr


title: Hot Spots_5_07_04_sir2_312_c

description: Spo11-ZZ Chip


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, miR_15a, 100 nM, 24 hr


title: Hot Spots_9_03_04_sir2_REV_312


title: Hot Spots_5_07_04_sir2_312_a


title: HCT116 Dicer -/- #2, mock, 10 hr vs. HCT116 Dicer -/- #2, miR_106b, 100 nM, 10 hr

tag: HCT116 Dicer -/- #2, mock, 10 hr


title: Hot Spots_5_20_04_sir2_312_1


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, Luc, 100 nM, 24 hr


title: HCT116 Dicer -/- #2, mock, 10 hr vs. HCT116 Dicer -/- #2, miR_15a, 100 nM, 10 hr


title: Hot Spots_5_20_04_sir2_311_c


title: HCT116 Dicer -/- #2, mock, 14 hr vs. HCT116 Dicer -/- #2, miR_16, 100 nM, 14 hr


title: HCT116 Dicer -/- #2, mock, 10 hr vs. HCT116 Dicer -/- #2, miR_16, 100 nM, 10 hr


title: HCT116 Dicer -/- #2, mock, 6 hr vs. HCT116 Dicer -/- #2, miR_16, 100 nM, 6 hr

tag: HCT116 Dicer -/- #2, mock, 6 hr


title: Hot Spots_5_20_04_sir2_308_1


title: HCT116 Dicer -/- #2, mock, 6 hr vs. HCT116 Dicer -/- #2, Luc, 100 nM, 6 hr

tag: HCT116 Dicer -/- #2, Luc, 100 nM, 6 hr


title: Hot Spots_5_07_04_sir2_311_b

hyb_protocol: Hybridization was performed in 4XSSC, 0.2% SDS and 15 g of poly A DNA.


title: Hot Spots_5_20_04_sir2_308_2


title: HCT116 Dicer -/- #2, mock, 24 hr vs. HCT116 Dicer -/- #2, miR_16, 100 nM, 24 hr


title: Hot Spots_9_03_04_sir2_REV_311


title: Hot Spots_9_03_04_sir2_REV_308


title: HCT116 Dicer -/- #2, mock, 6 hr vs. HCT116 Dicer -/- #2, miR_106b, 100 nM, 6 hr


title: HCT116 Dicer -/- #2, mock, 10 hr vs. HCT116 Dicer -/- #2, Luc, 100 nM, 10 hr


title: Calu3-GemR_G+B_2

status: Public on Jan 31 2007

city: San Diego

first: Thomas

last: Hermann

state: CA

address: 10275 Science Center Dr.

department: Molecular Oncology

institute: Ligand Pharmaceuticals

description: gemcitabine resistant Calu3 cells, treated for 10 cycles of 10 days each with gemcitabine plus bexarotene (still largely gemcitabine resistant since cells resensitized during culturing were eliminated from the culture by gemcitabine before harvest)

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Human U133A Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Affymetrix GeneArray scanner 2500.

data_processing: The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 250.

label: biotin

source_name: Calu3 human non-small lung cancer cells, gemcitabine resistant (Ligand)

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 2 ug total RNA (Expression Analysis Technical Manual, 701021 Revision 4, Affymetrix).

growth_protocol: cells were cultured in RPMI 1640 supplemented with 10% FBS and 2 mM glutamine in 95% air, 5% CO2

extract_protocol: Rneasy column (Qiagen) purification of total RNA was performed according to the manufacturer's instructions.

treatment_protocol: gemcitabine resistant Calu3 were treated over 10 consecutive 10-day treatment cycles with 50 nM gemcitabine and 1 micromolar bexarotene for the first 3 days of the cycle followed by 7 days of 1 micromolar bexarotene alone

tag: gemcitabine resistant Calu3 derivative generated from the parental Calu3 line by consecutive treatments with 50 nM gemcitabine over a period of 3 months


title: Calu3-GemR_G+B_4


title: Calu3-GemR_G_3

description: gemcitabine resistant Calu3 cells, treated for 10 cycles of 10 days each with gemcitabine (still gemcitabine resistant)

treatment_protocol: gemcitabine resistant Calu3 were treated over 10 consecutive 10-day treatment cycles with 50 nM gemcitabine for the first 3 days of the cycle followed by 7 days of vehicle


title: Hot Spots_5_20_04_sir2_311_1


title: Calu3-GemR_G_4


title: Calu3-GemR_G+B_3


title: HCT116 Dicer -/- #2, mock, 6 hr vs. HCT116 Dicer -/- #2, miR_15a, 100 nM, 6 hr


title: Calu3-GemR_G_1


title: Calu3-GemR_G+B_1


title: Calu3-GemR_G_2


title: Calu3-GemR_B_3

description: gemcitabine resistant Calu3 cells, resensitized to gemcitabine after treatment for 10 cycles of 10 days each with bexarotene

treatment_protocol: gemcitabine resistant Calu3 were treated over 10 consecutive 10-day treatment cycles with continuous exposure to 1 micromolar bexarotene


title: Muscle of trout fasted for one month cplx_J0-10

city: Rennes

first: Pierre-Yves

last: Rescan

address: Campus de Beaulieu

country: France

institute: INRA

hyb_protocol: A first hybridization was performed using a 33P-labelled oligonucleotide (TAATACGACTCACTATAGGG which is found at the extremity of each PCR product) to monitor the amount of cDNA present in each spot. After stripping (3 hours 68°C, 0.1 SSC, 0.2% SDS), arrays were prehybridized for 4 h at 65°C in hybridization solution (5X Denhardt's, 5 SSC, 0.5% SDS) and hybridized with denatured labeled cDNAs for 48 h at 65°C in hybridization solution.

scan_protocol: arrays were exposed for 65 hours to phosphor-imaging plates before scanning using a FUJI BAS 5000 at 25 µm

data_processing: Signal intensities were quantified using BZScan software. Low oligonucleotide signals (lower than three times the background level) were excluded from the analysis. After this filtering step, signal for each spot was divided by the hybridisation signal obtained with the vector oligonucleotide. After this correction, signals were then normalized by dividing each gene expression value by the median value of the array.

organism: Oncorhynchus mykiss

source_name: A transverse slice of fast-twitch  muscle of  Onchorynchus mykiss (trout) situated beneath the dorsal fin was taken for RNA extraction.

label_protocol: Complex target were prepared from 5 µg of RNA of each sample by simultaneous reverse transcription and labelled for 2 hours at 42°C in the presence of 30 µCi [alpha-33P] dCTP, 120 µM dCTP, 20 mM each dATP, dTTP, dGTP and 400 units Superscript II reverse transcriptase (Invitrogen). RNA was degraded by treatment at 68°C for 30 min with l µl 10% SDS, l µl 0.5M EDTA and 3 µl 3M NaOH, and then equilibrated at room temperature for l5 min. Neutralization was done by adding 10 µl 1M Tris-HCI plus 3 µl 2N HCl.

extract_protocol: Total RNA was purified using TRIzol reagent (Invitrogen, Carlsbad, CA). Chomczynski P. and Sacchi N. (1987): Single step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal. Biochem. 162: 156-159

tag: Tissue

value: Muscle


title: Calu3-GemR_B_4


title: Calu3-GemR_B_2


title: Calu3-GemR_B_1


title: Calu3-GemR_Veh_4

description: gemcitabine resistant Calu3 cells, treated for 10 cycles of 10 days each with vehicle (still gemcitabine resistant)

treatment_protocol: gemcitabine resistant Calu3 were treated with vehicle in analogy to cultures treated with compounds


title: Calu3-Parental_1

description: parental Calu3 cells, no treatment

source_name: Calu3 human non-small lung cancer cells, parental (ATCC)

treatment_protocol: parental Calu3 were maintained in regular growth medium and not subjected to treatment

tag: Morphology

value: epithelial, Organ:lung, Disease:adenocarcinoma (pleural effusion), Gender:male, Ethnicity:caucasian


title: Calu3-GemR_Veh_2


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-5


title: Muscle of trout fasted for one month cplx_J0-7


title: DOA1-mutant, part 1

city: Jena

first: Bernhard

last: Hube

address: Beutenbergstr. 11a

country: Germany

institute: Hans Knoell Institute

description:
for further analysis of the raw data, shown in the data table:
An intensity-dependent data normalization (LOWESS) was performed in GeneSpring 6.0. The different sets of data were compared to each other by one-way analysis of variance (ANOVA) test with a P-value cut-off of < 0.05 for genes that had an intensity in both channels higher than 100. Each gene that passed this test and showed at least 1.4-fold change in two arrays was defined as differentially expressed.

hyb_protocol: Labelled cDNA was hybridized to the C. albicans arrays as described by Sigle et al. (Sigle et al., 2005)

scan_protocol: Hybridized slides were scanned with an Axon 4000B scanner at a 10 μm resolution.

data_processing: Data were extracted by GenePix 4.1 software (Axon).

organism: Candida albicans

source_name: Candida albicans mutant

label_protocol: RNA was reverse transcribed into cDNA and labelled with Cy5 (Sigle et al., 2005)

growth_protocol: Cells from the Δdoa1 mutant and the wild type strain CAF2-1 were precultured in SD at 30 °C and 106 cells ml-1 were inoculated in SD medium at 37 °C for 7 h before RNA was isolated.

extract_protocol: RNA isolated as described previously (Felk et al., 2002)

tag: Δdoa1 mutant

source_name: Candida albicans control strain

label_protocol: RNA was reverse transcribed into cDNA and labelled with Cy3 (Sigle et al., 2005)

tag: control strain CAF2-1 of Candida albicans


title: Calu3-Parental_4

description: parental Calu3 cells, no treatment


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-4


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-9


title: Calu3-GemR_Veh_3


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-9


title: Calu3-Parental_2

description: parental Calu3 cells, no treatment


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-2


title: DOA1-mutant, part 2


title: Calu3-Parental_3

description: parental Calu3 cells, no treatment


title: Calu3-GemR_Veh_1


title: DOA1-mutant, part 3


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-3


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-9


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-4


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-6


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-2


title: Muscle of trout fasted for one month cplx_J0-4


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-10


title: Muscle of trout fasted for one month cplx_J0-5


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-10


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-7


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-8


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-6


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-2


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-8


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-6


title: Muscle of trout fasted for one month cplx_J0-6


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-3


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-2


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-5


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-8


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-5


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-7


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-7


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-3


title: Muscle of trout fasted for one month cplx_J0-8


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-4


title: Muscle of trout fasted for one month cplx_J0-2


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-6


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-5


title: Muscle of trout fasted for one month cplx_J0-3


title: Muscle of trout fasted for one month and then refed for 7 days cplx_J7-9


title: Muscle of trout fasted for one month and then refed for 36 days cplx_J36-7


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-10


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-3


title: Gen-5

city: Little Rock

first: Ying

last: Su

state: AR

address: 1120 Marshall St. South Campus

department: physiology

institute: UAMS

description: Bacterial sequence-derived probes on the arrays served as external controls for hybridization, whereas the housekeeping genes β-actin and GAPDH served as endogenous controls and for monitoring the quality of the RNA target.

hyb_protocol: 10 ug of cRNA was hybridized for 16 hours to an Affymetrix (Santa Clara, CA) rat RAE230A GeneChip (3-4 chips used per diet group), followed by incubations with streptavidin-conjugated phycoerythrin, and then with polyclonal anti-streptavidin antibody coupled to phycoerythrin.

scan_protocol: GeneChips were scanned using an Agilent GeneArray laser scanner. Images were analyzed using Affymetrix MAS 5.0 software.

data_processing: Value column are raw signal intensity generated from MAS 5.0.

organism: Rattus norvegicus

source_name: rat mammary epithelial cells

label_protocol: Pooled RNA (equal amounts of RNA from #4 mammary gland of two animals; 8 ug total) was used for cDNA synthesis using a T7-(deoxythymidine)24 primer and Superscript II (Life Technologies, Inc., Gaithersburg, MD). The resulting cDNA was used with the ENZO BioArray High Yield RNA Transcript labeling kit (ENZO, Farmingdale, NY) to synthesize biotin-labeled cRNA. The cRNA was purified on RNeasy spin columns (QIAGEN) and subjected to chemical fragmentation (size range of 35 to 200 bp). Three replicate cRNA targets were made in parallel starting from each RNA pool.

extract_protocol: TriTRIzol extraction procedure, followed by a cleanup using QIAGEN RNeasy Mini kit.

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet with addition of Genistein (250mg/kg)


title: Muscle of trout fasted for one month and then refed for 4 days cplx_J4-10


title: SPI-4

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet SPI diet


title: GEN-3

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet with addition of Genistein(250mg/kg).


title: CAS-2

label_protocol: Pooled RNA (equal amounts of RNA from #4 mammry gland of two animals; 8 ug total) was used for cDNA synthesis using a T7-(deoxythymidine)24 primer and Superscript II (Life Technologies, Inc., Gaithersburg, MD). The resulting cDNA was used with the ENZO BioArray High Yield RNA Transcript labeling kit (ENZO, Farmingdale, NY) to synthesize biotin-labeled cRNA. The cRNA was purified on RNeasy spin columns (QIAGEN) and subjected to chemical fragmentation (size range of 35 to 200 bp). Three replicate cRNA targets were made in parallel starting from each RNA pool

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet (CAS)


title: CAS-3

hyb_protocol: 10 ug of cRNA was hybridized for 16 hours to an Affymetrix (Santa Clara, CA) rat RAE230 GeneChip (3 chips used per diet group), followed by incubations with streptavidin-conjugated phycoerythrin, and then with polyclonal anti-streptavidin antibody coupled to phycoerythrin.

label_protocol: Pooled RNA (equal amounts of RNA from each of 7 animals; 8 ug total) was used for cDNA synthesis using a T7-(deoxythymidine)24 primer and Superscript II (Life Technologies, Inc., Gaithersburg, MD). The resulting cDNA was used with the ENZO BioArray High Yield RNA Transcript labeling kit (ENZO, Farmingdale, NY) to synthesize biotin-labeled cRNA. The cRNA was purified on RNeasy spin columns (QIAGEN) and subjected to chemical fragmentation (size range of 35 to 200 bp). Three replicate cRNA targets were made in parallel starting from each RNA pool

extract_protocol: TriTRIzol extraction procedure, followed by a cleanup using QIAGEN RNeasy Mini kit

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet (CAS ) or SPI diet.


title: Muscle of trout fasted for one month and then refed for 11 days cplx_J11-4


title: SPI-5

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of SPI diet.


title: genomic L1-24

status: Public on Jan 30 2007

city: Burjassot

first: Jose

middle: E.

last: Perez-Ortin

state: Valencia

address: Dr. Moliner 50

department: Bioquimica y Biologia Molecular

country: Spain

institute: Universitat de Valencia

description: L1-24 filter hybridization with labeled yeast genomic DNA. DNA labelling by random priming procedure and 33P-dCTP.

data_processing: raw image quantitation background substracted

source_name: Yeast genomic DNA

growth_protocol: Exponential growth in YPD medium

tag: Total genomic DNA from FY1679 yeast strain


title: unstimulated embryonic fibroblasts KO, biological rep2

city: Cambridge

first: Benjamin

middle: Robert

last: tenOever

address: 7 Divinity Avenue

department: MCB

institute: Harvard University

description: Gene expression data from unstimulated EF cells.

hyb_protocol: Following fragmentation, 15 microg of cRNA were hybridized for 16 hr at 45C on GeneChip Hybridization Oven 640.

scan_protocol: GeneChips were scanned using the GeneChip Array Scanner 7G

data_processing: the data were analyzed with GeneChip Operating Software (GCOS) ver 1.3.

organism: Mus musculus

source_name: embryonic fibroblast basal transcription

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 5 microg total RNA

growth_protocol: Cells were grown in DMEM (GibcoBRL) supplemented with 10% Fetal Bovine Serum

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions.

treatment_protocol: Fibroblasts were left untreated and RNA was extracted using Trizol solution (GibcoBRL).

tag: Genotype

value: ikbke -/- BL/6

tag: Primary cell extract


title: CAS-1

hyb_protocol: 10 ug of cRNA was hybridized for 16 hours to an Affymetrix (Santa Clara, CA) rat RAE230A GeneChip (3-4chips used per diet group), followed by incubations with streptavidin-conjugated phycoerythrin, and then with polyclonal anti-streptavidin antibody coupled to phycoerythrin.

label_protocol: Pooled RNA (equal amounts of RNA from #4 mammary gland from 2 animals; 8 ug total) was used for cDNA synthesis using a T7-(deoxythymidine)24 primer and Superscript II (Life Technologies, Inc., Gaithersburg, MD). The resulting cDNA was used with the ENZO BioArray High Yield RNA Transcript labeling kit (ENZO, Farmingdale, NY) to synthesize biotin-labeled cRNA. The cRNA was purified on RNeasy spin columns (QIAGEN) and subjected to chemical fragmentation (size range of 35 to 200 bp). Three replicate cRNA targets were made in parallel starting from each RNA pool

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet (CAS )


title: SPI-2

hyb_protocol: 10 ug of cRNA was hybridized for 16 hours to an Affymetrix (Santa Clara, CA) rat RAE230 GeneChip (3-4 chips used per diet group), followed by incubations with streptavidin-conjugated phycoerythrin, and then with polyclonal anti-streptavidin antibody coupled to phycoerythrin.


title: genomic L2-1

description: L2-1 filter hybridization with labeled yeast genomic DNA. DNA labelling by random priming procedure and 33P-dCTP.


title: genomic L2-24

description: L2-24 filter hybridization with labeled yeast genomic DNA. DNA labelling by random priming procedure and 33P-dCTP.


title: GEN-1

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based diet with addition of Genistein (250mg/kg).


title: H37Rv-Mbovis_rep2

city: Valdivia

first: Germán

last: Rehren

state: Valdivia

address: Campus Isla Teja

department: Biochemistry

country: Chile

institute: Universidad Austral de Chile

description: Synthesis and hybridization of labelled samples was performed as described in previously published protocols (Walters et al. Mol Microbiol (2006) 60(2):312-330)

hyb_protocol: Total purified cDNA was added to the arrays in a hybridization solution containing a final concentration of 0.5 ug/ul tRNA, 2.0x SSC, 0.25% formamide and 0.1% SDS.  For each array, cDNA prepared from the M. tuberculosis RNA was mixed with cDNA from M. bovis. The slides were covered by a flat 22 x 22 mm coverslip and hybridized in sealed hybridization chambers for sixteen hours at 50°C in a water bath.

scan_protocol: Prepared slides were immediately scanned using an Axon 4000A Scanner (Axon Instruments). Fluorescence intensities of Cy3 and Cy5 dies at each spot were quantified using the GenePix Pro 4.0 software.

data_processing: Intensity values obtained from spots on the chip were background subtracted and negative corrected intensities were set to +1. Corrected data were then normalized using the Lowess implementation included in GeneSpring 7.2 software (Silicon Genetics). The values are expressed as Ch2/Ch1 ratio determined from the normalized fluorescence intensity.

organism: Mycobacterium tuberculosis

source_name: M. tuberculosis culture

growth_protocol: Mid log phase culture was shortly centrifuged and the pellet quickly frozen with dry ice

extract_protocol: Manganelli et al. Mol Microbiol (2001) 41(2):423-437

treatment_protocol: Culture grew in Middlebrook 7H9 media supplemented with ADS, glycerol and Tween 80.

tag: Strain H37Rv

organism: Mycobacterium tuberculosis variant bovis

source_name: M. bovis culture

treatment_protocol: Culture grew in Middlebrook 7H9 media supplemented with ADS, pyruvate and Tween 80.

tag: Strain Ravenel (TMC 401)


title: unstimulated embryonic fibroblasts KO, biological rep3


title: unstimulated embryonic fibroblasts WT, biological rep2

value: wild type BL/6


title: H37Rv-Mbovis_rep3

label_protocol: Walters et al. Mol Microbiol (2006) 60(2):312-330


title: H37Rv-Mbovis_rep1

treatment_protocol: Culture grew in Middlebrook 7H9 media supplemented with ADS, glycerol ans Tween 80.


title: H37Rv-Mbovis_rep4


title: SPI-3

hyb_protocol: 10 ug of cRNA was hybridized for 16 hours to an Affymetrix (Santa Clara, CA) rat RAE230 GeneChip ( 3-4 chips used per diet group), followed by incubations with streptavidin-conjugated phycoerythrin, and then with polyclonal anti-streptavidin antibody coupled to phycoerythrin.

tag: Female Sprague-Dawley rat, postnatal day 50, life-time ingestion of casein based SPI diet


title: unstimulated embryonic fibroblasts WT, biological rep3


title: Swirm2_1_WT

city: Bethesda

first: Shiv

last: Grewal

state: MD

address: NCI bldg 37 Rm 6068 9000 Rockville Pike

department: LBMB

institute: NCI

description: Schizosaccharomyces pombe cells, Swirm2 ChIP(chromatin immunoprecipitation)-on-chip, replicate 1

hyb_protocol: 500 ng of Cy3 labeled WCE DNA was mixed with 500 ng of Cy5 labeled ChIP DNA plus 50 ul Agilent 10X control targets and 250 ul of 2X Agilent Hybridization buffer

scan_protocol: Slides were scanned using Agilent scanner and image intensity data were extracted and analyzed with Agilent Feature Extraction software version 7.5

data_processing: Log ratios of ChIP over WCE were obtained by dividing normalized median Cy5 value over its corrresponding Cy3 value

organism: Schizosaccharomyces pombe

source_name: Schizosaccharomyces pombe whole cell extract (WCE) genomic DNA

label_protocol: WCE DNA after amplified by random priming PCR incorporating amino-allyl dUTP was covalently coupled with Cy3 mono NHS ester in 100 mM sodium bicarbonate for 1 hr at room temperature. Labeled DNA was recovered by Qiagen QIAquick PCR column

growth_protocol: Asynchronous growing fission yeast cells at 30 C and cross-linked at 18C with paraformaldehyde for 30 minutes

extract_protocol: Chromatin was extracted from cross-linked cells following lysis with bead beater and sonication.  After reverse cross-linked and digested with Rnase A and proteinase K, DNA was recovered by phenol/chloroform extraction and ethanol precipitation

treatment_protocol: Schizosaccharomyces pombe cells crosslinked with paraformadehyde

source_name: Schizosaccharomyces pombe chromatin immunoprecipitated DNA

label_protocol: ChIP DNA after amplified by random priming PCR incorporating amino-allyl dUTP was covalently coupled with Cy5 mono  NHS ester in 100 mM sodium bicarbonate for 1 hr at room temperature. Labeled DNA was recovered by Qiagen QIAquick PCR column

growth_protocol: Asynchronous growing fission yeast cells at 30 C and cross-linked with paraformaldehyde at 18C for 30 minutes

tag: Chromatin DNA isolated from paraformaldehyde cross-linked wild type Schizosaccharomyces pombe cells and enriched for FLAG-Swirm2 chromatin with anti-FLAG (M2) antibody.


title: genomic L1-2

description: L1-2 filter hybridization with labeled yeast genomic DNA. DNA labelling by random priming procedure and 33P-dCTP.


title: Swirm1_2_WT

description: Schizosaccharomyces pombe cells, Swirm1 ChIP(chromatin immunoprecipitation)-on-chip, replicate 2

tag: Chromatin DNA isolated from paraformaldehyde cross-linked wild type Schizosaccharomyces pombe cells and enriched for FLAG-Swirm1 chromatin with anti-FLAG (M2) antibody.


title: Swirm2_2_WT

description: Schizosaccharomyces pombe cells, Swirm2 ChIP(chromatin immunoprecipitation)-on-chip, replicate 2

tag: Chromatin DNA isolated from paraformaldehyde cross-linked wild type Schizosaccharomyces pombe cells


title: genomic L1-1

description: L1-1 filter hybridization with labeled yeast genomic DNA. DNA labelling by random priming procedure and 33P-dCTP.


title: Retinoblatoma M22233

status: Public on Jan 29 2007

city: Essen

first: Ludger

last: Klein-Hitpass

address: Virchowstr. 173

department: Universitaetsklinikum

institute: Institut fuer Zellbiologie

description: Aim: to compare the transcriptomes of retinoblastoma with LOH on 16q to tumors without alterations on 16q.

hyb_protocol: Following fragmentation, 10 microg of cRNA were hybridized for 16 hr at 45C on HG-U133A Arrays. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G2500A.

data_processing: The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 1000. Differentially expressed target genes were identified using Significance Analysis of Microarrays (SAM).

source_name: retinoblastoma

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 2.5 microg total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

growth_protocol: no in vitro culture

extract_protocol: Isolation of total RNA was performed by a modified Qiagen RNeasy Mini Protocol for the isolation of totel RNA from animal tissues (QIAgen, Hilden, Germany). All the buffers and columns that are contained in this kit were used, except when specified. A 30 mg tumour sample is placed in 800 µl Buffer RLT (ß-ME 10µl/ml, Sigma-Aldrich, Taufkirchen, Germany) in a Lysing Matrix D column (Q.BIOgene, Heidelberg, Germany) and grinded in the Lysing Matrix for 45 s at 6.5 (Fast Prep, FP 120, Bio 101 Systems, Q.BIOgene). Dry ice is used to cool down the sample after grinding. At room temperature, the samples are shaken, vortexed and spun down for 5 min at 13,000 rpm at 4°C. The supernatant is taken out and placed in a new 1.5 ml tube, and spun down again in the same conditions as before. Leaving the protein pellet intact, the supernatant is placed in a 1.5 ml tube to which 750 µl Ethanol 70% is added, and then vortexed. The 1.5 ml solution obtained in this step is then placed on an RNeasy mini spin column and centrifuged for 15 s at 10,000 rpm at 4°C (in 2 steps). The flow-through is discarded. Digestion with DNaseI (QIAgen) is used to avoid the presence of DNA in the final extract. For this, 350 µl Buffer RW1 is added to the spin column and centrifuged for 15 s at 10,000 rpm. The flow-through is discarded. A mix of 10 µl DNaseI stock solution and 70 µl buffer RDD is thoroughly pipetted on the membrane of the column and incubated for 15 min at room temperature. After adding 350 µl buffer RW1 the column is centrifuged for 15 s at 10,000 rpm. The column is then placed on a new collection tube, 500 µl buffer RPE (with Ethanol) is added and spun down for 2 min at 13,000 rpm. The column placed on a new collection tube and the RNA is eluted with 50 µl RNase free water by centrifugation for 1 min at 10,000 rpm. The elution step is repeated one time..

treatment_protocol: no treatment

tag: Patient

value: sporadic unilateral Rb, male, age at dx in days:897, primary enucleation, original RNA ID M22234

tag: LOH 16q

value: no


title: Retinoblatoma M23978

value: sporadic unilateral Rb, female, age at dx in days:326, primary enucleation, original RNA ID M23979


title: Retinoblatoma M22641

value: sporadic unilateral Rb, male, age at dx in days:1051, primary enucleation, original RNA ID M22642

value: yes


title: Retinoblatoma M22590

value: sporadic unilateral Rb, female, age at dx in days:603, primary enucleation, original RNA ID M22591


title: Swirm1_1_WT

description: Schizosaccharomyces pombe cells, Swirm1 ChIP(chromatin immunoprecipitation)-on-chip, replicate 1


title: Retinoblatoma M23869

value: sporadic unilateral Rb, male, age at dx in days:448, primary enucleation, original RNA ID M23870


title: Retinoblatoma M22860

value: sporadic unilateral Rb, female, age at dx in days:1725, primary enucleation, original RNA ID M22861


title: Retinoblatoma M23449

value: sporadic unilateral Rb, female, age at dx in days:579, primary enucleation, original RNA ID M23450


title: Retinoblatoma M19484

value: sporadic unilateral Rb, female, age at dx in days:985, primary enucleation, original RNA ID M19485


title: Retinoblatoma M20517

value: sporadic bilateral Rb, male, age at dx in days:240, primary enucleation, original RNA ID M20519


title: Retinoblatoma M23209

value: sporadic bilateral Rb, male, age at dx in days:522, primary enucleation, original RNA ID M23210


title: Retinoblatoma M22067

value: sporadic unilateral Rb, male, age at dx in days:120, primary enucleation, original RNA ID M22069


title: Retinoblatoma M23818

value: sporadic unilateral Rb, female, age at dx in days:942, primary enucleation, original RNA ID M23818


title: Tumor_26

status: Public on Jan 26 2007

city: Buenos Aires

first: Michele

last: Bianchini

address: Cramer 1180

department: Centro de Investigaciones Oncologicas

country: Argentina

institute: Instituto Alexander Fleming

description: high quality RNA with minimal degradation and clear 28S/18S ribosomal bands

data_processing: MIDAS (Microarray Data Analysis System)

source_name: Tumor Left Colon Tissue

tag: Sex M

tag: Age 82

tag: TNM T3N0M0

tag: Dukes B

tag: AJCC 2

source_name: Normal Colon Tissue

tag: Pool of 13 colon mucosa:

tag: 4 right normal colon tissue

tag: 4 rectum normal tissue

tag: 5 left normal colon tissue


title: top1, A2S26a.gpr

city: Acton

first: Anthony

last: Millgate

state: ACT

address: PO Box 1600

country: Australia

institute: CSIRO

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 2, Technical Replicate: 1, Dye Swapped: false
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism

organism: Papaver somniferum

source_name: RNA from whole leaf/stem of D233 mutant at running up stage

source_name: RNA from whole leaf/stem of Wild Type CO52 at running up stage


title: Tumor_17

source_name: Tumor Right Colon Tissue

tag: Sex F

tag: Age 81

tag: TNM T3N1M0

tag: Dukes C

tag: AJCC 3


title: Tumor_24

tag: Age 35


title: kidney sea bream-2

city: Heraklion

first: Elena

middle: Athina

last: Sarropoulou

department: Marine Biology and Genetics

country: Greece

institute: HCMR

description: Hybridization of one kidney of cortisol treated fish vs. one kidney of control fish.

organism: Sparus aurata

source_name: cortisol treated fish (Cy3)

source_name: control fish (Cy5)


title: top1, A2S23.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 2, Technical Replicate: 2, Dye Swapped: true
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: top1, A2S24a.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 3, Technical Replicate: 1, Dye Swapped: true
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: neurula (Cy5) vs eye pigmentation (Cy3)

description: eye pigmentation (168hpf) (Cy3) vs neurula (24hpf) (Cy5) of sea bream

source_name: neurula (Cy5)

source_name: eye pigmentation (Cy3)


title: eye development (Cy5) vs mouth opening stage (Cy3)

description: eye development and pectoral fin budding stage (96hpf) (Cy5) vs mouth opening stage (144 hpf) (Cy3)

source_name: eye development (Cy5)

source_name: mouth opening stage (Cy3)


title: top1, A2S25b.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 1, Technical Replicate: 4, Dye Swapped: false
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: top1, A2S27.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 3, Technical Replicate: 2, Dye Swapped: false
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: top1, A2S22a.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 1, Technical Replicate: 3, Dye Swapped: true
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: neurula  (Cy3) vs  eye development (Cy5)

description: eye development and pectoral fin budding stage (96hpf) (Cy5) vs neurula (24hpf) (Cy3)

source_name: neurula  (Cy3)


title: neurula (Cy3) vs eye pigmentation (Cy5)

description: eye pigmentation (168hpf) (Cy5) vs neurula (24hpf) (Cy3) of sea bream

source_name: neurula (Cy3)

source_name: eye pigmentation (Cy5)


title: top1, A2S15a.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 3, Technical Replicate: 4, Dye Swapped: false
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: mouth opening stage  (Cy5) vs eye pigmentation (Cy3)

description: eye pigmentation (168hpf)(Cy5) stage vs mouth opening stage (144 hpf) (Cy3)

source_name: mouth opening stage  (Cy5)


title: neurula  (Cy5) vs  hatching (Cy3)

description: Hatching stage  (48hpf) (Cy3) vs neurula (24hpf) (Cy5) of sea bream

source_name: neurula  (Cy5)

source_name: hatching (Cy3)


title: neurula (Cy3) vs Hatching stage (Cy5)

source_name: Hatching stage (Cy5)


title: mouth opening (Cy3) vs eye pigmentation (Cy5)

description:
eye pigmentation (Cy5) stage vs mouth opening stage (144 hpf) (Cy3)
of sea bream

source_name: mouth opening (Cy3)


title: top1, A2S14b.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 2, Technical Replicate: 4, Dye Swapped: false
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism

source_name: RNA from whole leaf/stem of D233 mutant at running up stage


title: top1, A2S13.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 1, Technical Replicate: 1, Dye Swapped: false
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism

source_name: RNA from whole leaf/stem of Wild Type CO52 at running up stage


title: kidney sea bream-1

source_name: cortisol treated fish (Cy5)

source_name: control fish (Cy3)


title: top1, A2S12a.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 3, Technical Replicate: 3, Dye Swapped: true
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: eye development (Cy5) vs eye pigmentation (Cy3)

description:
eye pigmentation (168hpf) (Cy3)
eye development and  pectoral fin budding 96hpf (Cy5)


title: eye development (Cy3) vs mouth opening stage (Cy5)

description: mouth opening stage (Cy5)  (144hpf) vs eye development  and pectoral fin budding stage (96 hpf)(Cy3)

source_name: eye development (Cy3)

source_name: mouth opening stage (Cy5)


title: top1, A2S8a.gpr

description:
Thebaine and oripavine accumulating mutant (Norman) verses wildtype.
leaf RNA extracted from preflowering material of the top1 mutant of Papaver somniferum
leaf RNA extracted from preflowering Papaver somniferum cv C052.  Plants were grown in the glasshouse All RNA isolation was accomplished with the NTES method Higgins et al., Vol 260: 166-168 1976. RNA (100 µg) was reverse-transcribed into cDNA and flurescently labeled for microarray hybridization as per the methods in Schenk, P.M. et al. "Coordinated Plant Defense Responses in Arabidopsis Revealed by Microarray Analysis." Proceedings of the National Academy of Sciences of the United States of America 97, 11655-11660 (2000).
Hybridisation protocols are as per Protocols for Corning Gaps II coated slides Refer: http://www.corning.com/Lifesciences/technical_information/techDocs/gaps_ii_manual_protocol_5_02_cls_gaps_005.pdf
Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner (Axon Instruments, Union, CA) and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, the data was normalised using the spatial-normalisation protocol from the tRMA package Wilson, D.L , Buckley, M.J., Helliwell, C.A., and Wilson, I.W. "New Normalization Methods for cDNA Microarray Data." Bioinformatics 19, 1325-1332 (2003).   Using the relevant function in tRMA the spatially normalised back to ratio differentially-expressed genes were extracted from the complete list of 17,000 clones' median values.  Gaussian distribution outliers were selected.  Log Red dye/log Green dye [log(R/G)] data were compared using GenePix Pro 3 software (after spatial normalisation).  A ratio threshold cut-off of 2 was used.
Biological Replicate: 2, Technical Replicate: 3, Dye Swapped: true
Normalization Method: Spatial Normalization. Ref: Wilson DL, Buckley MJ, Helliwell CA, Wilson IW (2003) New normalization methods for cDNA microarray data. Bioinformatics 19, 1325-32.
Keywords = Opium Poppy, Secondary Metabolism


title: Mammary gland WT NT rep2

city: Houston

first: Kathryn

last: Schwertfeger

state: TX

address: 1 Baylor Plaza

institute: Baylor College of Medicine

description: WT mammary gland not treated

data_processing: MAS 5.0

label: Affymetrix

source_name: WT mammary gland not treated

tag: Strain

value: FVB

tag: Genotype

tag: Gender

value: Female

tag: Age

value: 6 weeks

tag: Tissue

value: Mammary gland

tag: Treament

value: not treated


title: Mammary gland WT NT rep1

description: WT mammary gland NT

source_name: WT mammary gland NT


title: hatching stage (Cy5) vs eye development stage (Cy3)

description:
stage of eye development   and pectoral fin budding (Cy3) (96hpf) of sea bream.
Stage of hatching (48hpf)(Cy5)

source_name: hatching stage (Cy5)

source_name: eye development stage (Cy3)


title: HUVEC transcriptional response to cigarette smoke extract after 24 hours_10441

city: Innsbruck

first: Blair

last: Henderson

address: Fritz-Pregl Strasse 3

department: Biocenter

country: Austria

institute: Innsbruck Medical University

description: No additional information.

hyb_protocol: Hybridisation of the labelled RNA to the microarray was carried out for 16 hours at 42°C in a moistened CMT-Hybridisation chamber (Corning, Acton, MA).  Non-specifically bound RNA was removed by washing for 5 min each with buffers 1-3 at 30°C respectively (wash buffer 1: 2x SSC, 0.1% SDS; wash buffer 2: 1x SSC, 0.1% SDS; wash buffer 3: 0.5x SSC)

scan_protocol: Microarrays were scanned using a GenePix 4000B array scanner and GenePix Pro software (Axon, Union City, CA).

data_processing: Initial analysis of the arrays was performed using GenePix Pro software, before normalisation using LimmaGUI software.  The normalisation process used was a print-tip group Loess method based on median values with background correction.  The LimmaGUI algorithm normalises the features within each block on the array, based on the assumption that the global expression within each block is unchanged.  Further data analysis was performed using the Excel programme from Microsoft.

label: Cy 5

source_name: human umbilical vein endothelial cells, control

label_protocol:
A single round of T7 RNA polymerase directed linear amplification with direct incorporation of Cy5 or Cy3 after cDNA synthesis was performed on
Total RNA (2µg) using the MessageAmp aRNA Kit (Ambion, Austin, Texas), before in vitro transcription using T7 RNA polymerase according to the manufacturer’s directions.  After determination of the RNA concentration, up to 40 µg was fragmented in Affymetrix fragmentation buffer by heating at 94°C for 15 minutes, and subsequently stored on ice.  Samples were post-labelled with Cy5  dye (CyScribe Direct mRNA labelling kit, Amersham Pharmacia Biotech, Uppsala, Sweden), according to the manufacturer’s instructions.  For each of the control and exposed (MF) samples, 2µg of the amplified RNA was labelled.  Labelled RNA was ethanol precipitated before being resuspended in hybridisation buffer (50% formamide, 5x SSC, 0.1% SDS).

growth_protocol: The cells were routinely passaged in 0.2% gelatin coated (2% solution, Sigma, Steinheim, Germany) polysterene culture flasks (Becton Dickinson, Meylan Cedex, France) in Endothelial Cell Basal Medium (CC-3121, BioWhittaker, Inc., Walkersville, MD, USA) supplemented with EGM SingleQuots Supplements and Growth Factors (CC-4133, BioWhittaker, Inc.) in a humidified atmosphere containing 5% CO2.

extract_protocol: cells were collected using a plastic cell scraper, and washed twice in ice cold phosphate buffered saline (PBS, pH 7.2) before total RNA was isolated using TriReagent according to the manufacturer’s instructions (Sigma, Vienna, Austria).  To ensure total removal of contaminants, RNA was further purified using RNeasy spin columns and the “RNA clean-up” protocol (Qiagen, Hilden, Germany).

treatment_protocol: Untreated control cells

tag: human umbilical vein endothelial primary cells, freshly isolated and cultivated for 5 passages.

source_name: human umbilical vein endothelial cells, treated with 8% cigarette smoke extract for 24 hours

label_protocol:
A single round of T7 RNA polymerase directed linear amplification with direct incorporation of Cy5 or Cy3 after cDNA synthesis was performed on
Total RNA (2µg) using the MessageAmp aRNA Kit (Ambion, Austin, Texas), before in vitro transcription using T7 RNA polymerase according to the manufacturer’s directions.  After determination of the RNA concentration, up to 40 µg was fragmented in Affymetrix fragmentation buffer by heating at 94°C for 15 minutes, and subsequently stored on ice.  Samples were post-labelled with Cy3  dye (CyScribe Direct mRNA labelling kit, Amersham Pharmacia Biotech, Uppsala, Sweden), according to the manufacturer’s instructions.  For each of the control and exposed (MF) samples, 2µg of the amplified RNA was labelled.  Labelled RNA was ethanol precipitated before being resuspended in hybridisation buffer (50% formamide, 5x SSC, 0.1% SDS).

treatment_protocol:
Cells were treated with an aqueous cigarette smoke extract (8% v/v) made by bubbling cigarette smoke through cell culture medium (1 cigarette in 4 mL medium).  The extract was sterile filtered before being added to cells.
The extract has been described in the following publications:
Bernhard, D., Pfister, G., Huck, C. W., Kind, M., Salvenmoser, W., Bonn, G. K. & Wick, G. (2003) Faseb J 17, 2302-4. Epub 2003 Oct 02.
Bernhard, D., Huck, C. W., Jakschitz, T., Pfister, G., Henderson, B., Bonn, G. K. & Wick, G. (2004) J Pharmacol Toxicol Methods 50, 45-51.


title: Mammary gland R1 24 hr S rep1

description: iFGFR1 mammary gland 24 hr solvent

source_name: iFGFR1 mammary gland 24 hr solvent

value: 24 hrs Solvent


title: HUVEC transcriptional response to cigarette smoke extract after 24 hours_10432

description: No additional information.

label: Cy 3


title: hatching stage  (Cy3) vs mouth opening stage (Cy5)

description: mouth opening stage(144hpf)(Cy5)  and hatching stage (48hpf) (Cy3) of sea bream.

source_name: hatching stage  (Cy3)


title: eye development (Cy3) vs eye pigmentation (Cy5)

description: eye development and pectoral budding (96hpf)(Cy3)  and eye pigmentation (168hpf) (Cy5) of sea bream.

source_name: eye pigmentation (Cy5)


title: stage neurula (Cy5) vs eye development (cy3)

description:
stage neurula (24h pf) (Cy5) vs
stage eye development and budding of pectoral fin (96hpf) (Cy3) of sea bream.

source_name: neurula stage of sea bream

source_name: eye development stage of sea bream


title: HUVEC transcriptional response to cigarette smoke extract after 3 hours_10426

description: No additional information.

label_protocol:
A single round of T7 RNA polymerase directed linear amplification with direct incorporation of Cy5 or Cy3 after cDNA synthesis was performed on
Total RNA (2µg) using the MessageAmp aRNA Kit (Ambion, Austin, Texas), before in vitro transcription using T7 RNA polymerase according to the manufacturer’s directions.  After determination of the RNA concentration, up to 40 µg was fragmented in Affymetrix fragmentation buffer by heating at 94°C for 15 minutes, and subsequently stored on ice.  Samples were post-labelled with Cy3  dye, respectively, (CyScribe Direct mRNA labelling kit, Amersham Pharmacia Biotech, Uppsala, Sweden), according to the manufacturer’s instructions.  For each of the control and exposed (MF) samples, 2µg of the amplified RNA was labelled.  Labelled RNA was ethanol precipitated before being resuspended in hybridisation buffer (50% formamide, 5x SSC, 0.1% SDS).

source_name: human umbilical vein endothelial cells, treated with 8% cigarette smoke extract for 3 hours

label_protocol:
A single round of T7 RNA polymerase directed linear amplification with direct incorporation of Cy5 or Cy3 after cDNA synthesis was performed on
Total RNA (2µg) using the MessageAmp aRNA Kit (Ambion, Austin, Texas), before in vitro transcription using T7 RNA polymerase according to the manufacturer’s directions.  After determination of the RNA concentration, up to 40 µg was fragmented in Affymetrix fragmentation buffer by heating at 94°C for 15 minutes, and subsequently stored on ice.  Samples were post-labelled with Cy5  dye, respectively, (CyScribe Direct mRNA labelling kit, Amersham Pharmacia Biotech, Uppsala, Sweden), according to the manufacturer’s instructions.  For each of the control and exposed (MF) samples, 2µg of the amplified RNA was labelled.  Labelled RNA was ethanol precipitated before being resuspended in hybridisation buffer (50% formamide, 5x SSC, 0.1% SDS).


title: S._cerevisiae_RM11-1a/BY4741_OD600=1_8-replicates-dye-swapping

city: Taipei

first: Huang-Mo

last: Sung

address: 128 Sec.2 Academia Rd.

department: Genomics Research Center

country: Taiwan

institute: Academia Sinica, Taiwan

description:
We could like to know the expression divergency of S. cerevisiae strains RM11-1a and BY4741.
Yeast strains were grown in YPAD and harvested at the mid-log phrase.  Overnight yeast cultures were used to prepare the starting cultures with OD600=0.1 and were grown in YPAD media at 30oC with 250 rpm shaking.  The yeast cells were harvested at the OD600=1.0 and the total RNAs were extracted by the hot acid phenol-chloroform method.

hyb_protocol: The hybridization of cDNA samples to the probes were done with the MAUI hybridization System (Biomicro) at 42oC for 16 hours. Pre-hybridization 1. Incubate arrays in pre-hybridization buffer, 42℃, 30-60 min Final conc.	           Volume 5X SSC	            12.5 ml 0.1% SDS	             0.5 ml 0.1 mg/ml BSA	0.5 ml ddH2O	                        36.5 ml Total	                           50 ml 2. Wash array by immersing in water 3. Dry by centrifugation ~ 5 min Hybridization 1. Prepare probe in fresh hybridization solution in 0.5 ml tube Final conc.	  Volume Labeled cDNA	  42 μl 5X SSC	  15 μl 0.1% SDS	   3 μl Total	              60 μl 2. denature the probe at 98℃ for 3 min, pop-spin, and cool to R/T 3. Place in hybridization chamber 4. Hybridize in 42℃ hybridizer (MAUI hybridization system) for 16 hr Post-hybridization (Washing) (Pre-warm the washing buffers in 30℃) 1. Dissemble the hybridization chamber 2. Immerse in 30℃-prewarmed (2X SSC+0.1% SDS), peel off the coverslip 3. Wash in 30℃-prewarmed (2X SSC+0.1% SDS) in R/T, 5 min 4. Wash in 30℃-prewarmed (0.1X SSC, 0.1% SDS) in R/T, 5 min 5. Wash in 30℃-prewarmed 0.1X SSC, in R/T, 5 min 6. Dry by centrifuge, scan

scan_protocol: The microarray was scanned with GenePix 4000B microarray scanner (Axon Instruments) with the GenePix 5 software package.

data_processing: See linked supplementary file: GSM157844_Data_processing.pdf

molecule: polyA RNA

label_protocol:
amino-allyl dye coupling procedure
1. 1 μl 0.5 M NaHCO3, pH 9.0 + 9 μl Aminoallyl-cDNA solution (final 50 mM)
2. Transfer 1. to microfuge tube containing dried down dye**
3. Pipet up and down
4. R/T 1 hr in the dark
5. Mix and spin every 15 min

growth_protocol: S. cerevisiae RM11-1a strain was grown to OD600=1 with YPAD media.

extract_protocol:
The yeast cells were harvested at the OD600=1.0 and the total RNAs were extracted by the hot acid phenol-chloroform method.
The mRNA was isolated by using the QIAGEN Oligotex mRNA purification kit (Qiagen) following the manufacturer’s instructions.

tag: S. cerevisiae RM11-1a strain

growth_protocol: S. cerevisiae BY4741 strain was grown to OD600=1 with YPAD media.

tag: S. cerevisiae BY4741 strain


title: HUVEC transcriptional response to cigarette smoke extract after 3 hours_10440

description: No additional information.


title: HUVEC transcriptional response to cigarette smoke extract after 7 hours_10437

description: No additional information.

source_name: human umbilical vein endothelial cells, treated with 8% cigarette smoke extract for 7 hours


title: Medicago truncatula (DZA315.16)  12 hours after E. pisi infection, rep3

status: Public on Jan 25 2007

city: St Paul

middle: A

last: VandenBosch

state: MN

address: Room 250 BioSci 6022 1445 Gortner Ave

department: Plant Biology

institute: University of Minnesota

description: Medicago truncatula  12 hours after no pathogen treatment (control)3

hyb_protocol: The hybridization and washing procedures of 3DNA Array 50 Expression Array Detection Kit were followed. Briefly, the hybridization mix consisted of 2.5 ó L of cDNAs from inoculated and uninoculated roots, 2.5 ó L of Cy3 and Cy5 dyes, 2 ó L of LNA dT blocker and 9 ó L of 2X SDS hybridization buffer. Hybridization was carried out at 62.0oC for 20- 23 h. Hybridized slides were washed in 2X SSC, 0.2% SDS for 10 min at 55oC, 2X SSC for 10 min at room temperature, and 0.2X SSC for 10 min at room temperatures with gentle agitation. Each hybridization was repeated a total of 6 times to sample the technical variability, with 3 repeats of each dye combination to control for dye effects.

scan_protocol: Microarray slides were scanned using an GenePix4000B (Axon) two-laser scanner and image analysis was performed using GenePix 5.0 (Axon) software

data_processing: Background-subtracted mean intensities for both tissues were log2 transformed and normalized using lowess and ANOVA

organism: Medicago truncatula

label_protocol: 11 ug RNA from each of 3 biological replicates (9 plants total) was pooled to make a total of 33 ug for cDNA synthesis.  cDNA synthesis and fluorescent labeling was performed using the Genisphere Array 50 kit (Hatfield, PA USA) with a modified bulk-synthesis protocol that generated cDNA sufficient for 10 slide hybridizations.  32-40 mg total RNA in a volume of 8  l was mixed with 2  l of the Cy5 capture RT primer, then heated to 80oC for 10 min.  Tubes were immediately transferred to ice and then 1  l Superase-In RNase Inhibitor was added.  In a separate tube, 4  l 5X Superscript II RT buffer, 1  l 10 mM dNTPs, 2  l 0.1 M DTT, 1  l (200 U) Superscript II Reverse Transcriptase enzyme (Invitrogen, Carlsbad, CA USA) and nuclease-free ddH20 were mixed before addition to the RNA/primer combination.  Each tube was incubated for 2 h at 42oC, before the addition of 3.5  l of 0.5M NaOH/50mM EDTA and transfer to 65oC for 10 minutes to stop the reaction.  Finally, 5  l of 1 M Tris-HCL (pH 7.5) was added to neutralize the reaction.  cDNAs were stored at -20oC.  11 ug RNA from each of 3 biological replicates.

growth_protocol: Surface-sterilized Medicago truncatula seeds were vernalized at 40 C for 36 h, and then transferred to water-soaked Jiffy-7 peat pellets (Jiffy Product of America, Norwalk, OH, USA) that were placed inside clay pots containing SB-300 soil mix (Sungrow USA, Pine Bluff, AS, USA).  A pure culture of Erysiphe pisi that was established from a naturally-infected pea plant was obtained from Dr. Tim L.W. Carver, Department of Environmental Biology,  Institute of Grassland and Environmental Research, Plas Gogerddan, Aberystwyth, Ceredigion SY23 3EB, Wales, UK.  Cultures were maintained on Perfection 8771 pea plants grown in isolation at 22°C with a 16 h photoperiod (300 µmol/min/m2) and 8 h of darkness at 16°C.  Four-week-old M. truncatula plants with 2-3 fully expanded compound leaves were inoculated with approximately 300-500 spores/cm2 leaflet area using a 180 cm settling tower.  After 20 min, plants were returned to the growth chamber.  Control plants were grown in parallel in a separate chamber under the same conditions.

extract_protocol: Trifoliate leaf samples from 3 experimental replicates (9 plants total) were collected for the experiment. RNA from frozen leaf samples following the manufacturer's instructions.

tag: Medicago truncatula; lines - DZA315.16; tissue-trifoliate leaves; age - 4 weeks; treatment  12 hours after pathogen inoculation

source_name: Medicago truncatula (DZA315.16)  12 hours after no pathogen treatment (control)

label_protocol: 11 ug RNA from each of 3 biological replicates (9 plants total) was pooled to make a total of 33 ug for cDNA synthesis.  cDNA synthesis and fluorescent labeling was performed using the Genisphere Array 50 kit (Hatfield, PA USA) with a modified bulk-synthesis protocol that generated cDNA sufficient for 10 slide hybridizations.  32-40 mg total RNA in a volume of 8  l was mixed with 2  l of the Cy3 capture RT primer, then heated to 80oC for 10 min.  Tubes were immediately transferred to ice and then 1  l Superase-In RNase Inhibitor was added.  In a separate tube, 4  l 5X Superscript II RT buffer, 1  l 10 mM dNTPs, 2  l 0.1 M DTT, 1  l (200 U) Superscript II Reverse Transcriptase enzyme (Invitrogen, Carlsbad, CA USA) and nuclease-free ddH20 were mixed before addition to the RNA/primer combination.  Each tube was incubated for 2 h at 42oC, before the addition of 3.5  l of 0.5M NaOH/50mM EDTA and transfer to 65oC for 10 minutes to stop the reaction.  Finally, 5  l of 1 M Tris-HCL (pH 7.5) was added to neutralize the reaction.  cDNAs were stored at -20oC.  11 ug RNA from each of 3 biological replicates.

tag: Medicago truncatula; lines - DZA315.16; tissue-trifoliate leaves; age - 4 weeks; treatment  12 hours after no pathogen treatment (control)


title: stage hatching (Cy3) vs stage eye pigmentation (Cy5)

description: stage of hatching (Cy3) (48hpf) and stage of eye pigmentation (Cy5) (168hpf) of sea bream.

source_name: stage hatching (Cy3)

source_name: stage eye pigmentation (Cy5)


title: HUVEC transcriptional response to cigarette smoke extract after 7 hours_10443

description: No additional information.