title: Norm.1.col.Ref1.2 (Exiqon)

description: Hy3: CHIP # 2 flipUS22502706_13974287_S01_Exiqon1.txt

value: Reference 1

value: pool of equal amounts of total RNA from mouse testicle, ovary and 10-12 day embryo.


title: Myeloma-18

status: Public on Oct 18 2010

type: genomic

city: Sutton

first: Brian

middle: A

last: Walker

address: 15 Cotswold Road

department: Haemato-Oncology Research Unit

country: United Kingdom

institute: The Institute of Cancer Research

description: bisulfite modified genomic DNA

data_processing: files were analyzed in GenomeStudio (Illumina) as per manufacturers (Illumina) instructions using the methylation module v1.5.5, and are described as beta values

label: DNP/biotin

molecule: genomic DNA

organism: Homo sapiens

source_name: CD138+ bone marrow cells

growth_protocol: cell lines were grown in RPMI media with 10% FCS

extract_protocol: DNA was extracted using the AllPrep protocol (Qiagen)

treatment_protocol: all cells were derived from patients prior to treatment with chemotherapy

tag: tissue

value: bone marrow

tag: cell type

value: Multiple myeloma plasma cell

tag: myeloma translocation


title: Myeloma-19

description: bisulfite modified genomic DNA

hyb_protocol: as per manufacturers (Illumina) instructions


type: RNA

city: Seattle

first: Ilsa

last: Coleman

state: WA

address: 1100 Fairview Ave N, E2-112

department: Human Biology

country: USA

institute: Fred Hutchinson Cancer Center

description: benign epithelium from age- and BMI-matched control prostate needle biopsy #1.

hyb_protocol: Hybridizations to Agilent 44K whole human genome expression oligonucleotide microarray slides (Agilent Technologies, Inc., Santa Clara, CA) following the manufacturer’s suggested protocols.

scan_protocol: Fluorescence array images were collected for Cy3 and Cy5 using the Agilent DNA microarray scanner G2505B (Agilent Technologies, Inc., Santa Clara, CA).

data_processing: Agilent Feature Extraction (FE) software was used to grid, extract, and normalize data. Spots of poor quality or average intensity levels <300 were removed from further analysis.   The Agilent FE file "LogRatio" column data were converted from log10 scale to log2 scale.

molecule: total RNA

source_name: HGS [reference]

label_protocol: WTA amplified amino-allyl cDNA from each experimental sample was labeled with Cy3 fluorescent dye (reference amino-allyl cDNA was labeled with Cy5) following the manufacturer’s suggested protocols.

extract_protocol: To provide a reference standard RNA for use on two-color oligo microarrays, we pooled equal amounts of total RNA isolated from LNCaP, DU145, PC3, and CWR22 cell lines (American Type Culture Collection, Manassas, VA) growing at log phase in dye-free RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS) (Invitrogen, Carlsbad, CA).  Reference RNA was purified using TRIzol (Invitrogen, Carlsbad, CA) following the manufacturer’s protocol. Reference RNA was then further purified by RNeasy maxi kit and treated with DNAse using the RNase-Free DNase Set. This RNA was amplified using the TransPlex WTA Kit following procedures used for the study.

tag: gender

value: male

tag: reference pool

value: LNCaP, DU145, PC3, and CWR22 cell lines

source_name: 1_BABE [test]

extract_protocol: Total RNA from laser-capture microdissected experimental samples was isolated using the Arcturus PicoPure RNA Isolation kit (Molecular Devices, Sunnyvale, CA) and DNAse-treated using the RNase-Free DNase Set (Qiagen Inc, Valencia, CA.) This RNA was converted to a cDNA library and amplified and amino-allyl labeled using the TransPlex WTA Kit (Sigma-Aldrich Corp, St. Louis, MO) and its purity was confirmed by spectrophotometric analysis.

tag: specimen name

tag: ethnicity

value: Caucasian

value: benign epithelium

tag: disease state

value: control


title: Norm.2.col.Ref1.2 (Exiqon)

city: Ottawa

first: Andrew

last: Williams

state: ON

address: 50 Columbine

country: Canada

institute: Health Canada

description:
Hy3: CHIP # 2 flipUS22502706_13974287_S01_Exiqon1.txt
Hy5: CHIP # 4 flipUS22502706_13974278_S01_Exiqon1.txt

hyb_protocol: Hybridisations were performed in SureHyb chambers (Agilent) for 16 hours at 56ºC. Slides were washed according to manufacturer’s instructions

scan_protocol: Arrays were scanned at a resolution of 5 mm using an Agilent G2505B scanner. Data were acquired using Agilent Feature Extraction software version 9.5.3.1, using an FE protocol available on-demand from Exiqon.

data_processing: Data were extracted from scanned images using Agilent Feature Extraction software version 9.5.3.1. Median signal intensities were lowess normalized for the 2 colour analysis and cyclic lowess normalized for the one colour analysis.  Technical replicates for each miRNA level were averaged using the median.  Data was flagged if the raw signal intensity was less than the mean plus three standard deviations of the negative controls or if the median of the technical replicates was flagged to be in the background.

label: Hy3 ,Hy5

organism: Mus musculus

source_name: Total RNA with small fraction

label_protocol: One microgram of each mouse Reference RNA sample was labelled with a Hy5 fluorophore (Exiqon, Woburn, MA, USA) and co-hybridised with 1 g of the other Reference RNA standard in Hy3. Labelling reactions were performed using Exiqon’s miRCURY LNA miRNA power labelling kit, according to the manufacturer’s protocol.

extract_protocol: Total RNA containing the small fraction was isolated from each sample using the miRVana miRNA Isolation Kit (Ambion), Streetsville, ON, Canada). Quality was confirmed using the Agilent 2100 Bioanalyzer and RNA Nano 6000 chips.

tag: sample

tag: sample type

value: Reference RNA was developed from FirstChoice® mouse Total RNA including the small fraction (Catalogue # AM7800-AM7828, Ambion, Streetsville, ON)


title: Norm.2.col.Ref2.2 (Exiqon)

description:
Hy3: CHIP # 4 flipUS22502706_13974278_S01_Exiqon1.txt
Hy5: CHIP # 2 flipUS22502706_13974287_S01_Exiqon1.txt

value: Reference 2

value: pool of equal parts RNA from heart, lung and liver


title: FO-regenerated5

city: Hong Kong

first: Mary

last: Lee

address: 21 sassoon Road

country: Hong Kong

institute: The University of Hong Kong

hyb_protocol: The biotinylated cRNA targets were then isolated with an RNeasy Mini Kit (Qiagen) and precipitated with ethanol. The cRNA was fragmented to 35 to 200 base fragments before hybridization onto GeneChip expression arrays. The GeneChip® porcine genome array used contained 23,937 probe sets that interrogate approximately 23,256 transcripts from 20,201 S. scrofa genes for target hybridization.

scan_protocol: Probe arrays were scanned using an Affymetrix® GeneChip® Scanner 3000.

data_processing:
Comparison analysis of two single arrays from native and regenerated endothelial cells of the same heart was done to evaluate the differential gene expression in the regenerated cells. The intensity for each feature of the array was captured with Affymetrix GeneChip Expression Analysis (GeneChip Operating System) Software according to standard Affymetrix procedures. Expression values for each transcript in native and regenerated endothelial cells were normalized using GeneSpring GX10.0.2 Analysis Software (Agilent Technologies, Santa Clara, CA, USA). Two analytical algorithms (standard guided workflow and MAS5.0) for gene expression analysis were used.  The detailed description of each probe set ID of the GeneChip® porcine genome array can be found at the Affymetrix website (http://www.affymetrix.com/index.affx).
Paired comparison of cultures derived from native and regenerated endothelial cells of the same pig hearts: The present microarray findings are represented as a paired analysis of grouped experiments. For each pair of microarray data, the mRNA expressions from native and regenerated endothelial cells of the same heart were compared, to minimize the genetic background noise. The raw data were normalized following two different strategies (GeneSpring GX 10.0.2) for data processing and statistical analysis. First, the standard guided workflow for gene expression analysis was used. Briefly, after the experimental grouping of native and regenerated cells, interpretation of native database to regenerated database was created, the data quality check to understand the similarity of gene changes among samples of the same group was performed, and the grouped data were normalized. The data sets (both native and regenerated cells) were filtered using Student's t-test for paired observations to produce lists of statistically significantly changed genes from both CHL and FO fed pigs. In addition, a list of genes was also produced, without performing any statistical analysis, by filtering the average fold-changes greater than two-fold. Second, the MAS5.0 for gene expression analysis was applied, using a similar statistical analysis (Student's t-test for paired observation) to produce a list of significantly changed genes for subsequent analysis. The three lists could then be combined to produce a complete list of gene changes due to endothelial regeneration in both CHL and FO groups.
Unpaired comparison of cultures derived from native endothelial cells of pig heart from control and CHL/FO diets: Similarly, a list of statistical significant genes was produced by comparing the native cells of animals with control diet to those with CHL or FO diet and analyzed using Student's t-test for unpaired observations, based on the two strategies described above (guided work flow and MAS5.0 system).

MAS5.0 data presented below.

label: biotin

organism: Sus scrofa

source_name: regenerated endothelial cells from farm pig

label_protocol: High quality RNAs from native and regenerated endothelial cells were examined with GeneChips® Porcine Genome Arrays (Affymetrix; Santa Clara, CA, USA), according to the manufacturer’s instructions. Less than 250 ng of total RNA was first reverse transcribed using a T7-Oligo(dT) Promoter Primer in the first-strand cDNA synthesis reaction. Following RNase H-mediated second-strand cDNA synthesis, the double-strand cDNA was purified and served as a template in the subsequent in vitro transcription (IVT) reaction. The IVT reaction was carried out in the presence of T7 RNA Polymerase and a biotinylated nucleotide analog/ribonucleotide mix for complementary RNA (cRNA) amplification and biotin labeling.

growth_protocol: The pig hearts were collected and transferred into cold Earle’s balanced salt solution (EBSS) supplemented with 5% fetal bovine serum (FBS). Native (left circumflex artery) and regenerated (previously denuded part of the LAD) endothelial cells were collected as follows. After clearing all the adhering fat, the arteries were cut longitudinally, exposing the luminal area. Isolation of endothelial cells was done by gently scraping the luminal surface with a scalpel blade and collecting the fragments of endothelial lining in culture medium. They were centrifuged at 1,250 rpm for five minutes and resuspended in Earle’s minimum essential medium (EMEM) containing 15% FBS in a 60 mm collagen-coated Petri dish. The cells were cultured at 37C in a humidified gas mixture (5% CO2, 95% O2). The medium was changed every two days to facilitate cell growth. The cultures derived from native and regenerated endothelial cells are termed native and regenerated cells, respectively.

extract_protocol: Native and regenerated endothelial cells were cultured for ten days to reach approximately 80% confluence before isolation of the RNA for microarray analysis. Primary cultures (cells at passage zero) were used for the analysis. Briefly, the cells were rinsed with cold phosphate buffer solution (PBS) twice and total RNA was extracted directly with RNA lysis thiocyanate (RLT) buffer (containing 25-50% guanidinium thiocyanate) using the RNeasy Mini kit (Qiagen; Valencia, CA, USA). Samples were stored immediately at -70C until use. The total amount of RNA was quantified by measuring the optical density at 260 nm. The ratio of absorption (260/280) of all samples was between 1.7 and 2.1. The integrity of RNA samples was assessed qualitatively [rRNA ratio for 28s/18s between 1.9 and 2.1].

value: female

tag: age

value: 4 months old

value: endothelial cell

tag: endothelial cell status

value: regenerated (LAD)

tag: agent

value: FO


title: FO-native6

source_name: native endothelial cells from farm pig

value: native (LCX)


title: Norm.1.col.Ref2.1 (Exiqon)

description: Hy3: CHIP # 3 flipUS22502706_13974276_S01_Exiqon1.txt


title: FO-regenerated4


title: FO-regenerated6


title: Norm.2.col.Ref2.1 (Exiqon)

description:
Hy3: CHIP # 3 flipUS22502706_13974276_S01_Exiqon1.txt
Hy5: CHIP # 1 flipUS22502706_13974284_S01_Exiqon1.txt


title: Norm.1.col.Ref1.1 (Exiqon)

description: Hy3: CHIP # 1 flipUS22502706_13974284_S01_Exiqon1.txt

value: Reference 1

value: pool of equal amounts of total RNA from mouse testicle, ovary and 10-12 day embryo.


title: FO-native4


title: Norm.2.col.Ref1.1 (Exiqon)

description:
Hy3: CHIP # 1 flipUS22502706_13974284_S01_Exiqon1.txt
Hy5: CHIP # 3 flipUS22502706_13974276_S01_Exiqon1.txt


title: Myeloma-17

description: bisulfite modified genomic DNA

scan_protocol: as per manufacturers (Illumina) instructions

value: bone marrow

value: Multiple myeloma plasma cell


title: D_D23

description: no additional information

hyb_protocol: Hy3 and Hy5 labelled samples were cohybridised to Exiqon multi-species miRCURY LNA miRNA array slides (version 11.0), containing ~1700 capture probes covering all human, mouse and rat miRNAs annotated in miRBase 11.0. Hybridisations were performed in SureHyb chambers (Agilent) for 16 h at 56oC and slides were washed according to manufacturer’s instructions.

scan_protocol: Arrays were scanned at a resolution of 5 um using an Agilent G2505B scanner.

data_processing:
Data were acquired using Agilent Feature Extraction software version 9.5.3.1, using an FE protocol available on-demand from Exiqon. Technical replicates were averaged using the median removing probes flagged as population background outliers.  The averaged signal intensities were then quantile normalized.  Data was flagged if the averaged signal intensity signal intensity was less than the mean plus three standard deviations of the negative controls (hsa_negative_control_1).

Sample Data table header definitions:
RawHy5 and RawHy3 is the average (over the technical replicates) of the median signal intensities for the Hy5 and Hy3 channels.
NormHy5 and NormHy3 is the quantile normalized signal intensity (log2).
Flag indicates whether the averaged signal intensity included a spot that had a median signal intensity within the background (mean plus three standard deviations of the negative controls).
Ratio (i.e., VALUE) is the log2 of the Ratio Hy3/Hy5.

source_name: Total RNA, mouse liver, BaP exposure

label_protocol: Labelling reactions were performed using Exiqon’s miRCURY LNA miRNA power labelling kit, according to the manufacturer’s protocol.

value: Median liver lobe

value: Male

tag: strain

tag: exposure

value: 150 mg/kg Benzo(a)pyrene (CAS number 50-32-8)

tag: time post exposure

value: 4 h

tag: rna storage

value: <7 days at -80C

source_name: Pooled mouse reference RNA

extract_protocol: Purchased from Ambion

tag: reference

value: Pooled total RNA (containing small fraction) derived from 10 mouse tissues


title: C_D03

description: no additional information

source_name: Total RNA, mouse liver

tag: tissue

value: 0 mg/kg BaP


title: FO-native5

value: female

tag: cell type


title: D_D21

description: no additional information

tag: gender


title: Myeloma-15

description: bisulfite modified genomic DNA

label_protocol: as per manufacturers (Illumina) instructions

tag: myeloma translocation


title: Myeloma-16

description: bisulfite modified genomic DNA


title: Myeloma-14

description: bisulfite modified genomic DNA


title: Myeloma-128

description: bisulfite modified genomic DNA

value: Not known


title: FO-regenerated3

value: 4 months old

value: endothelial cell


title: Myeloma-13

description: bisulfite modified genomic DNA


title: FO-native3

value: FO


title: D_D27

description: no additional information


title: D_D25

description: no additional information


title: C_D01

description: no additional information


title: C_D05

description: no additional information


title: C_D07

description: no additional information


title: FO-native1

value: native (LCX)


title: Myeloma-127

description: bisulfite modified genomic DNA

value: No split


title: FO-native2


title: FO-regenerated2

value: regenerated (LAD)


title: Myeloma-126

description: bisulfite modified genomic DNA

value: bone marrow


title: D_D26

description: no additional information


title: CHL-regenerated6

value: CHL


title: Myeloma-125

description: bisulfite modified genomic DNA

value: Multiple myeloma plasma cell


title: C_D06

description: no additional information

value: Median liver lobe

value: Male


title: FO-regenerated1


title: D_D22

description: no additional information

value: 150 mg/kg Benzo(a)pyrene (CAS number 50-32-8)

value: 4 h


title: Myeloma-124

description: bisulfite modified genomic DNA


title: C_D02

description: no additional information

value: <7 days at -80C

value: Pooled total RNA (containing small fraction) derived from 10 mouse tissues


title: Myeloma-123

description: bisulfite modified genomic DNA


title: CHL-native5

value: female


title: Myeloma-122

description: bisulfite modified genomic DNA


title: CHL-regenerated5


title: Reference_Slide 5

description: Pool

hyb_protocol: Hybridisations were performed in Agilent SureHyb chambers for 20 hours at 55C and washed according to manufacturer's instructions.

scan_protocol: Arrays were scanned at a resolution of 5 mm using an Agilent G2505B scanner.

data_processing:
Data were extracted from scanned images using Agilent Feature Extraction software version 9.5.3.1. Median signal intensities for the 4th and 24hr time points were quantile normalized separately.  Technical replicates for Probe level and miRNA level were averaged using the median.  Data was flagged if the raw signal intensity was less than the mean plus three standard deviations of the negative controls or if the median of the technical replicates was flagged to be in the background.

Sample Data table header definitions:
Raw is the median signal intensity.
Norm (i.e., VALUE) is the quantile normalized signal intensity of the log2 of the median signal intensity.
Flag indicates whether the raw signal intensity for the spot is within the background (mean plus three standard deviations of the negative controls).
Norm.Collapsed.Probe is the median of the probe technical replicates.
Flag.Probe indicates whether the median of the probe technical replicates was within the background.
Norm.Collapsed.miRNA is the median of the miRNA technical replicates.
Flag.miRNA indicates whether the median of the miRNA technical replicates was within the background.

source_name: Total RNA, mouse liver, pooled 6 control + 6 BaP treated

label_protocol: Total RNA samples were labelled using the Agilent miRNA Complete Labelling and Hybridisation Kit (Agilent Tech, Mississauga, ON, Canada).

value: Pooled mouse reference RNA (equal proportions all samples)

value: 6-8 weeks at -80C


title: CHL-regenerated4

value: 4 months old

value: endothelial cell


title: CHL-native4

value: native (LCX)


title: CHL-regenerated3


title: Myeloma-121

description: bisulfite modified genomic DNA

value: No split


title: CHL-native6


title: Myeloma-120

description: bisulfite modified genomic DNA


title: Myeloma-119

description: bisulfite modified genomic DNA


title: CHL-regenerated2

value: regenerated (LAD)


title: Myeloma-12

description: bisulfite modified genomic DNA

value: bone marrow


title: CHL-native3

value: CHL


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #30.

value: male

tag: reference pool

value: LNCaP, DU145, PC3, and CWR22 cell lines

source_name: 30_CABE [test]

tag: specimen name

value: Caucasian

value: benign epithelium

value: prostate cancer

tag: tumor grade

value: Gleason 8-10


title: CHL-native2


title: Myeloma-118

description: bisulfite modified genomic DNA

value: Multiple myeloma plasma cell


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #29.

source_name: 29_CABE [test]

tag: ethnicity

tag: disease state


title: CHL-native1

value: female

tag: age


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #24.

source_name: 24_BABE [test]

value: control


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #28.

source_name: 28_CABE [test]


title: Myeloma-117

description: bisulfite modified genomic DNA


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #27.

source_name: 27_CABE [test]

value: non-Caucasian


title: Myeloma-116

description: bisulfite modified genomic DNA


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #26.

source_name: 26_CABE [test]


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #25.

source_name: 25_BABE [test]


title: Myeloma-111

description: bisulfite modified genomic DNA

source_name: CD138+ bone marrow cells


title: Myeloma-112

description: bisulfite modified genomic DNA


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #18.

source_name: HGS [reference]

source_name: 18_CABE [test]


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #22.

source_name: 22_BABE [test]


title: control-regenerated8

source_name: regenerated endothelial cells from farm pig

value: 4 months old

tag: endothelial cell status


title: Myeloma-113

description: bisulfite modified genomic DNA


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #19.

source_name: 19_CABE [test]


title: control-native6

source_name: native endothelial cells from farm pig

value: endothelial cell

tag: agent


title: Myeloma-115

description: bisulfite modified genomic DNA


title: Myeloma-114

description: bisulfite modified genomic DNA


title: control-regenerated7


title: control-native7

value: native (LCX)


title: CHL-regenerated1


title: Myeloma-110

description: bisulfite modified genomic DNA

value: No split


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #21.

source_name: 21_BABE [test]


title: control-regenerated6

value: regenerated (LAD)


title: control-native8


title: Myeloma-11

description: bisulfite modified genomic DNA

value: bone marrow


title: control-native5


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #20.

source_name: 20_CABE [test]


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #15.

source_name: 15_BABE [test]


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #17.

source_name: 17_CABE [test]


title: Myeloma-109

description: bisulfite modified genomic DNA


title: control-regenerated5


title: Myeloma-108

description: bisulfite modified genomic DNA


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #14.

source_name: 14_BABE [test]


title: Myeloma-107

description: bisulfite modified genomic DNA


title: control-native4


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #12.

source_name: 12_BABE [test]


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #13.

source_name: 13_BABE [test]


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #10.

source_name: 10_CABE [test]


title: Myeloma-106

description: bisulfite modified genomic DNA


description: benign epithelium from prostate needle biopsy with high grade(Gleason 8-10) prostate cancer #16.

source_name: 16_CABE [test]


title: Myeloma-103

description: bisulfite modified genomic DNA


description: benign epithelium from age- and BMI-matched control prostate needle biopsy #5.

source_name: 5_BABE [test]


title: control-regenerated4


title: control-regenerated3


title: control-native3


title: control-native2


title: Myeloma-104

description: bisulfite modified genomic DNA

source_name: CD138+ bone marrow cells


title: Myeloma-105

description: bisulfite modified genomic DNA


title: Norm.1.col.Ref1.2 (NCode)

description: Alexa Fluor 5: 3780_reformat.txt

hyb_protocol: Arrays were hybridized 10 hours at 52°C. In a modification of the standard protocol, hybridizations were performed using Agilent hybridization chambers within a rotisserie oven, which required dilution of the hybridization mix to a total volume of 120 ul, by addition of BSA (8 mg/ml) and Enhanced Hybridization Buffer (Invitrogen). Arrays were washed and dried by centrifugation according to manufacturer’s instructions. To protect dyes against ozone-mediated degradation, 0.1 mM dithiothreitol was added to each wash solution, and slides were dried and stored in sealed tubes containing 20 ul of ≥ 98% 2-mercaptoethanol, ensuring that there was no contact between the reducing agent and the array.

scan_protocol: Arrays were scanned with a GenePix 4000B laser scanner (Molecular Devices) with a scan resolution of 5 um, and PTM between 620 to 660 V. Signal intensity was determined using Imagene 8.0 image analysis software (Biodiscovery, Inc.).

data_processing: Median signal intensities were lowess normalized for the 2 colour analysis and cyclic lowess normalized for the one colour analysis Alexa Fluor 5 channel only).  Technical replicates for each miRNA level were averaged using the median.  Data was flagged if the raw signal intensity was less than the mean plus three standard deviations of the negative controls or if the median of the technical replicates was flagged to be in the background.

label: Alexa Fluor 5

source_name: Total RNA with small fraction

label_protocol: 0.5 ug of total reference RNA was spiked with 40 fmoles NCode Microarray Positive Control (Invitrogen) and labelled using the NCode Rapid miRNA Labelling System according to manufacturer’s instruction (Invitrogen). Briefly, polyA polymerase is used to add a poly(A) tail to the RNA, to which is then ligated a DNA polymer carrying ~15 molecules of the fluorophore (Alexa Fluor® 3 or 5). Ligation is mediated by an oligo(dT) bridge whose sequence matches both the poly(A) tail of the RNA, and the fluorophore-labelled DNA polymer.

tag: sample

value: Reference 1

value: pool of equal amounts of total RNA from mouse testicle, ovary and 10-12 day embryo.

tag: sample type

value: Reference RNA was developed from FirstChoice® mouse Total RNA including the small fraction (Catalogue # AM7800-AM7828, Ambion, Streetsville, ON)


title: Norm.1.col.Ref2.2 (NCode)

description: Alexa Fluor 5: 3781_reformat.txt

value: Reference 2

value: pool of equal parts RNA from heart, lung and liver


title: Norm.2.col.Ref2.2 (NCode)

description:
Alexa Fluor 3: 3780_reformat.txt
Alexa Fluor 5: 3781_reformat.txt

label: Alexa Fluor 3 , Alexa Fluor 5


title: control-regenerated2


title: Norm.2.col.Ref1.2 (NCode)

description:
Alexa Fluor 3: 3781_reformat.txt
Alexa Fluor 5: 3780_reformat.txt


title: Norm.2.col.Ref2.1 (NCode)

description:
Alexa Fluor 3: 3863_reformat.txt
Alexa Fluor 5: 3779_reformat.txt


title: control-native1


title: Norm.1.col.Ref2.1 (NCode)

description: Alexa Fluor 5: 3779_reformat.txt


title: Myeloma-102

description: bisulfite modified genomic DNA


title: control-regenerated1


title: Myeloma-1

description: bisulfite modified genomic DNA


title: Myeloma-101

description: bisulfite modified genomic DNA


title: Norm.2.col.Ref1.1 (NCode)

description:
Alexa Fluor 3: 3779_reformat.txt
Alexa Fluor 5: 3863_reformat.txt


title: Myeloma-10

description: bisulfite modified genomic DNA


title: Myeloma-100

description: bisulfite modified genomic DNA


title: MGUS4

description: bisulfite modified genomic DNA


title: MGUS3

description: bisulfite modified genomic DNA


title: U3A_IRF7+IKKe_20h_rep1

city: New York

first: Benjamin

last: tenOever

state: NY

address: 430 East 29th Street, Room 540

department: Microbiology

institute: New York University

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 7GGeneChips were scanned using the GeneChip Array Scanner 7G

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

source_name: U3A cells, treated with type I IFN and transfected with IRF7 and IKKe, 20h

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

growth_protocol: See Schmid et. Al. JBC 2010

extract_protocol: Trizol

treatment_protocol: See Schmid et. Al. JBC 2010

tag: cell line

value: U3A cells (fibrosarcoma), deficient in STAT1

tag: treatment

value: 25 IU/ml type I IFN

tag: transfection

value: IRF7 + IKKe

tag: time post transfection


title: MGUS2

description: bisulfite modified genomic DNA

tag: tissue

tag: cell type

value: Multiple myeloma plasma cell


title: MGUS1

description: bisulfite modified genomic DNA


title: U3A_IRF7+IKKe_20h_rep2


title: Norm.1.col.Ref1.1 (NCode)

description: Alexa Fluor 5: 3863_reformat.txt


title: Bcell-6

description: bisulfite modified genomic DNA

source_name: CD19+ peripheral blood cells

value: peripheral blood

value: B cell


title: Bcell-5

description: bisulfite modified genomic DNA


title: LCM mammary gland tumor from Her2/Neu mouse #12

hyb_protocol: A total of 825 ng of amplified amino-allyl aRNA from each experimental sample was hybridized to Agilent 44K whole human genome expression oligonucleotide microarray slides (Agilent Technologies, Inc., Santa Clara, CA) following the manufacturer’s suggested protocols.

scan_protocol: Fluorescence array images were collected for Cy3 and Cy5 using the Agilent DNA microarray scanner G2565BA (Agilent Technologies, Inc., Santa Clara, CA).

data_processing: Agilent Feature Extraction software was used to grid, extract, and normalize data. Spots of poor quality or average intensity levels <300 were removed from further analysis.

source_name: MGS

label_protocol: Reference total RNA was amplified two rounds using the Ambion MessageAmp aRNA Kit (Ambion Inc, Austin, TX), incorporating amino-allyl UTP into amplified antisense RNA. A total of 825 ng of amplified amino-allyl aRNA from each sample was labeled with Cy5 fluorescent dye following the manufacturer's suggested protocols.

extract_protocol: To provide a reference standard RNA for use on two-color cDNA microarrays, we used pooled total RNA from normal adult male Swiss-Webster mice (10% prostate and 30% each testis, liver, and kidney.) Reference Total RNA was purified using Trizol (Life Technologies, Rockville, MD) following the manufacturer's protocol and further purified by RNeasy maxi kit incorporating DNAse treatment using the RNase-Free DNase Set (Qiagen Inc, Valencia, CA.)

tag: gender

value: male

tag: strain

value: Swiss-Webster

value: reference pool of normal adult tissues (10% prostate and 30% each testis, liver, and kidney)

label_protocol: Experimental total RNA was amplified two rounds using the Ambion MessageAmp aRNA Kit (Ambion Inc, Austin, TX), incorporating amino-allyl UTP into amplified antisense RNA. A total of 825 ng of amplified amino-allyl aRNA from each sample was labeled with Cy3 fluorescent dye following the manufacturer's suggested protocols.

growth_protocol: All mouse work was performed under IACUC regulations as approved by the FHCRC animal use committee.  For these studies, a doxycycline-inducible, bitransgenic MMTV-rtTA/TetO-NeuNT (Her2/Neu) mouse model was used.  Corresponding control mice were transgenic for TetO-NeuNT only and were littermates of the bitransgenic mice.  The procedures for animal husbandry and tissue and plasma sample collection were previously reported.  Briefly, all mice were fed standard laboratory chow (Harland Teklad, 8664) and acidified water ad libitum and kept on a 12 hour light-dark cycle.

extract_protocol: Total RNA from laser-capture microdissected experimental samples was isolated using the Arcturus PicoPure RNA Isolation kit (Molecular Devices, Sunnyvale, CA) incorporating DNAse treatment using the RNase-Free DNase Set (Qiagen Inc, Valencia, CA.)

treatment_protocol: All mice received doxycycline (2 mg/mL + 5% sucrose) in the drinking water starting at 8 weeks of age.  Doxycycline was used to induce tumor development in the bitransgenic mice and breast tumors were allowed to develop to 1.0 cm in diameter after which each tumor-bearing mouse and its corresponding control littermate were euthanized by CO2 inhalation.  For the 3 day and 6 day regression experiments, doxycycline was withdrawn from the drinking water after the tumor sizes had reached ~1 cm, causing a visible decrease in mammary tumor size, followed by euthanization. Tissue samples were collected when the tumors reached ~1 cm size and the samples were snap-frozen in liquid nitrogen.

value: female

value: FVB

tag: genotype

value: mammary gland tumor (5000 cells laser-capture microdissected from frozen OCT block)


title: LCM mammary gland tumor from Her2/Neu mouse #6

value: doxycycline-inducible, bitransgenic MMTV-rtTA/TetO-NeuNT (Her2/Neu) mouse model


title: Ref1.2 (Agilent)

description: Reference RNA was developed from FirstChoice® mouse Total RNA including the small fraction (Catalogue # AM7800-AM7828, Ambion, Streetsville, ON)

data_processing: Data were extracted from scanned images using Agilent Feature Extraction software version 9.5.3.1. Median signal intensities were cyclic lowess normalized.  Technical replicates for each miRNA level were averaged using the median.  Data was flagged if the raw signal intensity was less than the mean plus three standard deviations of the negative controls or if the median of the technical replicates was flagged to be in the background.

value: pool of equal amounts of total RNA from mouse testicle, ovary and 10-12 day embryo.

tag: microarray barcode


title: U3A_GFP_20h_rep2

source_name: U3A cells, treated with type I IFN and transfected with GFP, 20h

value: GFP


title: LCM mammary gland tumor from Her2/Neu mouse #4


title: Bcell-4

description: bisulfite modified genomic DNA


title: Ref2.2 (Agilent)


title: pool of LCM benign mammary gland epithelium from two Her2/Neu strain-matched control mice replicate #2

value: control mice transgenic for TetO-NeuNT only and littermates of the bitransgenic mice

value: pool of benign mammary gland epithelium from two mice (5000 cells laser-capture microdissected from each frozen OCT block)


title: pool of LCM benign mammary gland epithelium from two Her2/Neu strain-matched control mice replicate #1


title: Bcell-3

description: bisulfite modified genomic DNA


title: U3A_GFP_20h_rep1

value: U3A cells (fibrosarcoma), deficient in STAT1


title: Bcell-1

description: bisulfite modified genomic DNA


title: Bcell-2

description: bisulfite modified genomic DNA


title: Ref2.1 (Agilent)

value: pool of equal parts RNA from heart, lung and liver


title: S_5_xeno:ALL case#5 :xenograft splenocyte

city: Bethesda

first: Sean

last: Davis

state: MD

address: 37 Convent Drive, Room 6138

institute: National Cancer Institute

description: ALL case#5 :xenograft splenocyte

hyb_protocol: Hybridization was to HumanWG-6_v3 arrays per manufacturer's instructions.

scan_protocol: Images were scanned using the Illumina BeadScan software.

data_processing: Data were processed using the Bioconductor package, lumi, using default values for background correction, variance stabilization transformation, and quantile normalization.

source_name: ALL case#5 :xenograft splenocyte

label_protocol: Biotinylated cRNA targets were amplified by in vitro transcription using the TotalPrep-96 RNA Amplification Kit (Ambion)

extract_protocol: Thawed cell suspension was pelleted, resuspended in cold PBS and lysed in TRIzol Reagent (Invitrogen).  RNA was isolated using a combined protocol, where the TRIzol extraction aqueous phase was mixed with an equal volume of 70% EtOH, applied to a Rneasy Midi column (Qiagen) and purified according to the manufactures instructions.  RNA integrity was verified on a BioAnalyzer (Agilent). cDNA was synthesized by reverse transcription from 200 ng of total RNA and biotinylated cRNA targets were amplified by in vitro transcription using the TotalPrep-96 RNA Amplification Kit (Ambion)

tag: sample tissue of origin

value: xenograft splenocyte

tag: clinical status at collection

value: Diagnosis

value: M

tag: clinical status

value: CCR (4)

tag: major cytogenetic abnormality

tag: percent blasts in human

tag: cd10 in human

tag: cd19 in human

tag: cd20 in human

tag: cd22 in human

tag: cd34 in human

tag: cd45 in human

tag: cd13 in human

tag: cd33 in human


title: Ref1.1 (Agilent)

tag: sample type


title: S_4_xeno:ALL case#4 :xenograft splenocyte

description: ALL case#4 :xenograft splenocyte

source_name: ALL case#4 :xenograft splenocyte

tag: age

tag: gender

value: DOD


title: PB428_Nsp_peripheral blood white cells

hyb_protocol: 90 ug of fragmented labeled DNA is hybridised to the array for 16 hours at 49C accoring to Affymetrix protocols.

scan_protocol: Genechips were scanned on an Affymetrix GeneChip 3000 7G/4 color early acces scanner and processed using GCOS 1.4.0.

data_processing: SNP genotype calls were generated using Affymetrix Powertools  v1.8.0 (apt-probeset-genotype).  Calls were generated using the default parameters and the brlmm calling algorithm.

source_name: non-tumour peripheral blood white cells

label_protocol: 250 ng of DNA is digested with NspI/StyI, and processed accorodng to Affymetrix 500K protocol.  DNA is end-labeled with biotin using TdT.

extract_protocol: DNA was extracted from peripheral white blood cell pellets using the Qiagen Flexigene kit, as per manufacturer's instructions.

tag: extraction

value: Flexigene

tag: cell type

value: non-tumour peripheral blood white cells


title: PB418_Nsp_peripheral blood white cells


title: S_3_xeno:ALL case#3 :xenograft splenocyte

description: ALL case#3 :xenograft splenocyte

source_name: ALL case#3 :xenograft splenocyte

tag: sample tissue of origin

tag: clinical status at collection

value: Relapse #2

value: F

value: Not done


title: PB409_Nsp_peripheral blood white cells


title: PB429_Nsp_peripheral blood white cells


title: S_2_xeno:ALL case#2 :xenograft splenocyte

description: ALL case#2 :xenograft splenocyte

source_name: ALL case#2 :xenograft splenocyte

value: Relapse #1

tag: clinical status

tag: major cytogenetic abnormality


title: PB401_Nsp_peripheral blood white cells


title: PB407_Nsp_peripheral blood white cells


title: PB400_Nsp_peripheral blood white cells


title: S_5_human:ALL case#5 :Bone Marrow

description: ALL case#5 :Bone Marrow

source_name: ALL case#5 :Bone Marrow

value: Bone Marrow

tag: percent blasts in human

tag: cd10 in human


title: S_4_human:ALL case#4 :Bone Marrow

description: ALL case#4 :Bone Marrow

source_name: ALL case#4 :Bone Marrow

tag: cd19 in human

tag: cd20 in human


title: MM428_Nsp_CD138_selected_plasma_cells_presentation

source_name: pre-treatment bone marrow

extract_protocol: Sample was CD138 positively selected using MACS beads and stored in RLT, extracted using Qiagen AllPrep as per manufacturer's protocol.

value: AllPrep

value: pre-treatment bone marrow


title: PB418_Sty_peripheral blood white cells


title: PB403_Nsp_peripheral blood white cells


title: S_1_xeno:ALL case#1 :xenograft splenocyte

description: ALL case#1 :xenograft splenocyte

source_name: ALL case#1 :xenograft splenocyte

tag: cd22 in human

tag: cd34 in human


title: PB429_Sty_peripheral blood white cells


title: PB407_Sty_peripheral blood white cells


title: PB428_Sty_peripheral blood white cells


title: PB404_Nsp_peripheral blood white cells


title: S_3_human:ALL case#3 :Bone Marrow

description: ALL case#3 :Bone Marrow

source_name: ALL case#3 :Bone Marrow

tag: cd45 in human

tag: cd13 in human


title: MM429_Nsp_CD138_selected_plasma_cells_presentation


title: MM421_Nsp_CD138_selected_plasma_cells_presentation


title: PB409_Sty_peripheral blood white cells


title: MM422_Nsp_CD138_selected_plasma_cells_presentation


title: MM424_Nsp_CD138_selected_plasma_cells_presentation


title: MM426_Nsp_CD138_selected_plasma_cells_presentation


title: S_1_human:ALL case#1 :Peripheral Blood

description: ALL case#1 :Peripheral Blood

source_name: ALL case#1 :Peripheral Blood

value: Peripheral Blood

tag: cd33 in human


title: PB400_Sty_peripheral blood white cells


title: PB403_Sty_peripheral blood white cells


title: PB401_Sty_peripheral blood white cells


title: MM427_Nsp_CD138_selected_plasma_cells_presentation


title: MM423_Nsp_CD138_selected_plasma_cells_presentation


title: MM420_Nsp_CD138_selected_plasma_cells_presentation


title: S_2_human:ALL case#2 :Peripheral Blood

description: ALL case#2 :Peripheral Blood

source_name: ALL case#2 :Peripheral Blood

tag: age


title: MM428_Sty_CD138_selected_plasma_cells_presentation


title: MM429_Sty_CD138_selected_plasma_cells_presentation


title: MM425_Nsp_CD138_selected_plasma_cells_presentation


title: MM417_Nsp_CD138_selected_plasma_cells_presentation


title: MM419_Nsp_CD138_selected_plasma_cells_presentation


title: eto1-4_3-day-old_rep2

city: Taipei

first: Kevin Long-Chi

last: WANG

address: 128 Section 2, Academia Road

department: Institute of Plant and Microbial Biology

country: Taiwan

institute: Academia Sinica

hyb_protocol: Standard Affymetrix GeneChip hybridization protocol

scan_protocol: Standard Affymetrix GeneChip scanning protocol

data_processing: MAS5.0 using default settings

organism: Arabidopsis thaliana

label_protocol: Standard Affymetrix GeneChip labeling protocol

growth_protocol: Seedling were growth on 0.5x MS medium

extract_protocol:
Pine Tree method, for preparation of total RNA from etiolated seedlingss
Plant Molecular Biology Reporter (1993) 11: 113-116

tag: ecotype

tag: genome/variation

value: ethylene mutant (eto1-4)

tag: developmental stage

tag: tissue

value: whole seedling

tag: growth condition

value: dark


title: MM418_Nsp_CD138_selected_plasma_cells_presentation

tag: extraction

tag: cell type


title: PB404_Sty_peripheral blood white cells


title: MM427_Sty_CD138_selected_plasma_cells_presentation


title: MM426_Sty_CD138_selected_plasma_cells_presentation


title: MM415_Nsp_CD138_selected_plasma_cells_presentation


title: MM416_Nsp_CD138_selected_plasma_cells_presentation


title: MM414_Nsp_CD138_selected_plasma_cells_presentation


title: MM425_Sty_CD138_selected_plasma_cells_presentation


title: MM412_Nsp_CD138_selected_plasma_cells_presentation


title: MM419_Sty_CD138_selected_plasma_cells_presentation


title: MM424_Sty_CD138_selected_plasma_cells_presentation


title: MM422_Sty_CD138_selected_plasma_cells_presentation


title: MM410_Nsp_CD138_selected_plasma_cells_presentation


title: MM420_Sty_CD138_selected_plasma_cells_presentation


title: MM418_Sty_CD138_selected_plasma_cells_presentation


title: MM409_Nsp_CD138_selected_plasma_cells_presentation


title: MM421_Sty_CD138_selected_plasma_cells_presentation


title: C0T2_D0-13

description: no additional information

hyb_protocol: Labelled cRNA was hybridized to Agilent mouse 4 x 44 oligonucleotide microarrays (Agilent Technologies) at 60ºC overnight. Arrays were washed according to the manufacturer’s specifications.

scan_protocol: Arrays were scanned on an Agilent G2505B scanner using default settings at 5mm, PMT 100%, extended dynamic range 0.1. Data were acquired using Agilent Feature Extraction software version 9.5.3.1.

data_processing:
The median signal intensity of each microarray sample was used. Technical replicates were not averaged.  Data for each probe was flagged (flag=1) if the median signal intensity for that probe was less than the mean of the negative controls ((-)3xSLv1) plus three standard deviations.
BAP_*.txt processed data files: RawCy5 and RawCy3 are the median signal intensities for the Cy5 and Cy3 channels on each microarray. NormCy5 and NormCy3 are the loess normalized signal intensities for the Cy5 and Cy3 channels within each array; this is a within-array normalization performed separately for each array. Flag indicates whether the median signal intensity falls in the background, i.e below threshold calculated from the negative controls (mean plus three standard deviations of the negative controls.) Ratio is calculated as NormCy5/NormCy3 (Relative Intensity, not logged.)
KJ_*.txt files are raw data files.

source_name: Total RNA, mouse liver

label_protocol: Double-stranded cDNA and cyanine labelled cRNA was synthesized according using Agilent Linear Amplification Kits. Experimental samples were labelled with Cyanine 5-CTP, and reference RNA with Cyanine 3-CTP (Perkin-Elmer Life Sciences). Cyanine-labelled cRNA targets were in vitro transcribed using T7 RNA polymerase and purified by RNeasy Mini Kit (Qiagen).

extract_protocol: RNA was isolated using the TRIzol reagent (Invitrogen Canada Inc., Burlington, ON, Canada), and further purified using RNeasy Mini Kits (Qiagen Inc., Mississauga, ON, Canada).  RNA was quantified using UV spectrophotometer, and quality was confirmed using the Agilent 2100 Bioanalyzer and RNA 6000 NanoLab Chip Kit (Agilent Technologies Canada Inc., Mississauga, ON, Canada).

tag: exposure

value: Control

tag: time post exposure

tag: microarray slide block

tag: microarray barcode

tag: gender

value: Male

tag: strain

value: Median liver lobe

source_name: universal mouse reference total RNA (Catalog #740100; Stratagene, La Jolla, CA, USA)

extract_protocol: Material was purchased as total RNA (universal mouse reference total RNA (Stratagene, CA, USA))

tag: reference

value: Total RNA derived from 11 mouse cell lines


title: MM411_Nsp_CD138_selected_plasma_cells_presentation


title: MM423_Sty_CD138_selected_plasma_cells_presentation


title: D2T2_D2-14

description: no additional information

value: 150 mg/kg Benzo(a)pyrene (CAS number 50-32-8)


title: D2T1_D2-7

description: no additional information


title: MM417_Sty_CD138_selected_plasma_cells_presentation


title: C0T1_D0-7

description: no additional information


title: MM415_Sty_CD138_selected_plasma_cells_presentation


title: MM407_Nsp_CD138_selected_plasma_cells_presentation


title: C0T2_D0-12

description: no additional information


title: MM403_Nsp_CD138_selected_plasma_cells_presentation


title: MM404_Nsp_CD138_selected_plasma_cells_presentation


title: Wild type embryos 2

city: Baltimore

first: Deborah

middle: J

last: Andrew

address: 725 N. Wolfe St., Hunterian G-1

department: Cell Biology

institute: Johns Hopkins University School of Medicine

description: Wild type embryos st. 11-17 isolated following passage through a COPAS biosorter

hyb_protocol: Samples and controls were prepared using the Affymetrix GeneChip hybridization, wash and stain kit (P/N 900720).  Arrays were prehybridized for 10 minutes prior to sample addition.  Following sample addition, arrays were hybridized for 16 hours at 45 Celsius, and shaken at 60 rpm.  Arrays were stained and washed using the Affymetrix GeneChip Fluidics Station 450 and FS450_002 fluidics script.

scan_protocol: Arrays were scanned with the Affymetrix GeneChip Scanner 3000 and raw analysis was performed with Affymetrix Command Console and Expression Console.

data_processing: Data was normalized by RMA and statistical analysis was performed using Partek software.  Further analysis was performed using Spotfire software.

organism: Drosophila melanogaster

source_name: St 11-15 embryos

label_protocol: Amplification and labeling were performed using the Affymetrix GeneChip 3' IVT Express kit (P/N 901229) following the manufacturer's protocol.

growth_protocol: Embryos were collected for eight hours and aged an additional 11 hours by the time of RNA extraction.

extract_protocol: RNA was isolated by TriZol extraction and cleaned up using the Qiagen Rneasy kit.

treatment_protocol: Embryos were sorted on a COPAS biosorter prior to RNA extraction.

value: embryo

tag: genotype/variation

value: wild type


title: D2T1_D2-5

description: no additional information


title: MM414_Sty_CD138_selected_plasma_cells_presentation


title: Wild type embryos 1


title: MM402_Nsp_CD138_selected_plasma_cells_presentation


title: MM406_Nsp_CD138_selected_plasma_cells_presentation


title: MM416_Sty_CD138_selected_plasma_cells_presentation


title: C0T1_D0-5

description: no additional information


title: MM411_Sty_CD138_selected_plasma_cells_presentation


title: MM413_Nsp_CD138_selected_plasma_cells_presentation


title: MM412_Sty_CD138_selected_plasma_cells_presentation


title: MM188_CD138_selected_plasma_cells_presentation

description: pre-treatment bone marrow plasma cells selected for CD138+ using microbeads and magnet-assisted cell sorting (MACS; Miltenyi Biotech, Bergisch Gladbach, Germany), according to the manufacturer's instructions.

hyb_protocol: Affymetrix Fluidics Station Protocol: Hybridization-EukGE-WS2v5_450, 15ug of fragmented cRNA were hybridised to U133 Plus 2.0 arrays using the standard Affymetrix conditions for 16 hours at 45C.

data_processing: The data were analyzed with dChip (April 2007 build) using default analysis settings for normalization and summarization methods.

label_protocol: 100 ng of RNA was used in the Affymetrix 2-cycle IVT reaction and cRNA was labelled with biointylated rUTP during the second IVT reaction as per Affymetrix protocol.

extract_protocol: Sample was CD138 positively selected using MACS beads and stored in RLT, extracted using Qiagen AllPrep DNA/RNA mini kit as per manufacturer's protocol.

value: myeloma cells

tag: storage

value: RLT


title: MM213_CD138_selected_plasma_cells_presentation

growth_protocol: 100 ng of RNA was used in the Affymetrix 2-cycle IVT reaction and cRNA was labelled with biointylated rUTP during the second IVT reaction as per Affymetrix protocol.


title: D2T2_D2-13

description: no additional information

tag: age


title: MM413_Sty_CD138_selected_plasma_cells_presentation


title: D2T2_D2-12

description: no additional information


title: C0T1_D0-6

description: no additional information


title: MM410_Sty_CD138_selected_plasma_cells_presentation


title: MM131_CD138_selected_plasma_cells_presentation


title: MM400_Nsp_CD138_selected_plasma_cells_presentation


title: MM405_Nsp_CD138_selected_plasma_cells_presentation


title: MM194_CD138_selected_plasma_cells_presentation


title: MM167_CD138_selected_plasma_cells_presentation


title: C0T2_D0-10

description: no additional information


title: Wild type embryos 3


title: MM408_Sty_CD138_selected_plasma_cells_presentation


title: D2T1_D2-6

description: no additional information


title: D2T2_D2-10

description: no additional information


title: MM408_Nsp_CD138_selected_plasma_cells_presentation


title: MM401_Nsp_CD138_selected_plasma_cells_presentation


title: D2T2_D2-9

description: no additional information


title: MM407_Sty_CD138_selected_plasma_cells_presentation


title: MM409_Sty_CD138_selected_plasma_cells_presentation


title: MM405_Sty_CD138_selected_plasma_cells_presentation


title: MM116_CD138_selected_plasma_cells_presentation


title: C0T1_D0-2

description: no additional information


title: D2T1_D2-2

description: no additional information


title: Creb3L1 sorted 3

description: Creb3L1 T/GFP expressing cells isolated by FACS using a FACSaria flow cytometer

source_name: HeLa cells; lipofectamine transfected with Creb3L1 T and GFP

growth_protocol: HeLa cells were grown at 37 C with 5%CO2 and humidity.  Confluent cultures were split 24 hours prior to transfection.  Following transfection, cells were grown in culture for 20 hours prior to removal for cell sorting.

extract_protocol: RNA was isolated using the Qiagen Rneasy kit.

treatment_protocol: Cells were transfected with 1 ug of DNA using the Fugene 6.0 lipofectamine reagent (Roche) 20 hours prior to sorting.  The amount of Creb3L1 T DNA was 0.8ug and DNA was 0.2ug.

tag: cell line

value: HeLa

tag: treatment


title: MM018_CD138_selected_plasma_cells_presentation


title: MM067_CD138_selected_plasma_cells_presentation


title: Wild type 3

description: Mock transfected HeLa cells that underwent sorting on a FACSaria flow cytometer

source_name: HeLa cells; lipofectamine only

value: lipofectamine only


title: MM014_CD138_selected_plasma_cells_presentation

value: AllPrep


title: Creb3L1 sorted 1

value: lipofectamine transfected with Creb3L1 T and GFP


title: C0T2_D0-9

description: no additional information


title: C0T1_D0-1

description: no additional information


title: MM016_CD138_selected_plasma_cells_presentation


title: MM406_Sty_CD138_selected_plasma_cells_presentation

value: pre-treatment bone marrow


title: D2T1_D2-1

description: no additional information


title: MM163_CD138_selected_plasma_cells_presentation


title: Creb3L1 sorted 2


title: MM404_Sty_CD138_selected_plasma_cells_presentation


title: Wild type 1


title: MM403_Sty_CD138_selected_plasma_cells_presentation


title: MM402_Sty_CD138_selected_plasma_cells_presentation


title: Fibroblasts infected with active virus, 6h timepoint, dendrimer labeling, replicate 2

status: Public on Oct 16 2010

first: Falko

last: Schmeisser

address: 8800 Rockville Pike, building 29A, room 3C-05

department: DVP

institute: FDA/CBER

hyb_protocol: A two-step hybridization procedure was initiated with the paired, concentrated cDNAs. The first hybridization was performed in the Array 350™ formamide-based hybridization buffer to the oligo array slides in HybChambers™ (GenomicSolutions) under 25 x 40 mm LifterSlip coverslips (Erie Scientific) for 18 h at 42° C. After washing the slides, the second hybridization was performed in HybChambers™ under 25 x 40 mm LifterSlip coverslips with the fluorescent 3DNA Cy3 and Cy5 reagents in the formamide-based hybridization buffer for 3 h at 42° C. Washes were performed in between the hybridization steps and after the second hybridization at 45° C, 15 min each wash, in the following order:  2X SSC with 0.2% SDS; 2X SSC; 0.2X SSC. After the first hybridzation only, the slides were immersed in 95% ethanol for 2 minutes after the third wash. Slides were dried by centrifugation at 200 x g in a RT7 Plus tabletop centrifuge (Sorvall) for 2 minutes at room temperature.

scan_protocol: For image acquisition, an Axon GenePix 4000B scanner (Molecular Devices) was utilized, with power settings fixed at 100%, PMT gain variable for Cy3 and Cy5 channels (500-800). Microarray slides were scanned at 10-μm resolution at 532 nm (Cy3) and 635 nm (Cy5) wavelengths using GenePix Pro software (MolecularDevices).

data_processing: Scanned microarray images were aquired as tif files for data analysis using the corresponding GAL file for block alignment, block mask corrections and spot evaluation. The quantification of spot intensities, qualities, and local background was performed automatically by Genepix software using variable spot diameter in the range 70–180 µm and manual supervision and correction for any inaccuracies in the automatic spot detection. Raw images were collected at 16-bit/pixel resolutions that displayed all pixels in a 0 to 65 535 count dynamic range. The area surrounding each spot image was used to calculate a local background and subtracted from each spot before Cy5/Cy3 ratio calculation. The average of the resulting total Cy3 and Cy5 signal gave a ratio that was used to normalize the signals. Each microarray experiment was globally normalized to make the median value of the log-2 ratio equal to zero. The normalization process corrects for dye bias, PMT (photo multiplier tube) voltage imbalance, and variations between channels in the amounts of the labeled cDNA probes hybridized. Data files in gpr format and images in jpg format were imported into the microarray database (mAdb), and analyzed using software tools provided by the National Institutes of Health, Center for Information Technology (http://nciarray.nci.nih.gov). Spots that were indicated as bad or not found were excluded. Only spots with a fluorescence intensity of at least 500 in both channels and having 25% target pixels at least one standard deviation above background were considered as good spots for analysis. Spot diameter was limited to the range between 60 and 300 µm. Matrix Data files were then created using a greater than or equal to threefold signal intensity change criteria. Only spots with a confidence interval of 99% (> 3- fold),were considered good spots for analysis. Genes were required to have values in two out of three experiments analyzed.

source_name: Fibroblasts, uninfected

label_protocol: The Array 350™ Cy3 Kit and the Array 350™ Cy5 Kit (Genisphere) were used to label the cDNA made from the respective RNA: The kits also contained reagents necessary for hybridization. Dye-swap experiments (reverse labeling using independent biological replicates) were performed twice. Each cDNA made from control and virus-infected cell RNA was prepared in a reaction containing 5 μg of total RNA, respective capture-dendrimer primer, dNTPs (kit provided), and 100 U of StrataScript RT in 1X StrataScript buffer (Stratagene) at 42°C for 2 h. Following reaction termination, denaturation at 65°C, and neutralization, the reactions containing the control cDNA and cDNA produced with RNA isolated from infected cells were combined just prior to concentration through a Microcon YM-30 (Millipore) centrifugal filter unit.

growth_protocol: Detroit 551 fibroblasts were cultured in Eagle's Minimum Essential Medium (MEM) supplemented with fetal bovine serum to a final concentration of 10%, as recommended by the supplier. For subculturing, medium was removed and the cell layer rinsed with 0.25% trypsin, 0.53 mM EDTA solution. An additional 1 to 2 ml of trypsin-EDTA solution was added, and the culture vessel (flask) was allowed to sit at t 37°C until the cells detach. Fresh culture medium was added and the appropriate amount of cells was dispensed into new culture flasks. Subcultivation was performed at a ratio of 1:2 to 1:5, medium was renewed 2 to 3 times per week. The cells have a finite lifespan of about 25 serial passages from the tissue of origin. Cells were at the same passage number at time of treatment for each of the three independent biological replicates.

extract_protocol: Total RNA extracted using Trizol (Invitrogen) following manufacturer's instructions. RNA pellets were resuspended in deionized water containing 2% SDS and further processed using RNeasy spin columns (Qiagen), including on-column DNase treatment with RNase-free Dnase (Qiagen) as described by the supplier. Quality of RNA was confirmed using an Agilent 2100 Bioanalyzer (Agilent Technologies).

treatment_protocol: Detroit 551 fibroblasts were either cultured uninfected, or were infected with Herpes Simplex Virus 1 (HSV-1) at a multiplicity of infection (moi) of 10. Samples were harvested at 6 hours post infection.

value: Detroit 551

tag: cell line description

value: Detroit 551 human foreskin fibroblasts   ATCC Number CCL-110,

tag: cell line reference

value: Sugarman BJ, et al. (1985) Recombinant human tumor necrosis factor-alpha:effects on proliferation of normal and transformed cells in vitro. Science 230:943-945. PubMed:3933111

tag: infection

value: uninfected

tag: virus description

value: no virus

source_name: Fibroblasts, infected with HSV-1, F-strain

value: infected with HSV-1 (F) at a multiplicity of infection (moi) = 10

value: Herpes Simplex Virus serotype 1, F-strain, ATCC Number VR-733


title: Fibroblasts infected with active virus, 6h timepoint, dendrimer labeling, replicate 3


title: Fibroblasts infected with active virus, 6h timepoint, dendrimer labeling, replicate 1


title: NTNeurons infected with active virus, 24h timepoint, aminoallyl labeling, replicate 3

hyb_protocol: Arrays were prehybridized with 50 µl prehybridization buffer (25 µl 20x SSC, 20 µl 5% BSA, 54 µl water, and 1 µl 10% SDS) under a coverslip (22 mm x 40 mm) for 1 hour at 42°C, washed with water and isopropanol (2 minutes each), and spin-dried. For hybridization, 34 µl hybridization mixture (24 µl cDNA mixture, 1 µl [10 µg] COT-1 DNA, 1 µl [8-10 µg] poly(dA), 1 µl [4 µg] yeast tRNA, 6 µl 20x SSC, and 1 µl 10% SDS) was preheated to 100°C for 2 minutes and quickly spun down for 1 minute . The entire volume of probe was added onto prehybridized array and covered with coverslip. Slides were placed in HybChamber hybridization chambers and incubated at 65°C in a water bath overnight (10-16 hours). Then, slides were washed for 2 minutes each in 2x SSC, 1x SSC, and 0.2x SSC. Slides were dried by centrifugation at 200 x g in a RT7 Plus tabletop centrifuge (Sorvall) for 2 minutes at room temperature.

data_processing: Scanned microarray images were aquired as tif files for data analysis using the corresponding GAL file for block alignment, block mask corrections and spot evaluation. The quantification of spot intensities, qualities, and local background was performed automatically by Genepix software using variable spot diameter in the range 70–180 µm and manual supervision and correction for any inaccuracies in the automatic spot detection. Raw images were collected at 16-bit/pixel resolutions that displayed all pixels in a 0 to 65 535 count dynamic range. The area surrounding each spot image was used to calculate a local background and subtracted from each spot before Cy5/Cy3 ratio calculation. The average of the resulting total Cy3 and Cy5 signal gave a ratio that was used to normalize the signals. Each microarray experiment was globally normalized to make the median value of the log-2 ratio equal to zero. The normalization process corrects for dye bias, PMT (photo multiplier tube) voltage imbalance, and variations between channels in the amounts of the labeled cDNA probes hybridized. Data files in gpr format and images in jpg format were imported into the microarray database (mAdb), and analyzed using software tools provided by the National Institutes of Health, Center for Information Technology (http://nciarray.nci.nih.gov). Spots that were indicated as bad or not found were excluded. Only spots with a fluorescence intensity of at least 500 in both channels and having 25% target pixels at least one standard deviation above background were considered as good spots for analysis. Spot diameter was limited to the range between 60 and 300 µm. Matrix Data files were then created using a greater than or equal to threefold signal intensity change criteria. Only spots with a confidence interval of 99% (> 3- fold),were considered good spots for analysis. Genes were required to have values in three out of four experiments analyzed.

source_name: NTNeurons, uninfected

label_protocol: 20 µg total RNA was incubated at 70°C for 5 minutes along with 1 µl oligo dT and quickly chilled for 3 minutes. Then, 3 µl of 10x first-strand buffer, 2 µl StratascriptRT enzyme (Stratagene), 2 µl of 20x aa-dUTP/dNTP (containing 10 mM of dATP, dCTP, and dGTP, 6 mM of dTTP, and 4 mM of aminoallyl-dUTP), and 3 µl of 0.1 M dithiothreitol (DTT) were added and incubated for 90 minutes at 42°C for reverse transcription. After incubation, the volume of the mixture was increased to 50 µl with 20 µl water. cDNA was purified using MinElute columns (Qiagen). After washing, the probe was eluted with 15 µl elution buffer, spun for 1 minute, and dried by speed-vac for 14 to 15 minutes, avoiding overdrying the probe. Finally, 5 µl of 2x coupling buffer and 5 µl Cy3 and Cy5 dye were mixed with the respective cDNAs, and incubated at room temperature in the dark for 1 hour. After incubation, the volume was raised to 50 µl with water. The cDNA was purified on MinElute column once again, eluted with 12 µl elution buffer, centrifuged to collect the cDNA probes, and then both probes were combined.

growth_protocol: Undifferentiated NT2 cells were passaged at relatively high density (>5x10^6 per 75 cm^2 flask) in Opti-MEM (Gibco/Life Technologies, Gaithersburg, MD, USA) containing 10% fetal bovine serum. For differentiation, 5x10^6 cells (162 cm^2 flask) were treated for 5 weeks with retinoic acid (0.01 mM), then replated at a 1:5 dilution and treated with mitotic inhibitors. After 10 days, the NTNeurons were mechanically dislodged and replated on poly-L-lysine-coated dishes, wells, or slides treated with MATRIGEL (Collaborative Biomedical, Bedford, MA, USA) and cultured in conditioned DMEM containing fetal bovine serum without retinoic acid or mitotic inhibitors.

treatment_protocol: NTNeurons were either cultured uninfected, or were infected with Herpes Simplex Virus 1 (HSV-1) at a multiplicity of infection (moi) of 10. Samples were harvested at 24 hours post infection.

value: NTERA-2 cl.D1 [NT2/D1] ATCC Number CRL-1973, embryonal carcinoma cell line derived from a human teratocarcinoma

tag: cells line differentiation

value: irreversibly differentiated into NTNeurons in the presence of retinoic acid.

value: Andrews PW (1984) Retinoic acid induces neuronal differentiation of a cloned human embryonal carcinoma cell line in vitro. Dev Biol 103:285-293

source_name: NTNeurons, infected with HSV-1, F-strain


title: Alveolar Rhabdomyosarcoma tumor PAIZIY 50K_Xba

city: Philadelphia

first: Frederic

last: Barr

state: PA

address: 422 Curie Blvd

institute: University of Pennsylvania

description: Hybridized to 50K_Xba

hyb_protocol: DNA was digested with the appropriate restriction enzyme, PCR amplified, fragmented, labeled and hybridized to each array according to the manufacturer's instructions.

scan_protocol: The hybridized arrays were washed using Affymetrix fluidics stations, and scanned using the GeneChip Scanner 3000.

data_processing:
CEL files and TXT files were generated by Affymetrix GeneChip DNA Analysis Software version 3.0, Affymetrix Copy Number Analysis Tool (CNAT) versions 2.2 and 3.0, Affymetrix GeneChip Genotyping Operating Analysis software version 4.1 and Partek Genomics Suite version 6.3.  The 50K data were calculated in CNAT using an algorithm developed to identify genome wide gains and losses on high density oligonucleotide arrays based on SNP genotyping metods and whole genome sampling analysis.  The raw data were analyzed with CNAT to calculate a copy number at each locus, which was then normalized relative to a reference set of normal individuals.  The copy number estimates were displayed with a genome smoothed analysis, which incorporates Gaussian kernel smoothing over the default genomic filter length of 0.5 Mb.  Data from the 250K array were analyzed with the Hidden Markov Model default settings in the Copy Number work flow in Partek Genomics Suite.  The respective Hap Map samples were used as normals for both the 50K and 250K chips.
The 50K CNAT data contains probe set id, physical chromosome position, genotype call (SNP call) AA, AB, BB, NC no call, GSA_CN genomic smoothed analysis of the copy number, GSA_pVal genomic smoothed analysis of the significance value and Log2 Ratio.  The 250K data .txt files analyzed on Partek contain the probe set ID and the copy number generated using the Copy Number workflow in Partek Genomic Suite.

label: Biotin

source_name: Alveolar Rhabdomyosarcoma tumor PAIZIY

label_protocol: As per manufacturer (Affymetrix)

extract_protocol: Genomic DNA was extracted from fresh frozen tumors using the Qiagen QIAamp DNA Mini Kit. DNA quality and quantity was assessed using a Nanodrop Spectrophotometer and agarose gel electrophoresis.

tag: tissue

value: Alveolar rhabdomyosarcoma tumor

tag: case

value: PAIZIY


title: MM401_Sty_CD138_selected_plasma_cells_presentation

tag: extraction

tag: cell type


title: Alveolar Rhabdomyosarcoma tumor PAMJZD 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAMJZD

value: PAMJZD


title: Alveolar Rhabdomyosarcoma tumor PALENF 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALENF

value: PALENF


title: Alveolar Rhabdomyosarcoma tumor PAMEHR 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAMEHR

value: PAMEHR


title: NTNeurons infected with active virus, 24h timepoint, aminoallyl labeling, replicate 4


title: Alveolar Rhabdomyosarcoma tumor PALADB 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALADB

value: PALADB


title: NTNeurons infected with active virus, 24h timepoint, aminoallyl labeling, replicate 2


title: Alveolar Rhabdomyosarcoma tumor PALTNM 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALTNM

value: PALTNM


title: Alveolar Rhabdomyosarcoma tumor PALKXV 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALKXV

value: PALKXV


title: Alveolar Rhabdomyosarcoma tumor PALDIF 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALDIF

value: PALDIF


title: Wild type 2

value: HeLa

value: lipofectamine only


title: Alveolar Rhabdomyosarcoma tumor PALIRD 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALIRD

value: PALIRD


title: Alveolar Rhabdomyosarcoma tumor PAKIET 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKIET

value: PAKIET


title: Alveolar Rhabdomyosarcoma tumor PALKKG 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALKKG

value: PALKKG


title: Alveolar Rhabdomyosarcoma tumor PALITD 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALITD

value: PALITD


title: Alveolar Rhabdomyosarcoma tumor PAKJZV 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKJZV

value: PAKJZV


title: Alveolar Rhabdomyosarcoma tumor PAKIPK 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKIPK

value: PAKIPK


title: Alveolar Rhabdomyosarcoma tumor PALJPA 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALJPA

value: Alveolar rhabdomyosarcoma tumor

value: PALJPA


title: NTNeurons infected with active virus, 24h timepoint, aminoallyl labeling, replicate 1

source_name: NTNeurons, uninfected

value: NTERA-2 cl.D1 [NT2/D1] ATCC Number CRL-1973, embryonal carcinoma cell line derived from a human teratocarcinoma

value: irreversibly differentiated into NTNeurons in the presence of retinoic acid.

value: Andrews PW (1984) Retinoic acid induces neuronal differentiation of a cloned human embryonal carcinoma cell line in vitro. Dev Biol 103:285-293

value: uninfected

value: no virus

value: infected with HSV-1 (F) at a multiplicity of infection (moi) = 10

value: Herpes Simplex Virus serotype 1, F-strain, ATCC Number VR-733


title: Alveolar Rhabdomyosarcoma tumor PALFJB 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALFJB

value: PALFJB


title: Alveolar Rhabdomyosarcoma tumor PAKJHG 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKJHG

value: PAKJHG


title: Alveolar Rhabdomyosarcoma tumor PAKREB 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKREB

value: PAKREB


title: Alveolar Rhabdomyosarcoma tumor PAJALC  50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAJALC

value: PAJALC


title: NTNeurons infected with active virus, 6h timepoint, aminoallyl labeling, replicate 3

treatment_protocol: NTNeurons were either cultured uninfected, or were infected with Herpes Simplex Virus 1 (HSV-1) at a multiplicity of infection (moi) of 10. Samples were harvested at 6 hours post infection.

source_name: NTNeurons, infected with HSV-1, F-strain


title: MM400_Sty_CD138_selected_plasma_cells_presentation

source_name: pre-treatment bone marrow

value: AllPrep

value: pre-treatment bone marrow


title: NTNeurons infected with inactivated virus, 24h timepoint, dendrimer labeling, replicate 4

treatment_protocol: NTNeurons were either cultured uninfected, or were infected with inactivated Herpes Simplex Virus 1 (HSV-1) at a multiplicity of infection (moi) of 10. Virus was inactivated by psoralen treatment for 10 min at RT followed by UV-irradiation for 15 min. Briefly, Trioxsalen was added to a final concentration of 1 μg/ml, and the virus suspension was left at room temperature for 10 min. The Trioxsalen-treated virus suspension was transferred into 100-mm-diameter petri dishes (7–10 ml per dish), and exposed to ultraviolet light for 15 min using a Stratalinker-1800 (Stratagene.). The titer of an aliquot of untreated virus suspension was used to estimate the pfu-equivalent of the inactivated virus suspension. Inactivated virus preps were titered on Vero cells to confirm that no active virus remained in the preparation. Samples were harvested at 24 hours post infection.

source_name: NTNeurons, infected with inactivated HSV-1, F-strain

value: infected with inactivated HSV-1 (F) at a multiplicity of infection (moi) = 10

value: Herpes Simplex Virus serotype 1, F-strain, ATCC Number VR-733, inactivated by a combination of psoralen and UV treatment


title: NTNeurons infected with active virus, 6h timepoint, aminoallyl labeling, replicate 2


title: NTNeurons infected with active virus, 6h timepoint, aminoallyl labeling, replicate 1


title: NTNeurons infected with inactivated virus, 24h timepoint, dendrimer labeling, replicate 3


title: NTNeurons infected with inactivated virus, 24h timepoint, dendrimer labeling, replicate 2


title: NTNeurons infected with inactivated virus, 24h timepoint, dendrimer labeling, replicate 1


title: Alveolar Rhabdomyosarcoma tumor PALKPZ 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALKPZ

value: Alveolar rhabdomyosarcoma tumor

value: PALKPZ


title: NTNeurons infected with inactivated virus, 6h timepoint, dendrimer labeling, replicate 4

treatment_protocol: NTNeurons were either cultured uninfected, or were infected with inactivated Herpes Simplex Virus 1 (HSV-1) at a multiplicity of infection (moi) of 10. Virus was inactivated by psoralen treatment for 10 min at RT followed by UV-irradiation for 15 min. Briefly, Trioxsalen was added to a final concentration of 1 μg/ml, and the virus suspension was left at room temperature for 10 min. The Trioxsalen-treated virus suspension was transferred into 100-mm-diameter petri dishes (7–10 ml per dish), and exposed to ultraviolet light for 15 min using a Stratalinker-1800 (Stratagene.). The titer of an aliquot of untreated virus suspension was used to estimate the pfu-equivalent of the inactivated virus suspension. Inactivated virus preps were titered on Vero cells to confirm that no active virus remained in the preparation. Samples were harvested at 6 hours post infection.

value: NTERA-2 cl.D1 [NT2/D1] ATCC Number CRL-1973, embryonal carcinoma cell line derived from a human teratocarcinoma


title: Alveolar Rhabdomyosarcoma tumor PALKVA  50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALKVA

value: PALKVA


title: NTNeurons infected with inactivated virus, 6h timepoint, dendrimer labeling, replicate 1

value: irreversibly differentiated into NTNeurons in the presence of retinoic acid.

value: Andrews PW (1984) Retinoic acid induces neuronal differentiation of a cloned human embryonal carcinoma cell line in vitro. Dev Biol 103:285-293


title: NTNeurons infected with inactivated virus, 6h timepoint, dendrimer labeling, replicate 2

value: uninfected

value: no virus


title: NTNeurons infected with active virus, 24h timepoint, dendrimer labeling, replicate 3

description: reverse-label experiment, not a true dye-flip because samples used are biologically independent

value: infected with HSV-1 (F) at a multiplicity of infection (moi) = 10

value: Herpes Simplex Virus serotype 1, F-strain, ATCC Number VR-733


title: NTNeurons infected with active virus, 24h timepoint, dendrimer labeling, replicate 4


title: NTNeurons infected with inactivated virus, 6h timepoint, dendrimer labeling, replicate 3


title: Alveolar Rhabdomyosarcoma tumor PAKCBU 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKCBU

value: PAKCBU


title: Alveolar Rhabdomyosarcoma tumor PALMGG 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PALMGG

value: PALMGG


title: Alveolar Rhabdomyosarcoma tumor PAJAEG 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAJAEG

value: PAJAEG


title: Alveolar Rhabdomyosarcoma tumor PAJAFT 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAJAFT

value: PAJAFT


title: Alveolar Rhabdomyosarcoma tumor PAJADP 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAJADP

value: PAJADP


title: Alveolar Rhabdomyosarcoma tumor PAKDIF 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKDIF

value: PAKDIF


title: Alveolar Rhabdomyosarcoma tumor PAKBEN 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKBEN

value: PAKBEN


title: Alveolar Rhabdomyosarcoma tumor PAJAAP 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAJAAP

value: Alveolar rhabdomyosarcoma tumor

value: PAJAAP


title: Alveolar Rhabdomyosarcoma tumor PAIZZP 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZZP

value: PAIZZP


title: NTNeurons infected with active virus, 6h timepoint, dendrimer labeling, replicate 3

value: NTERA-2 cl.D1 [NT2/D1] ATCC Number CRL-1973, embryonal carcinoma cell line derived from a human teratocarcinoma

source_name: NTNeurons, uninfected


title: NTNeurons infected with active virus, 6h timepoint, dendrimer labeling, replicate 4

value: irreversibly differentiated into NTNeurons in the presence of retinoic acid.


title: NTNeurons infected with active virus, 6h timepoint, dendrimer labeling, replicate 1

value: Andrews PW (1984) Retinoic acid induces neuronal differentiation of a cloned human embryonal carcinoma cell line in vitro. Dev Biol 103:285-293


title: NTNeurons infected with active virus, 24h timepoint, dendrimer labeling, replicate 2

value: uninfected


title: NTNeurons infected with active virus, 24h timepoint, dendrimer labeling, replicate 1

value: no virus


title: Alveolar Rhabdomyosarcoma tumor PAIZZG 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZZG

value: PAIZZG


title: Alveolar Rhabdomyosarcoma tumor PAIZYP 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZYP

value: PAIZYP


title: Alveolar Rhabdomyosarcoma tumor PAIZWY 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZWY

value: PAIZWY


title: Alveolar Rhabdomyosarcoma tumor PAIZXF 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZXF

value: PAIZXF


title: Alveolar Rhabdomyosarcoma tumor PAIZUT 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZUT

value: PAIZUT


title: 2G1_Normal

first: quincy

last: mo

address: 1275 York Avenue

institute: Memorial Sloan-Kettering Cancer Center

description: Gene expression data from normal bone marrow sample.

hyb_protocol: Ten micrograms of labeled and fragmented cRNA were hybridized to the Human Genome U133 plus 2.0 array (Affymetrix) at 45°C for 16 h.

scan_protocol: Chips were scanned with a high-numerical Aperture and flying objective (FOL) lens in the GS3000 scanner (Affymetrix).

data_processing: The microarray data were quantile-normalized and the gene expression values were estimated using the RMA method. R 2.7.0 and package affy was used to process the data.

source_name: Normal CD34+ G1/S/G2/M cells

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 20ng total RNA using the GeneChip two-cycle target labeling kit (Affymetrix, Inc., Santa Clara, CA, U.S.)

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

value: bone marrow

tag: cell cycle phase

tag: disease state

value: normal


title: Alveolar Rhabdomyosarcoma tumor PAIZRA 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZRA

value: PAIZRA


title: 8G1_CML

description: Gene expression data from chronic myelogenous leukemia patient.

source_name: CML CD34+ G1/S/G2/M cells

value: peripheral blood

value: chronic myelogenous leukemia (CML)

tag: cml phase

value: chronic


title: 4G1_Normal


title: Alveolar Rhabdomyosarcoma tumor PAIZKG 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZKG

value: Alveolar rhabdomyosarcoma tumor

value: PAIZKG


title: Alveolar Rhabdomyosarcoma tumor PAIZPC 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZPC

value: PAIZPC


title: 6G1_CML

value: accelerated


title: Alveolar Rhabdomyosarcoma tumor PAIZNC 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZNC

value: PAIZNC


title: NTNeurons infected with active virus, 6h timepoint, dendrimer labeling, replicate 2

source_name: NTNeurons, infected with HSV-1, F-strain

value: NTERA-2 cl.D1 [NT2/D1] ATCC Number CRL-1973, embryonal carcinoma cell line derived from a human teratocarcinoma

value: irreversibly differentiated into NTNeurons in the presence of retinoic acid.

value: Andrews PW (1984) Retinoic acid induces neuronal differentiation of a cloned human embryonal carcinoma cell line in vitro. Dev Biol 103:285-293

value: infected with HSV-1 (F) at a multiplicity of infection (moi) = 10

value: Herpes Simplex Virus serotype 1, F-strain, ATCC Number VR-733

value: uninfected

value: no virus


title: 5G1_CML


title: Alveolar Rhabdomyosarcoma tumor PAIZRN 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZRN

value: PAIZRN


title: 3G1_Normal

value: bone marrow

value: normal


title: Alveolar Rhabdomyosarcoma tumor PAIZGD 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZGD

value: PAIZGD


title: Alveolar Rhabdomyosarcoma tumor PAIZNH 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZNH

value: PAIZNH


title: 7G1_CML

value: peripheral blood

value: chronic myelogenous leukemia (CML)

value: chronic


title: Alveolar Rhabdomyosarcoma tumor PAIZJI 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZJI

value: Alveolar rhabdomyosarcoma tumor

value: PAIZJI


title: 1G1_Normal


title: 4G1_CML


title: reconSCID, biological rep1

city: Boston

first: CBDM

last: Lab

state: MA

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

description: Gene expression data of MECs isolated from BM reconstituted SCID-NOD mouse

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

scan_protocol: GeneChips were scanned using  GeneArray Scanner.

data_processing: The data were normalized and analyzed with Genepattern software.

source_name: reconSCID

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

growth_protocol: Mice were reconstituted by BM transfer (IP injection into 2-4d pups) 6 hrs post-irradiation (200rad). Reconstituted thymi were analyzed 6-8w after transfer.

treatment_protocol: MECs were isolated from thymi by standard Collagenase Dispase protocol and sorted into Trizol on MoFlo instrument

tag: strain

value: NOD strain with SCID (DNAPK mutation)

value: MHCIIhi MECs


title: 3G1_CML

value: accelerated


title: reconSCID, biological rep2


title: Alveolar Rhabdomyosarcoma tumor PAIZAI 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIZAI

value: PAIZAI


title: reconRAG1KO, biological rep4

description: Gene expression data of MECs isolated from BM reconstituted RAG-NOD mouse

value: NOD strain with null  RAG1


title: Alveolar Rhabdomyosarcoma tumor PAIYYV 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYYV

value: PAIYYV


title: 1G1_CML


title: reconSCID, biological rep3


title: Alveolar Rhabdomyosarcoma tumor PAIYYE 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYYE

value: PAIYYE


title: Alveolar Rhabdomyosarcoma tumor PAKZRT 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAKZRT

value: PAKZRT


title: 2G1_CML


title: 3G0_Normal

source_name: Normal CD34+ G0 cells

value: bone marrow


title: 2G0_Normal

value: normal


title: Alveolar Rhabdomyosarcoma tumor PAIYSN 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYSN

value: PAIYSN


title: 1G0_Normal


title: Alveolar Rhabdomyosarcoma tumor PAIYWS 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYWS

value: PAIYWS


title: 8G0_CML

source_name: CML CD34+ G0 cells

value: peripheral blood


title: 7G0_CML

value: chronic myelogenous leukemia (CML)


title: 5G0_CML


title: reconRAG1KO, biological rep3


title: reconRAG1KO, biological rep2


title: Alveolar Rhabdomyosarcoma tumor PAIYXX 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYXX

value: PAIYXX


title: Alveolar Rhabdomyosarcoma tumor PAIYRA 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYRA

value: Alveolar rhabdomyosarcoma tumor

value: PAIYRA


title: 4G0_CML

value: chronic


title: 6G0_CML


title: 2G0_CML


title: 3G0_CML


title: Alveolar Rhabdomyosarcoma tumor PAIYRA sty 250K

description: Hybridized to 250K_Sty


title: 4G0_Normal


title: reconRAG1KO, biological rep1

value: MHCIIhi MECs


title: HD837-T/T

city: New Haven

first: Laurence

last: Menard

state: CT

address: 300 George st

department: Immunobiology-HTI

institute: Yale School of Medicine

description: Gene expression data from CD19+CD10-CD21+CD27- conventional mature naive B cells from T/T patients (carriers of 2 alleles of the polymorphism).

hyb_protocol: Labeled cDNA was hybridized on chips containing the whole human genome (Human Genome U133 Plus 2.0 from Affymetrix).

scan_protocol: GeneChip Scanner 3000 7G using Affymetrix GeneChip Operating Software. The target signal intensity from each chip was scaled to 500.

data_processing: The data normalization and statistical analysis were performed with GeneSpring GX7 software (Agilent). We used the per-chip and per-gene normalization method.

source_name: CD19+CD10-CD21+CD27- conventional mature naive B cells, T/T alleles

label_protocol: Using the Ovation biotin system kit from Nugen, 30-50ng of RNA was amplified and labeled to produce cDNA.

growth_protocol: CD19+CD10-CD21+CD27- conventional mature naive B cells were batch sorted by FACS.

extract_protocol: RNA was extracted from batch-sorted CD19+CD10-CD21+CD27- conventional mature naive B cells using the Absolutely RNA microprep kit (Stratagene).  100-200 ng of RNA was obtained per sample, and the quality of the purified RNA was assessed by the Bioanalyzer from Agilent.

tag: ptpn22 alleles

value: T/T


title: HD862-C/T

description: Gene expression data from CD19+CD10-CD21+CD27- conventional mature naive B cells from C/T patients (carriers of 1 allele of the polymorphism).

source_name: CD19+CD10-CD21+CD27- conventional mature naive B cells, C/T alleles

value: C/T


title: Alveolar Rhabdomyosarcoma tumor PAIYHS 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYHS

value: PAIYHS


title: HD13-C/T

value: CD19+CD10-CD21+CD27- conventional mature naive B cell


title: HD535-T/T


title: HD138-T/T


title: Alveolar Rhabdomyosarcoma tumor PAIYHS 250K_Sty

description: Hybridized to 250K_Sty


title: HD-CT2-C/T


title: Alveolar Rhabdomyosarcoma tumor PALKKG 250K_Sty

description: Hybridized to 250K_Sty

source_name: Alveolar Rhabdomyosarcoma tumor PALKKG

value: PALKKG


title: Alveolar Rhabdomyosarcoma tumor PAIZJI 250K_Sty

description: Hybridized to 250K_Sty

value: PAIZJI


title: 1G0_CML


title: HD12-C/T


title: Alveolar Rhabdomyosarcoma tumor PAMJZD 250K_Sty

description: Hybridized to 250K_Sty

source_name: Alveolar Rhabdomyosarcoma tumor PAMJZD

value: PAMJZD


title: Alveolar Rhabdomyosarcoma tumor PAIYHC 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYHC

value: PAIYHC


title: Alveolar Rhabdomyosarcoma tumor PAIYMI 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYMI

value: PAIYMI


title: 4302_CD133-, laminin treated

city: Durham

first: Igor

last: Shats

state: NC

address: 101 Science Drive

institute: Duke University

description: Gene expression data from breast cancer cell lines

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Human U133A plus 2.0 arrays. Affymetrix GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Affymetrix Scanner 3000 7G.

data_processing: The data were analyzed with RMA (Robust Multichip Average) using Bioconductor

source_name: Cultured cell line

label_protocol: Biotinylated cRNA were prepared using Ambion MessageAmp Premier labeling kit.

growth_protocol: CD133+ and CD133- glioma cells were separated from tumor xenografts (4105 and 4302), cultured in serum-free media for 48 hours in the presence of absence of laminin, which led to adherent growth of glioma stem cells while preserving stemness.

extract_protocol: RNeasy mini kit (Qiagen)

treatment_protocol: No treatment

value: glioblastoma xenografts


title: HD1400-C/C

description: Gene expression data from CD19+CD10-CD21+CD27- conventional mature naive B cells from C/C patients (non-carriers of the polymorphism).

source_name: CD19+CD10-CD21+CD27- conventional mature naive B cells, C/C alleles

value: C/C


title: Alveolar Rhabdomyosarcoma tumor PAIYPB 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYPB

value: PAIYPB


title: 4302_CD133+, laminin treated


title: HD750-C/T


title: Alveolar Rhabdomyosarcoma tumor PAIYGR 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIYGR

value: PAIYGR


title: Alveolar Rhabdomyosarcoma tumor PAIXXY 50K_Xba

description: Hybridized to 50K_Xba

source_name: Alveolar Rhabdomyosarcoma tumor PAIXXY

value: Alveolar rhabdomyosarcoma tumor

value: PAIXXY


title: HD10-C/C


title: HD1424-C/C


title: ko_4

city: Los Angeles

first: Giovanni

last: Coppola

state: CA

address: 1524 Gonda, 695 Charles Young Drive South

department: Psychiatry and Neurology

institute: UCLA

hyb_protocol: Illumina protocol

scan_protocol: Illumina protocol

data_processing: BeadStudio, quantile normalization

source_name: E12.5 DRG RNA from Erk1/2CKO(Wnt1) mice

label_protocol: Biotin

extract_protocol: Total RNA was extracted with Trizol (Invitrogen) and a Qiagen RNeasy Mini kit per manufacturer’s instructions.

treatment_protocol: E12.5 DRGs from control and Erk1/2CKO(Wnt1) embryos derived from three independent litters were dissected in PBS supplemented with 10% RNAlater (Qiagen) and frozen on dry ice.

tag: genotype/variation

value: E12.5 Erk1/2CKO(Wnt1)

value: E12.5 dorsal root ganglia (DRG)


title: HD1425-C/C


title: HD06-C/C


title: HD16-C/C


title: Alveolar Rhabdomyosarcoma tumor PAIZNC 250K_Sty

description: Hybridized to 250K_Sty

source_name: Alveolar Rhabdomyosarcoma tumor PAIZNC

value: PAIZNC


title: ko_2


title: HD07-C/C


title: 4105_CD133+, laminin treated


title: Ctr1

description: 293T cells transfected with pCMV empty expression vector

growth_protocol: 293T cells were grown on 10cm plates in DMEM containing 10% FBS under standard TC conditions

extract_protocol: Total RNA was extracted 48 hrs post transfection by Trizol extraction  according to the manufacturer's instructions.

treatment_protocol: 293T cells were transfected either by pCMV-Aire (6ug/plate) or empty pCMV (6ug/plate) using Mirus reagent, according to manufacturer's instructions


title: Alveolar Rhabdomyosarcoma tumor PAIZXF 250K_Sty

description: Hybridized to 250K_Sty

source_name: Alveolar Rhabdomyosarcoma tumor PAIZXF

value: PAIZXF


title: 4105_CD133-, laminin treated


title: 4302_CD133+, untreated


title: Ctr2

value: 293T cells transfected with pCMV empty expression vector


title: wt_3

source_name: E12.5 DRG RNA from WT mice

value: wild-type


title: Aire3

description: 293T cells transfected with pCMV-Aire expression vector


title: HD17-C/C

value: CD19+CD10-CD21+CD27- conventional mature naive B cell


title: Aire2

value: 293T cells transfected with pCMV-Aire expression vector


title: 4105_CD133+, untreated


title: TTHA0167-deficient strain_480_2

status: Public on Oct 15 2010

city: Hyogo

first: Akeo

last: Shinkai

address: 1-1-1, Kouto, Sayo

department: SPring-8 Center

country: Japan

institute: RIKEN Harima Institute

description: TTHA0167-deficient strain cultivated for 480 min_2

hyb_protocol: 3'-terminal-labeled cDNA (2 micro g) was hybridized to a TTHB8401a520105F GeneChip (Affymetrix, Santa Clara, CA). The array was incubated for 16 h at 50°C in a solution comprising 180 mM MES, pH 6.6, 40 mM EDTA, 0.02% Tween 20, 7% dimethyl sulfoxide, 20 micro g of herring sperm DNA (Promega), 0.1 mg of bovine serum albumin (BSA), the recommended amount of Eukaryotic Hybridization Control (Affymetrix), and Control Oligo B2 for the alignment signal (Affymetrix), and then the array was automatically washed and stained with streptavidin-phycoerythrin (Invitrogen) by using a GeneChip Fluidics Station, 450XP (Affymetrix).

scan_protocol: The Probe Array was scanned with a GeneChip Scanner 3000 (Affymetrix).

data_processing: The expression levels were summarized by one-step Tukey’s biweight algorithm and normalized by global scaling method using GeneChip Operating Software, version 1.4 (Affymetrix, Santa Clara, CA). The trimmed mean target intensity of each array was arbitrarily set to 500.

organism: Thermus thermophilus HB8

source_name: T. thermophilus HB8, TTHA0167 deletion mutant, rich medium, 480 min

label_protocol: cDNA was synthesized with SuperScript II (Invitrogen) reverse transcriptase in the presence of RNase inhibitor SUPERase (Ambion) and 6 base random primers (Invitrogen). The cDNA was fragmented with 1.2 units of DNase I (GE Healthcare Bio-Science Corp.) at 37°C for 10 min, and after inactivation at 98°C for 10 min, the cDNA fragments were labeled with GeneChip DNA Labeling Reagent (Affymetrix, Santa Clara, CA), using terminal transferase according to the manufacturerâ€™s instructions (Affymetrix).

growth_protocol: The TTHA0167-deficient T. thermophilus HB8 strain was pre-cultured at 70°C for 16 h in 3 ml of TT medium containing 0.8% polypeptone, 0.4% yeast extract, 0.2% NaCl, 0.4 mM CaCl2, and 0.4 mM MgCl2, which was adjusted to pH 7.2 with NaOH. The cells (2 ml) were inoculated into 1 liter of the same medium and then cultivated at 70°C for 480 min.

extract_protocol: Cells were collected from 10 ml of the culture medium, and then crude RNA was extracted by the addition of 1.4 ml of a solution comprising 5 mM Tris-HCl, pH 7.5, 5 mM EDTA, 0.25% SDS, and 50% of water-saturated phenol. This mixture was incubated at 65°C for 5 min, chilled on ice for 5 min, and then centrifuged at 4°C. Then, 750 micro l of TRIZOL LS (Invitrogen, Carlsbad, CA) was added to 0.2 ml of the aqueous phase. After incubation for 5 min at room temperature, the RNA was extracted with 0.2 ml of chloroform. The extraction was repeated with 0.5 ml of chloroform, and the aqueous phase was precipitated with isopropanol. The pellet was dissolved in 0.2 ml of nuclease-free water, precipitated with ethanol, and then resuspended in 0.2 ml of water. The RNA was treated with DNase I (Ambion, Austin, TX) at 37°C for 20 min in a 25-micro l reaction mixture. The reaction was terminated by the addition of 1 micro l of 0.5 M EDTA, followed by incubation at 70°C for 5 min. Thereafter, the RNA was precipitated with ethanol in the presence of 1% glycogen and 1 M NH4OAc.

tag: genotype

value: TTHA0167-deficient strain


title: wt_2


title: TTHA0167-deficient strain_480_3

description: TTHA0167-deficient strain cultivated for 480 min_3


title: wt_4


title: TTHA0167-deficient strain_480_1

description: TTHA0167-deficient strain cultivated for 480 min_1


title: wt_1