title: dcr-2_non-beta-eliminated

description:
This RNA sample was not beta-eliminated before adapter ligation.
This sample was not used in Ghildiyal et al. (2008) Science, where a better dataset (with optimized deep-sequencing protocol) was available (GEO sample #GSM278703).

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000323


title: WT_females_beta-eliminated

status: Public on Apr 11 2008

type: other

city: Worcester

first: Hervé

last: Seitz

state: MA

address: 364 Plantation Street

department: Biochemistry and Molecular Pharmacology

country: USA

institute: University of Massachusetts Medical School

description: This RNA sample was beta-eliminated as described in Horwich et al. (2007) Curr. Biol. 17(14):1265-1272 before adapter ligation.

hyb_protocol: not applicable

scan_protocol: not applicable

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000316

label: not applicable

molecule: other

organism: Drosophila melanogaster

source_name: WT female heads

label_protocol: not applicable

extract_protocol: Total RNA was extracted using the mirVana kit (Ambion), then short (18-30 nt) RNAs were gel-purified. 2S rRNA was depleted as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).

tag: Heads were dissected from Oregon R adult female flies as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).


title: head and neck squamous cell carcinoma (HNSCC) sampleNo.08

status: Public on Nov 24 2007

type: RNA

city: Chiba

first: Satoya

last: Shimizu

address: 1-8-1 Inohana, Chuo-ku

country: Japan

institute: Chiba university

hyb_protocol: Hybridized for 20 h at 50 C. Hybridization buffer and washing protocol was followed by the protocol supplied by Hitach Software Engineering Co., Ltd. http://www.dna-chip.co.jp/thesis/AceGeneProtocol.pdf

scan_protocol: ScanArray HT (PerkinElmer Japan Co., Ltd.) was used for scanning. Array images were analyzed with DNASIS Array (Hitachi Software Engineering Co., Ltd.).

data_processing: This data were analyzed by DNASIS array software(Hitachi Software Engineering), which converted the signal intensity of each spot into text format. Log2-ratios of the median subtracted background intensity levels were analyzed. Data from each microarray were normalized by LOWESS normalization.

molecule: total RNA

organism: Homo sapiens

label_protocol: Amino Allyl aRNA was synthesis by Amino Allyl MessageAmp™  aRNA Amplification Kit (Ambion). CyeDye Coupling and fragmentation were performed as the protocol supplied by Hitach Software Engineering Co., Ltd. http://www.dna-chip.co.jp/thesis/AceGeneProtocol.pdf

extract_protocol: TriZol procedure

tag: Non-tumorus tissue

tag: Tumor tissue

tag: Histlogical Type:moderately differentiated squamous cell carcinoma

tag: Stage:Ⅱ　(T2N0)


title: head and neck squamous cell carcinoma (HNSCC) sampleNo.10

tag: Histlogical Type:well differentiated squamous cell carcinoma

tag: Stage:ⅣA (T3N2c)


title: ago2_oxidized

description:
This RNA sample was oxidized before adapter ligation.
Protocol for oxidization:
Gel-purified 18-30 nt RNAs from 75 micrograms total RNA were incubated in 100 mM NaAcO (pH=5.5) and 33.3 mM freshly-dissolved NaIO4, for 5 minutes at room temperature, then ethanol-precipitated.

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000321

source_name: ago2 mutant heads

tag: Heads were dissected from ago2 mutant adult flies (ago2^{414} allele described in Okamura et al. (2004) Genes Dev. 18:1655-1666) as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).


title: head and neck squamous cell carcinoma (HNSCC) sampleNo.09

tag: Histlogical Type:poorly differentiated squamous cell carcinoma


title: ago2_untreated

description:
This RNA sample was not beta-eliminated before adapter ligation.
This sample was used in Ghildiyal et al. (2008) Science.

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000325


title: homozygous_dcr-2_untreated

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000324

source_name: homozygous dcr-2 mutant heads

tag: Heads were dissected from dcr-2 mutant adult flies (dcr-2^{L811fsX} allele described in Lee et al. (2004) Cell 117(1):69-81) as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).


title: ago2_beta-eliminated

description:
This RNA sample was beta-eliminated as described in Horwich et al. (2007) Curr. Biol. 17(14):1265-1272 before adapter ligation.
This sample was not used in Ghildiyal et al. (2008) Science, where simple oxidization (less prone to RNA degradation) was preferred to full beta-elimination.

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000319


title: WT_males_beta-eliminated

data_processing:
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure:
For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts".
Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt.
Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.

Raw data available at SRA under accession SRX000317

source_name: WT male heads

tag: Heads were dissected from Oregon R adult male flies as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).


title: oocytesmallRNA-19to24

status: Public on Mar 28 2008

type: SRA

city: Cold spring Harbor

first: ravi

last: Sachidanandam

state: NY

address: 1 bungtown road

institute: Cold Spring Harbor Laboratory

description: 19-24 nt fraction

library_source: transcriptomic

data_processing: The 3' linker, CTGTAGGCACCATCAAT, was identified and stripped out of the raw reads using a smith-waterman based alignment algorithm.

organism: Mus musculus

source_name: fully grown oocytes

extract_protocol: Total RNA was isolated from fully grown oocytes with Trizol, then separated PAGE to obtain three size fractions (19-24/24-30/19-30 nt). Adapters for Solexa sequencing were ligated, then the ligated product amplified by RT-PCR. The PCR product is gel-purified for sequencing

treatment_protocol: Mice were injected with 5 IU of PMSG 48 hours prior to sacrifice

tag: female 4-weeks old CD1 strain


title: oocytesmallRNA-24to30

description: 24-30 nt fraction


title: Day 2 High CO2

status: Public on Feb 08 2008

city: Plymouth

first: Jack

middle: Anthony

last: Gilbert

state: Devon

address: Prospect Place

country: United Kingdom

institute: Plymouth Marine Laboratory

description: Raw data for both the metagenomic and metatranscriptomic components are available at NCBI's Short Read Archive under accession SRP000101

data_processing:
Sequences were read and raw counts reported.

Raw sequence data files for this record are available for download from the SRA FTP site under accession SRX000133

organism: marine metagenome

source_name: Day 2 High CO2

growth_protocol: CO2 bubbled through 11,000 L mesocosm to simulate ocean acidification predicted conditions. Then phosphate and nitrate were added to induce a phytoplankton bloom.

extract_protocol: Total nucleic acid extraction was done as quickly as possible using the method of Neufeld et al. 2007.  For transcriptomics, total RNA was separated from the columns using the RNA MinEluteTM clean-up kit (Qiagen) and checked for integrity of rRNA using an Agilent bioanalyser (RNA nano6000 chip). High integrity rRNA is essential for subtractive hybridization. Samples were treated with Turbo DNA-free enzyme (Ambion) to remove contaminating DNA. The rRNA was removed from mRNA by subtractive hybridization (Microbe Express Kit, Ambion), and absence of rRNA and DNA contamination was confirmed using the Agilent bioanalyser. The mRNA was further purified with the MEGAclearTM kit (Ambion). Reverse transcription of mRNA was performed using the SuperScript® III enzyme (Invitrogen) with random hexamer primers (Promega). The cDNA was treated with RiboShredderTM RNase Blend (Epicentre) to remove trace RNA contaminants. To improve the yield of cDNA, samples were subjected to random amplification using the GenomiPhi V2 method (GE Healthcare). GenomiPhi technology produces branched DNA molecules that are recalcitrant to the pyrosequencing methodology. Therefore amplified samples were treated with S1 nuclease using the method of Zhang et al.2006. DNA and mRNA were sequenced using the 454 Corporation GS-FLX instrument at the NERC-funded University of Liverpool 454 Sequencing Facility. Extraneous sequences that resulted from > 1 template molecule per picotitre well were removed from the datasets.

treatment_protocol: Waters samples were prefiltered through a 1.6 um GF/A glass fibre filter to reduce Eukaryotic contamination. Filtrate was then collected on a 0.2 um Sterivex (millipore) filter which was frozen in liquid nitrogen until nucelic acid extraction.

tag: CO2, phytoplankton bloom, bacterial population


title: Day 1 High CO2

data_processing:
Sequences were read and raw counts reported.

Raw sequence data files for this record are available for download from the SRA FTP site under accession SRX000131

source_name: Day 1 High CO2


title: Day 2 Present Day

data_processing:
Sequences were read and raw counts reported.

Raw sequence data files for this record are available for download from the SRA FTP site under accession SRX000134

source_name: Day 2 Present Day

growth_protocol: Air bubbled through 11,000 L mesocosm. Then phosphate and nitrate were added to induce a phytoplankton bloom.

tag: air, phytoplankton bloom, bacterial population.


title: Day 1 Present Day

data_processing:
Sequences were read and raw counts reported.

Raw sequence data files for this record are available for download from the SRA FTP site under accession SRX000132

source_name: Day 1 Present Day


title: oocytesmallRNA-19to30

description: 19-30 nt fraction