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| Status |
Public on Jun 17, 2012 |
| Title |
3P_Seq_PreMZT_2 |
| Sample type |
SRA |
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| Source name |
whole embryo at 1.5-2 hours post fertilization
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| Organism |
Danio rerio |
| Characteristics |
genotype/variation: wild type
tissue: whole embryo
developmental stage: 1.5-2 hours post fertilization
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| Treatment protocol |
Anesthetized decorioneted embryos were washed three times in PBS (137 mM NaCl, 2.7 mM KCl, 1.5 mM KH2PO4, 8 mM Na2HP04, pH 7.4) and suspended in PBS containing 1% freshly added formaldehyde. Embryos were transferred to a dounce homogenizer, dounced several times and incubated at room temperature for 15 min. Formaldehyde was quenched by adding 1/20 volume 2.5 M glycine. Cells were pelleted at 400 x g for 5 min. The supernatant was removed, and pellets were rinsed twice with PBS, flash frozen in liquid nitrogen and stored at-80C.
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| Growth protocol |
Zebrafish embryos or adults grown under standard condition
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
3P-Seq; see http://web.wi.mit.edu/bartel/pub/protocols.html
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer II |
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| Description |
3P-Seq 1.5-2 hpf lane 2
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| Data processing |
For 3P-Seq: Reads were reverse complemented and aligned to the D. rerio genome (Zv9/danRer7) using Bowtie. Reads that aligned to up to four genomic locus and had one or more mismatches at their 3' end within a terminal adenylate-run were carried forward as 3P tags. Reads mapping to the same locus with the same number of terminal adenylates were consolidated in the processed data file. The BED file is as in Jan et al. (GSE24924)
For 3P-PE-Seq: In each read pair, read #1 captured the 3' end of the poly(A) tail in the antisense orientation, and read #2 captured a portion of the 3' region of the transcript, and occasionally, the beginning of the poly(A) tail, in the sense orientation. Read #2 began with an adapter of 26 bases. Reads with more than 10 mismatches to the adapter were discarded and the first 26 bases were removed before further processing. A read pair was considered informative only if read #1 began with Ts and read #2 contained 2–39 terminal As. After leading Ts and terminal As were removed from reads #1 and #2, respectively, they were mapped to the genome using Bowtie, allowing for up to two mismatches and requiring a unique mapping position in the zebrafish genome. The length of the tail encoded in read #2 was defined as the maximum number of trailing As allowing for up to one mismatch. Only cases in which this number was larger than the number of As encoded in the genome at the predicted cleavage position by at least 2 bases were carried forward. Cleavage and polyadenylation position was defined as the last non-A base in read #2. The length of the poly(A) tail at that position was estimated using the corresponding read #1, and defined as the maximal i for which >90% of the bases in the first i bases of read #1 were Ts. This criterion was used to allow for some sequencing errors expected when sequencing long homopolymers.
Genome_build: danRer7
Supplementary_files_format_and_content: The .bed files contain the positions where 3P-Seq reads were mapped, and the name of each track is the number of reads mapping to that position.
Supplementary_files_format_and_content: The bigWig (.bw) files contain a summary of the number of reads whose 3' ends mapped to each position in the genome.
Supplementary_files_format_and_content: For the paired-end sequencing for mapping poly(A) tail lengths, the text files contain the following columns : chromosome, position, strand, number of poly(A) length reads, followed by the individual measurements (lower bounds on poly(A) tail length)
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| Submission date |
Apr 20, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Igor Ulitsky |
| Organization name |
Whitehead Institute for Biomedical Research
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| Street address |
9 Cambridge Center
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| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02472 |
| Country |
USA |
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| Platform ID |
GPL9319 |
| Series (1) |
| GSE37453 |
Extensive alternative polyadenylation during zebrafish development |
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| Relations |
| SRA |
SRX143560 |
| BioSample |
SAMN00862027 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM919966_3P_Seq_PreMZT_2.bed.gz |
8.0 Mb |
(ftp)(http) |
BED |
| GSM919966_3P_Seq_PreMZT_2.bw |
4.1 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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