Pelechano 2013, Transcript Isoform mapping.

Description

TSS mapping in WT S. cerevisie cells grown in galactose or glucose media. Several technical (denoted as 'lib') and biological (denoted as 'bio') replicas are provided.

Source

Samples

From Yeast Apr 2011 (NCBI3.1/sacCer3) Assembly.

RNA-seq data:

Filename Description Feature GEO-ID
1 GSM956495.sga WT|TSS-seq|Galactose|lib1-bio1 TSSseq GSM956495
2 GSM956496.sga WT|TSS-seq|Galactose|lib1-bio2 TSSseq GSM956496
3 GSM956501.sga WT|TSS-seq|Galactose|lib2-bio3 TSSseq GSM956501
4 GSM956502.sga WT|TSS-seq|Galactose|lib2-bio4 TSSseq GSM956502
5 GSM956511.sga WT|TSS-seq|Galactose|lib3-bio3 TSSseq GSM956511
6 GSM956512.sga WT|TSS-seq|Galactose|lib3-bio4 TSSseq GSM956512
7 GSM956515.sga WT|TSS-seq|Galactose|lib4-bio1 TSSseq GSM956515
8 GSM956516.sga WT|TSS-seq|Galactose|lib4-bio4 TSSseq GSM956516
9 GSM956493.sga WT|TSS-seq|Glucose|lib1-bio1 TSSseq GSM956493
10 GSM956494.sga WT|TSS-seq|Glucose|lib1-bio2 TSSseq GSM956494
11 GSM956499.sga WT|TSS-seq|Glucose|lib2-bio1 TSSseq GSM956499
12 GSM956500.sga WT|TSS-seq|Glucose|lib2-bio2 TSSseq GSM956500
13 GSM956509.sga WT|TSS-seq|Glucose|lib3-bio1 TSSseq GSM956509
14 GSM956510.sga WT|TSS-seq|Glucose|lib3-bio2 TSSseq GSM956510
15 GSM956513.sga WT|TSS-seq|Glucose|lib4-bio1 TSSseq GSM956513
16 GSM956514.sga WT|TSS-seq|Glucose|lib4-bio2 TSSseq GSM956514

Technical Notes

The protocol to construct the sequencing library is quite different from the normal TSS mapping promotocol. In this case, it is designed to map both ends of a transcript. To do so, it links them using an adapter. Sequencing adapters (with barcodes) are then added within the transcript (not so far away form the 5' or 3' end) resulting in reads that start on the transcript, goes towards the ends and cover the adapter. They are then sequenced using paired-end reads. This results in the 5' end being sequence on the reverse, with the TSS located in the middle of the reads. Since we were only interested in the 5' ends (TSS-mapping), we mapped the 3' end of the adapter to the reads [TGAGCAATACC] (the reverse-complement of it - GGTATTGCTCA), extracted the sequence before its starting point (this is the TSS) and produced the reverse complement of it. Finally, the sequencing barcodes was removed from the reads (7 nucleotides at the beginning of each read). Once trimmed, the reads longer than 30 bp were mapped to the genome using Bowtie2 v2.2.9. SAM files were then converted into bam using samtools v0.1.14 and to bed using bamToBed v2.12.0 (bedtools). SGA conversion was carried out using bed2sga.pl (ChIP-Seq v. 1.5.2).

References

Last update: 21 Jun 2016