Hi everyone. I am interested to find out microsatellite from publicly available transcriptome data. I want to use sequencing data from SRA archive of NCBI. I am using CLC Bio workbench for processing of data. In the first very step I am having trouble. Is the sequence read of SRA file is adapter trimmed or not. Another problem I faced while using illumina paired data that using SRA tool kit fastqdump it can not split the data into two files.
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written
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rakeshmbb •
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I look at project (and run, sample, etc) metadata at the respective SRA pages, e.g. here for this run. Which version of fastq-dump are you using? Running:
fastq-dump --split-files SRR2163549
gave me just one fastq file, but also the following output:
Rejected 6856409 READS because READLEN < 1 Read 6856409 spots for SRR2163549 Written 6856409 spots for SRR2163549
So I suspect either read 2 was trimmed for some reason, or the metadata is incorrect, or this record is corrupted
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To check if the sequences are adapter trimmed you can use
FastQCtool: http://www.bioinformatics.babraham.ac.uk/projects/fastqc/Passing the data through a trimming program is not a bad idea. If it was trimmed then it should come through without changes. Only thing that you have to invest in is some time. This should apply to trimming tool in CLC as well.
As for fastq-dump you need to use the
--split-filesoption to get the two reads in two separate files. While you are at it may as well use the--origfmtoption to recover the original fastq file headers.What SRA # are you looking at?
Get the fastq files from EBI: http://www.ebi.ac.uk/ena/data/view/ERR1203908
Did you check the project metadata? The information if the reads have been trimmed or not may be available there. Also, maybe fastq-dump is not splitting files because it is a single end run?