Thursday, February 2, 2012

RNA-seq pipeline

Hello readers,
I've been working on a computational pipeline to analyze RNA-Seq data in order to detect alternatice splicing and chimeric transcripts. It got published last month in Nucleic Acid Research. Please find the paper at:
http://nar.oxfordjournals.org/content/early/2012/01/28/nar.gks047.long

The program is freely available at: www.mcdonaldlab.biology.gatech.edu/r-sap.htm.
The pipeline is actively under development so feel free to use it and any suggestions and feedbacks are most welcome.

5 comments:

  1. Hi Vinay,

    I am new to bioinformatic field. your blog help me a lot in analysis NGS data's. At present i started working on RNA-SEQ. I have come across about R-SAP through your blog. I am working on arabidopsis rna-seq data. Can you guide me how to convert the fastq file to Psl/Pslx file

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  4. Hi Saranya,
    Thanks for your interest. This pipeline works well and accurately for assembled contigs from the short-read data or longer reads (such as from 454 or PacBio sequencing technologies).
    Since you have FastQ files, I presumably you will have short-read sequence data. I suggest you to do the assembly first (reference based using Cufflinks or Scripture if you are interested in detecting novel splice variants ) or de-novo assembly using Trans-ABySS or trinity (or any other similar program) if you are interested in detecting fusion-gene transcripts.
    For reference-based assembly, you can directly supply resulting GTF or BED files to R-SAP but for de-novo assembly, you will have to align the assembled contigs to the reference genome using SSAHA2 or BLAT. Then you can use resulting psl/pslx files with R-SAP.

    If you are inclined to use your short-read sequence data directly with R-SAP then try aligning them to the reference genome using SSAHA2 and then run R-SAP on the alignment file. But this way you will get a lot of noise in the final output.
    Hope this helps.

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