Hello
I have WIG file and need to find genes regulated by the protein which is alternatively spliced (to be precise - the type of alternative splicing should be a5ss - alternative donor sites)
Please advise what programs should I use for this purpose.
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marina-orlova
MATS is my favorite tool for splicing event analysis
As mentioned in the previous comment, MISO is another option. In both cases, you need .bam files for RNA-Seq data (neither uses ChIP-Seq data, which is probably not a reliable tool for analyzing splicing events).
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Charles Warden
Thank you very much for the answer. I have both CHiP-seq and RNA-seq data. At first I should find peaks which are situated in introns and 3-UTR regions, and then check if the genes where peaks are situated are alternatavely spliced (a5ss). Could you please advise how to connect these two parts of the task?
I think you have the right idea. I don't know of a specific program to accomplish this all at once. For the ChIP-Seq peak calling, I assume you can use something standard like MACS. If you have a list of interesting coordinates, you could use bedtools to look for overlaps between peaks and regions
For an alternative 5` splice site, I would probably look for peaks around promoters where the ChIP-seq binding is upstream of the start of transcription, but maybe you have something different in mind.
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Charles Warden
Since this got mentioned in a more recent thread, I think I should probably say that I currently like QoRTS + JunctionSeq
If you have RNA-seq data you can use TopHat, however it uses fastq/bam/sam files: http://bioinformatics.oxfordjournals.org/content/25/9/1105.short
Or if you already know R language, you can use the Bioconductor package Rsubread, using the "subjunc" function.
Thank you! I will try using both
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marina-orlova
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Alternative splicing analysis tools require BAM files so that they can use both read abundance across exons and reads spanning splice junctions to identify isoforms. WIG files are for visualization purpose. I don't know of any tools that takes wig files as input and predicts alternative splicing events. Rather, WIG files are often produced as output files from RNA-seq analysis.
Thank you for your answer. I found also sam and fastq files for input and chip-seq, hope this is enough. Could you please advise what programs should I use now?
You can use MISO that will take the SAM files as input. You can also use Tophat and Cufflinks that will take fastq files as input. I would suggest you to use Tophat+Cufflinks.
Thank you a lot!