Run RNA-SeQC on tophat alignment result

[vivienne_lovely]

Hey guys,

I want to use RNA-SeQC to analyse my tophat output.I know that the default output (accepted_hits.bam) of tophat could not be as direct input of RNA-SeQC since it misses SM tag in the bam file header lines. I added the @RG line into the accepted_hits.bam, but the problem was not solved. Is there anyone can solve this problem?

Any comments are highly appreciated.
Thanks

vivienne

[vivienne_lovely]

I found the answer in this link:http://www.google.com.hk/url?sa=t&rc...kOLCuw&cad=rjt
but in China, I can't open this link, is there anyone can open it and copy the content "The default output (accepted_hits.bam) of tophat could not be as direct input of RNA-SeQC since it misses SM tag and doesn't sorted in..." for me? Thank you very much. Waiting for your help.
vivienne

[offspring]

The page you cited has the following commands:

#replace
java -jar biosoft/picard-tools-1.74/AddOrReplaceReadGroups.jar I=accepted_hits.bam O=accepted_hits_gr.bam LB=lane6 PL=illumina PU=lane6 SM=lane6 >AddorReplaceReadGrups2clippeddata.log 2>&1

#reorder
java -jar biosoft/picard-tools-1.74/ReorderSam.jar I=accepted_hits_gr.bam O=accepted_hits_gr_sort.bam R=database/index/hg19.fa

This is consistent with my findings trying to do the same thing (although I use the commands the other way round).
However, that only gives you the metrics for your mapped reads, since TopHat writes the unmapped ones to a seperate unmapped.bam file.

If you want to get metrics for your complete library you have to jump through a couple more hoops. See the following thread for details: http://seqanswers.com/forums/showthread.php?t=28155

Chris

[vivienne_lovely]

Thank you,Chris, this really helps a lot.
Vivienne