Hi JIwen,

As the seqanswers thread shows, there is some debate about this. One of the last posts there makes the most sense - where "fragment length" is defined as the total genome bases covered by the aligned paired sequences: tip of 5' start, through the gap, to the very 3' tail end and where "mean inner distance" is the gap in the middle (between the paired seqs) where there is no alignment. This could be tested with smaller samples of data and compared to your fragment selection, to see how it matches up. But, for a definitive answer, asking the tool authors is the best bet. The contact information is: tophat.cuffli...@gmail.com

If you are given a reply that explains the calculation, it would be great if the TopHat documentation itself were updated.
http://tophat.cbcb.umd.edu/manual.html

And we would be very glad to hear of the details, so that here at Galaxy we could add the information to our RNA-seq help and the searchable mailing list archives.

Great question! If we find out ourselves meanwhile, an update will be posted back here to the mailing list,

Best,

Jen
Galaxy team

On 3/6/12 12:23 PM, 杨继文 wrote:
Hi all,
When mapping pair end RNA-seq reads using tophat, we need to type in
"Mean Inner Distance between Mate Pairs". In galaxy, we can read the
following information:

This is the expected (mean) inner distance between mate pairs. For, example, 
for paired end runs with fragments
  selected at 300bp, where each end is 50bp, you should set -r to be 200. There 
is no default, and this parameter
  is required for paired end runs.

I think the size of fragment (here 300bp) includes not only the length of pair 
end reads, but also the length of adaptors. so, maybe the Mean Inner Distance 
between Mate Pairs should be : fragment length - pair end read length - adaptor 
length. Am I right? or did I miss something?

Is it a must to type in the accurate value?

Looking forward to your reply

JIwen





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