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Old 06-17-2009, 08:38 AM   #1
jwaage
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Default 75mer mapping with BOWTIE

Hey all,

I'm looking for some input on tuning BOWTIE for 75mer RNA-seq reads from mus - any suggestions? I'm currently allowing only unique hits, and using 68bp as seed length, but that may be too conservative.

Best,
Johannes Waage
Uni. of Copenhagen
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Old 06-17-2009, 11:37 AM   #2
Ben Langmead
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Hi Johannes,

That's certainly reasonable; I suggest doing a series of runs using your target data but also using the -u parameter to cut the job off after a certain number of reads so that it doesn't take forever. Doing several such runs with different sets of parameters (but the same -u) should help you determine the sweet spot w/r/t speed and sensitivity.

Ben
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Old 06-17-2009, 03:41 PM   #3
graveley
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Hi,

We are doing this routinely. We typically use the -v 2 option and use an index made from both the genome and splice junctions with 69 nt on either side of the junction. Surprisingly, we get a very large fraction of reads that map uniquely using these parameters.

Brent
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Old 06-17-2009, 08:23 PM   #4
xuying
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Hi Johannes:
How do you deal with exon junctions? some exons will be shorter than 75bps. Try tophat?

Xuying
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Old 06-18-2009, 06:00 AM   #5
drio
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Can you tell me what reference are you using?
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Old 06-18-2009, 08:08 AM   #6
jwaage
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Quote:
Originally Posted by drio View Post
Can you tell me what reference are you using?
mm9 combined with a combinatorial exon-exon DB (142mers, requiring overlap of 4nt)
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