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Old 04-12-2011, 05:59 AM   #1
LFM
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Location: Scandinavia

Join Date: Mar 2011
Posts: 5
Default Targeted Resequencing - Target file issues

Hello!
After an intensive month following getting my targeting resequencing data, and working with this kind of data for the first time, going from panic attacks to "it's not so bad, I just didn't analyse correctly"; I have finally come to the point were I can pose some questions that actually make sense.
I am using SureSelect target enrichment, in order to design my probes, I uploaded a file to the eArray service that looks like this (one line for each region):

chr1:1234-56789
chr1:2345-98765... etc (1124 regions).

Now, I wanted to try aligning my results with BWA to exactly these regions. What I did was I uploaded these regions as custom tracks, then downloaded the sequences from the UCSC browser.
Now, and this is where everything gets really stupid, all my little regions have the same identifier:
>hg19_ct_UserTrack_3545_1 range=chr1:13456-19087 5'pad=0 3'pad=0 strand=+ repeatMasking=none
so after I went through my whole workflow, and I tried to see my alignment using Tablet, things wouldn't work (no wonder).
So, what did I do?
As the lazy person I am (I wasn't going to do things manually, or with find and replace!), I decided to look in the threads here and found the one recommending Unix and Perl for Biologists, and I took a flash course during a weekend.
So, I made a little Perl script that would add sequential numbers at the end of the region identifier (yeah, I know pretty pathetic, but I am so proud, kind of like reaching the top of the Himalayas), and I solved my problem temporarily.
Now, I want my features in these target regions too, so I do the same procedure, download a GTF file and add numbers and try to open it in Tablet, but apparently there are no features matching to my sequences.

I probably have been doing this all wrong from the beginning, so if anyone could tell me, how to retrieve my target sequences so the identifiers are unique, and at the same time retrieve the features in this area - I also wanted to get information about variations in the target region, but I have not gotten that far, I guess it could be done all at the same time?
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chromosomal coordinates, chromosomal regions, download gtf, targeted resequencing

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