SEQanswers

Go Back   SEQanswers > Bioinformatics > Bioinformatics



Similar Threads
Thread Thread Starter Forum Replies Last Post
pre-filtering before mapping? NanYu SOLiD 9 03-29-2012 02:29 AM
pre-built indexes biofreak RNA Sequencing 2 07-26-2011 02:52 PM
PubMed: Reduction of non-insert sequence reads by dimer eliminator LNA oligonucleotid Newsbot! Literature Watch 0 05-10-2011 10:10 AM
filtering pre-mRNA chrisbala Bioinformatics 0 11-19-2010 08:15 AM
WGA pre-Next Gen Sequencing Suzanne Sample Prep / Library Generation 5 03-19-2009 09:04 PM

Reply
 
Thread Tools
Old 02-10-2012, 06:32 AM   #1
cerebralrust
Junior Member
 
Location: Sweden

Join Date: Jan 2012
Posts: 8
Exclamation Extent of reduction during pre-processing

I'm just curious as how much reduction others observe during pre-processing of ngs data.

I've got 454 rna-seq data of about 300,000 reads.
Based on the fastqc report and using fastx toolkit,
- removing reads with bases below quality score of 20
- removing reads containing Ns
-removing ribosomal rna sequences (167 using bwa)
-removing reads below 100bp length

the dataset reduced to 185,735. I felt like this is too small. Is such a reduction common?
I could retain the reads less than 100bp in length , align them separately and align the entire dataset again.

I appreciate any advice and/or insight. Thank you.
cerebralrust is offline   Reply With Quote
Old 02-10-2012, 07:50 AM   #2
westerman
Rick Westerman
 
Location: Purdue University, Indiana, USA

Join Date: Jun 2008
Posts: 1,104
Default

It does seem like you are being rather strict. While it depends on your project and the program you wish to use, most NGS programs are rather insensitive to poor quality and short reads. Thus I rarely eliminate these. Of course rRNA can mess up a project as can adapter sequences so it can be useful to remove (or trim) reads that match those sequences.
westerman is offline   Reply With Quote
Old 02-10-2012, 08:09 AM   #3
cerebralrust
Junior Member
 
Location: Sweden

Join Date: Jan 2012
Posts: 8
Default

Thank you very much for your reply, Rick!

I'm using both MIRA and Trinity for assembly. Trinity is supposedly less sensitive to low quality compared to MIRA.

Anyhow, i will stick to removal of rRna and trimming adaptor sequences prior to assembly.

Thanks a lot!
cerebralrust is offline   Reply With Quote
Old 04-30-2013, 10:53 PM   #4
Manal
Junior Member
 
Location: KSA

Join Date: Apr 2013
Posts: 8
Default

Hi All
I have one basic question, I am new in the NGS field / Illumina and little bit confused about the meaning of Phasing and Prephasing rate and how does it happen?
Can anyone explain ?

Thanks
Manal is offline   Reply With Quote
Old 05-01-2013, 04:17 AM   #5
mastal
Senior Member
 
Location: uk

Join Date: Mar 2009
Posts: 667
Default Extent of reduction during pre-processing

See the explanation in the Rationale section of this paper:

http://genomebiology.com/2009/10/8/R83

Basically phasing/prephasing refers to the problem that not all the molecules in a cluster on the flow cell will successfully incorporate a base during each cycle, or some may incorporate more than 1 base.
mastal is offline   Reply With Quote
Reply

Tags
454, pre-processing, rna-seq

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 06:58 PM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2019, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO