Question: Tophat Library-Type : Illumina Truseq Stranded Total Rna Sample Prep Kit
7
gravatar for Nicolas Rosewick
6.3 years ago by
Belgium, Brussels
Nicolas Rosewick7.7k wrote:

Hi,

Anybody know which type of tophat library-type do I have to use with IlluminaTruSeq Stranded Total RNA Sample Prep Kit (paired-end 2x50bp) ?

Thanks ,

N.

illumina tophat strand • 31k views
ADD COMMENTlink modified 5.2 years ago by Biostar ♦♦ 20 • written 6.3 years ago by Nicolas Rosewick7.7k

So I've to do :

tophat -library-type fr-firststrand index R2.fastq R1.fastq and not tophat -library-type fr-firststrand index R1.fastq R2.fastq because R2 reads are aligning on the sens of the RNA fragment that is sequenced ? Is it correct ?

Thanks

ADD REPLYlink written 6.3 years ago by Nicolas Rosewick7.7k
13
gravatar for Irsan
6.3 years ago by
Irsan6.9k
Amsterdam
Irsan6.9k wrote:

fr-firststrand

Library Types:

  • fr-unstranded: Standard Illumina Reads from the left-most end of the fragment (in transcript coordinates) map to the transcript strand, and the right-most end maps to the opposite strand.
  • fr-firststrand:dUTP, NSR, NNSR Same as above except we enforce the rule that the right-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during first strand synthesis is sequenced.
  • fr-secondstrand: Ligation, Standard SOLiD Same as above except we enforce the rule that the left-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during second strand synthesis is sequenced.
ADD COMMENTlink modified 6.3 years ago • written 6.3 years ago by Irsan6.9k
1

The funny thing is that what is fr-firststrand for Tophat is RF for Trinity (transcriptome assembler).

ADD REPLYlink written 5.3 years ago by Biomonika (Noolean)3.1k
7
gravatar for dbrg77
5.3 years ago by
dbrg7770
United Kingdom
dbrg7770 wrote:

If you use Truseq stranded protocol, the library type should be fr-firststrand

check this analysis guide from Illumina, in the pdf file, they clearly suggest using fr-firststrand http://res.illumina.com/documents/products/technotes/rnaseqanalysistophat.pdf

ADD COMMENTlink written 5.3 years ago by dbrg7770
1

As of 2016-09-07, this is the correct URL for that document: http://www.illumina.com/documents/products/technotes/RNASeqAnalysisTopHat.pdf

ADD REPLYlink written 2.8 years ago by maizemu10

I like the direct answer like this. So for Illumina Truseq Stranded Total Rna Sample Prep Kit, we can use tophat -library-type fr-firststrand, is that correct?

ADD REPLYlink written 3.9 years ago by camelbbs650

what would be the library type if data from Illumina HiSeq 2000

ADD REPLYlink written 7 weeks ago by shweta.jajoriya0
2
gravatar for Puriney
5.8 years ago by
Puriney330
New York City
Puriney330 wrote:

enter image description here

This could illustrate how to select your library-type if you really want to know why so. http://onetipperday.blogspot.com/2012/07/how-to-tell-which-library-type-to-use.html

ADD COMMENTlink modified 5.8 years ago • written 5.8 years ago by Puriney330

Could you please share the Zhao Zhang's title paper? Thanks in advance.

ADD REPLYlink written 2.6 years ago by germelcar20
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