Strandness setting for HISAT2 : BGI sequencing platform
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8 months ago
venura ▴ 70

Hi,

I have this question (prob a silly question). We have a set of RNASEq data that came from BGI sequencing platform. The dataset is PE 150bp (and stranded).

I am going to use HISAT2 for alignment and wondering about the strandness setting to be used. This is the exp procedure they used:

enter image description here

Any thoughts on what setting I should use R/RF or F/FR . By the Image I feel the latter should be used. Just trying to make sure. Thanks in advance.

RNA-Seq Hisat2 • 389 views
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Run GUESSmyLT and you should have your answer.

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Thank you for the link, Jacques!

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Is this even a stranded library? I would ask the facility that made the libraries for advise.

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I did. They said to follow the Illumina pipeline. :| When we placed the order we asked for stranded sequencing and PE 150

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Following Illumina pipeline is one thing but as ATPoint said earlier this library does not appear to be stranded (see: HISAT2 rna-strandness option )

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That is not an answer I would accept if I was you. You paid for it, so I personally would insist on details which means a protocol and details on the library prep. As of the picture you provide it does not seem that the library is stranded (at least based on the image) because (for me) it is not obvious whether the the first strand synthesis preserves the strand information. Ask for details, don't let them waive you off until you get the details you need for your analysis, you are the paying customer ;-)

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Writing back to them now! You are correct, I am going to give them a hard time! :)

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They replied and said it was a mistake :) and sent their original logs as proof. and asked me to replace the old image with this.

New Image

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Please edit the original post and replace the image in it.

Looks like this is a standard dUTP library.

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Just updated the original post.

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