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Hello to all,
I have one dataset that contains samples (2 conditions in 2 replicates) sequenced with "TruSeq Stranded mRNA Sample Prep Kit" : directional mRNA-seq.
How is it possible to study the antisense transcripts? In fact, I have theoretical and technical questions about it.
- antisense transcripts have as template the sense strand of DNA (and thus have the same sequence as the antisense strand of DNA) ? So antisense transcripts are transcribed from + strand ? and conversely, sense transcripts have as template the antisense strand of DNA (and thus have the same sequence as the sense strand of DNA) ?
- when I align my data with the TopHat tool (with fr-firstrand option, because this is a dUTP protocol), I have a XS:A tag by read, following by + or -. So, reads that are tagged by XS:A:+ correspond to sense transcripts and reads that are tagged by XS:A:- correspond to antisense transcripts ?
- in this topic "http://genomebytes.wordpress.com/2013/07/08/fixing-the-xs-tag-in-tophat-output-bug-fixing/", Chung-Chau Hon said : "If a read is mapped on a feature specified in the GTF, the XS tag would always be on the same strand of the feature not matter what strand the read is mapped to. This would create wrong counts is there are unannotated antisense transcripts.". How do you study these antisense transcripts?
Thank you in advance for your response.
I have one dataset that contains samples (2 conditions in 2 replicates) sequenced with "TruSeq Stranded mRNA Sample Prep Kit" : directional mRNA-seq.
How is it possible to study the antisense transcripts? In fact, I have theoretical and technical questions about it.
- antisense transcripts have as template the sense strand of DNA (and thus have the same sequence as the antisense strand of DNA) ? So antisense transcripts are transcribed from + strand ? and conversely, sense transcripts have as template the antisense strand of DNA (and thus have the same sequence as the sense strand of DNA) ?
- when I align my data with the TopHat tool (with fr-firstrand option, because this is a dUTP protocol), I have a XS:A tag by read, following by + or -. So, reads that are tagged by XS:A:+ correspond to sense transcripts and reads that are tagged by XS:A:- correspond to antisense transcripts ?
- in this topic "http://genomebytes.wordpress.com/2013/07/08/fixing-the-xs-tag-in-tophat-output-bug-fixing/", Chung-Chau Hon said : "If a read is mapped on a feature specified in the GTF, the XS tag would always be on the same strand of the feature not matter what strand the read is mapped to. This would create wrong counts is there are unannotated antisense transcripts.". How do you study these antisense transcripts?
Thank you in advance for your response.