We are developing a library preparation for directional mRNA-seq. All sequences will read 5' -> 3' direction. Our protocol isolates mRNA, fragments the messages and then these are re-purified with oligodT - resulting in a library of short 3' fragments for sequencing. This eliminates the need to adjust for transcript length when doing quantitative expression analysis. We're using bowtie to align the raw reads to a (cDNA) reference transcriptome (minus introns and intergenic regions). BUT - approximately 12% of the loci in this organism (Arabidopsis) have overlapping 3' tails (Watson/Crick strands) leading to ambiguities when using BowTie ( which looks for alignments on both strands). Basically we're looking for a short read alignment algorithm that would take our directional data and align it ONLY with transcripts in the 5' -> 3' direction. Illumina is developing a directional mRNA-seq library prep method similar to the one we are using (without the purification of the 3' ends). I wonder if anyone has experience analyzing directional mRNA-seq data?
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Originally posted by jmtepp View PostBasically we're looking for a short read alignment algorithm that would take our directional data and align it ONLY with transcripts in the 5' -> 3' direction.
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strand specific RNA-seq mapping
Does anyone know how do mapping for strand specific RNA-seq data? which tools can work well? what the parameter of tools for strand specific infomation of RNA-seq data?
Looking forward to your answer.
Thank you very much! Best wishes!
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Originally posted by qqtwee View PostDoes anyone know how do mapping for strand specific RNA-seq data? which tools can work well? what the parameter of tools for strand specific infomation of RNA-seq data?
Looking forward to your answer.
Thank you very much! Best wishes!
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Mappind strand specific RNAseq bacteria pair-end reads
Just following up on this thread, which mapper will be most ideal for mapping strand specific RNAseq bacteria pair-end reads,
I know tophat has the library type option but its mostly ideal for eukaryotic genomes.
Any thoughts will be appreciated.
Thanx
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