I have some questions about read orientation in Tophat that I cannot find the clear answers yet and I hope you can shed some light on these.
1 What are the orientation of reads for pair-end data for library fr-unstranded,fr-firststranded, or fr-secondstranded? I'm not sure whether these strand specific parameter related with pair orientation (ff, fr, rf in Bowtie). In Bowtie, this parameter (ff, fr, rf) need to be specified and Tophat use Bowtie for alignment. However, I cannot find the way to set this parameter in Tophat.
2 According to description about library type, "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." does left-most end of fragment in transcript coordinates correspond to the read in first first file in paired-end data?
3 Some of public data that I collected do not have information about library type except only platform (standard illumine or strand specific Illumina), can I assume that they are fr-unstranded and fr-secondstrand respectively?