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Old 02-05-2014, 01:46 AM   #2
Devon Ryan
Location: Freiburg, Germany

Join Date: Jul 2011
Posts: 3,480

Inputting the files like:
paired_1,unpaired_1 paired_2,unpaired_2
Will result in "unpaired_1" and "unpaired_2" being treated as paired, which is the opposite of what you want. To keep the strandedness correct you'll need to run things twice. Firstly using "paired_1,unpaired_1 paired_2" with library-type set to fr-firststrand and then just "unpaired_2" by itself with "fr-secondstrand". I should note that aligning unpaired_2 is usually not worthwhile (the reads are often crap), but perhaps you'll get luckier than I have with that.
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