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Old 02-20-2014, 10:51 PM   #1
snehaldk
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Location: Bangalore

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Question How to use unpaired reads from trimmomatic output in SOAPdenovo for assembly?

Hi all,

I have illumina paired end reads trimmed using Trimmomatic - resulting in 4 files - 2 paired, 2 unpaired (as expected). Now I want to do assembly using SOAPdenovo2. I want to incorporate singleton/unpaired files in addition to paired files for assembly.

There is an option('q') to include singleton files from 'single end files' (which I believe refers to 'single end sequencing' and not paired). But I am not sure if I can use this option for my files where the sequencing is forward-reverse(paired). And if I do use it, should I make reverse complement of my R2_unpaired file before adding it to R1_unapaired and using it further? Or the direction does not matter for singleton files? Do the assembly programs descriminate between complementary strands?

Sorry if the questions are too naive. I am rather new to this.

Thanks in advance.
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Old 01-24-2015, 12:19 PM   #2
NYGen
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I also have this question - if anyone has insight on this, could you offer some?

Thanks very much!
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Old 01-25-2015, 11:48 PM   #3
sarvidsson
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Just concatenate the singleton files and use them as a single end sequencing file. If you only trimmed adapters with Trimmomatic I'd throw away those reads, however. If they result from quality trimming, you can keep them.

The fragment orientation in Illumina shotgun libraries is random - unless you have stranded RNA libraries - so there is no meaning of what is forward and reverse sequence across different sequenced clusters, just between the paired reads from a single cluster.
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