5′ Adapter stripping was designed to be used with Nimbelgen Sequence Capture Arrays, (Okou ,et al. 2007), (Albert, et al. 2007)and also a discussion on SeqAnswers. The Nimblegen protocol includes a PCR step to amplify the fragments and it’s possible that some PCR primer is retained on the fragments and included in the reads.
The PCR adapter stripping is designed to remove full or partial 5′ PCR adapter sequence. Partial adapters may occur with some protocols designed to remove the PCR adapter prior to sequencing. With paired end reads, adapter sequence can be removed from both reads.
Command Line Option
|-a sequence1 sequence2||Strips sequence1 and sequence2 from paired-reads. If -a used without arguments then defaults to standard Illumina PE adaptor sequences|
|-5 sequence||Strips 5′ primer sequences from reads before aligning. Default is not to strip 5′ sequences.
This option is useful where the sample preparation protocol involved an additional PCR step with non-Solexa primers that may still be present on the 5′ ends of reads.
This option is similar to the -a except that it acts on the 5′ end of reads. It will also strip partial primer sequences.
The Novoalign adapter stripping is aggressive in that it can remove very short lengths of adapter, down to as little as 1 bp of adapter. This aggressiveness is deliberate in that removing the odd extra base is seen as preferable to calling a mismatches for a few bp of adapter that might not be stripped in less aggressive approaches.
Adapter stripping is performed by ungapped alignment of the adapter sequence to the 5′ end of the read. The alignment uses base qualities to calculate mismatch penalties