Hi all,
I am trying select sequences for capture enrichment using an unpublished genome of the same species available from NCBI as a reference. The reference genome seems to be pretty good quality (approximately right size, large scaffolds, etc).
I went through and picked my sequences of interest (~500kb), broke them up into ~100bp sequences, and then used BLASTn to see if there were any sequences that had multiple hits in the genome, and threw them out. I also BLASTed my selected sequences against themselves and threw out any that had multiple hits. Finally, I thought I would check these for low complexity regions, so I ran them through RepeatMasker online, selecting the most closely related species in the database. About 40% of my sequences were masked (~15% are LINE1).
RepeatMasker is obviously much more sensitive than BLAST, and repeats could potentially be more divergent than could be detected with BLASTn. But I would have thought I would have had more BLAST hits of my sequences to themselves or to the reference genome with such a large proportion being repetitive. Any thoughts about why this might be?
I am trying select sequences for capture enrichment using an unpublished genome of the same species available from NCBI as a reference. The reference genome seems to be pretty good quality (approximately right size, large scaffolds, etc).
I went through and picked my sequences of interest (~500kb), broke them up into ~100bp sequences, and then used BLASTn to see if there were any sequences that had multiple hits in the genome, and threw them out. I also BLASTed my selected sequences against themselves and threw out any that had multiple hits. Finally, I thought I would check these for low complexity regions, so I ran them through RepeatMasker online, selecting the most closely related species in the database. About 40% of my sequences were masked (~15% are LINE1).
RepeatMasker is obviously much more sensitive than BLAST, and repeats could potentially be more divergent than could be detected with BLASTn. But I would have thought I would have had more BLAST hits of my sequences to themselves or to the reference genome with such a large proportion being repetitive. Any thoughts about why this might be?
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