Hello all,
I am trying to figure out how to sequence amplicons I have made using a microsatellite DNA enrichement protocol.
Briefly, I took genomic DNA from an animal, digested it with restriction enzymes, ligated adapters on the fragments, hybridized them with biotinylated oligos with repetitive elements, recovered the hybridized DNA with streptavidin beads, and recovered the enriched DNA with PCR.
So now I have multiple amplicons from each animal, with an average length of 500bp, and each amplicon has a known adapter sequence on each end.
Illumina doesn't have a kit that supports this protocol, so I am trying to figure out what will be the best way to load my samples on a MiSeq for PE reads. I will also need multiplex several (6) samples on one flowcell.
Will the TruSeq sample prep kit work, if I skip the end repair stage (as my amplicons are blunt ended) and proceed from the adenylation stage?
Any input will help! This NGS is all new to me.
I am trying to figure out how to sequence amplicons I have made using a microsatellite DNA enrichement protocol.
Briefly, I took genomic DNA from an animal, digested it with restriction enzymes, ligated adapters on the fragments, hybridized them with biotinylated oligos with repetitive elements, recovered the hybridized DNA with streptavidin beads, and recovered the enriched DNA with PCR.
So now I have multiple amplicons from each animal, with an average length of 500bp, and each amplicon has a known adapter sequence on each end.
Illumina doesn't have a kit that supports this protocol, so I am trying to figure out what will be the best way to load my samples on a MiSeq for PE reads. I will also need multiplex several (6) samples on one flowcell.
Will the TruSeq sample prep kit work, if I skip the end repair stage (as my amplicons are blunt ended) and proceed from the adenylation stage?
Any input will help! This NGS is all new to me.
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