Hi, we would like to use a MiSeq to analyze amplicons (for barcoding projects- so there are thousands of slightly different sequences for each amplicon) longer than 300 bases, using Nextera library preparation kits. The tagmentation step in the Nextera kits results in fragments shorter than the original amplicons that can then be sequenced.
My question is, has someone devised software appropriate for re-assembling these amplicons, when there is a mixture containing thousands of different original amplicons, while avoiding misassemblies? We also probably need to use de novo methods of assembly since we do not have every sequence in our database that might be present in our (unknown composition) samples, so we don't have an appropriate reference for every sample. Because these sequences represent the same gene region from different organisms, and there are conserved regions, we are concerned about chimeric misassemblies from the data.
Liz
My question is, has someone devised software appropriate for re-assembling these amplicons, when there is a mixture containing thousands of different original amplicons, while avoiding misassemblies? We also probably need to use de novo methods of assembly since we do not have every sequence in our database that might be present in our (unknown composition) samples, so we don't have an appropriate reference for every sample. Because these sequences represent the same gene region from different organisms, and there are conserved regions, we are concerned about chimeric misassemblies from the data.
Liz
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