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lh3 / minimap2

A versatile pairwise aligner for genomic and spliced nucleotide sequences
https://lh3.github.io/minimap2
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How to map to combinatorial plasmid libraries #1178

Closed c-ruprecht closed 7 months ago

c-ruprecht commented 7 months ago

Hi, I am using golden gate assemblies to create modular plasmid libraries. I currently am looking at potentially a 80 combinations of plasmids that are quite similar. (4 Promoters x 8 Gene A x 8 Gene B ) backbone---PromoterRBS1---GeneA1---GeneB1---Barcode backbone---PromoterRBS2---GeneA1---GeneB1---Barcode ... backbone---PromoterRBS4---GeneA4---GeneB4---Barcode

I sequence the multiplexed plasmids using and demultiplex using the barcode. I want to identify the corresponding plasmids using the demultiplexed reads. Currently I am using:

minimap2 -x map-ont --secondary=no

I then take the mapping with the highest score for each read and assign the plasmid by the highest abundance of a mapping. This works somewhat ok with a control experiment but I want to work on my minimap settings and I am wondering what the right minimap settings are for my situation. Thanks for the help!

lh3 commented 7 months ago

You may consider to add -f 100 in case minimap2's automatic heuristic chooses an inappropriate k-mer threshold. As to other options, you have to try by yourself as I haven't worked with such data.