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frascom 05-22-2019 09:54 AM

TrueSeq PCR free vs Nextera Mitogenome seq
I wondered if anyone had any recommendations for TrueSeq PCR free kits vs Nextera XT for mitogenome sequencing around 17,000bp pooling PCR fragments and around 192 individual samples on a single Miseq run?

Are there benefits of considering one over the other or protocols to follow for sample size and barcoding

thanks for the help

SNPsaurus 05-22-2019 12:47 PM

Can you multiplex that much on a Miseq and get enough reads per sample to reconstruct the mitogenome? I guess if the nuclear genome isn't large then a reasonable fraction of reads will go to the mitogenome.

You can use partial reaction of just Nextera DNA Flex and get good results. That might be the cheapest route.

frascom 05-23-2019 05:02 AM

I have amplified the mitogenome separately so there should no concern of reads. The main aim is trying to save some costs due to barcoding 192 individuals whilst achieving a high quality of reads. The benefit of NexteraXT I think is the availability of barcodes index for up to 384 samples. Although another option would be to use propriety barcodes so save a lot of money

SNPsaurus 05-23-2019 11:11 AM

I see, that makes sense for amplified mtDNA.

You can combine the Nextera Flex and Nextera XT barcode sets for Nextera Flex preps. We do that for increased multiplexing.

nucacidhunter 05-23-2019 07:25 PM

plexWell 96 or 384 could fit your library prep. It seems to be cheap and also quick.

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