MinIon Prep Kit Selection (Ligation vs. Rapid)

[hathormaat]

Hey all,

I'm looking to sequence a handful of pure bacterial isolates for WGS and de novo assembly. Our group has decided to go with a combined Illumina/Oxford nanopore approach. While our Illumina protocols are well developed (MiSeq v3 using the Pulsenet protocol), our MinIon sequencing is something we're having to start (more or less) from the ground up. This would be an ongoing project sequencing around 2-12 isolates a month.

Does anyone have insight into which kits have worked best for them? A lot of the literature I see discusses using the Rapid Sequencing Kit (SQK-RAD004), but not much in the way of the Ligation Sequencing Kit (SQK-LSK109). For my purposes, will the Rapid sequencing kit (in combination with Illumina sequencing) be enough? Is the Ligation kit + Illumina effectively overkill? I don't want to suggest a kit that requires $1500 of reagents just for a $600 kit to get essentially the same results.

Thanks!

[vivianshaw]

NGS Library Prep kits

You may want to check if Creative Biolabs can provide a better suggestion.

Here are the links to their DNA Library Prep and RNA Library Prep kits:

https://www.creative-biolabs.com/suprecision/category-rna-library-prep-284.htm

https://www.creative-biolabs.com/suprecision/category-dna-library-prep-283.htm

[jhalpin]

ONT sequencing kits

In our lab, we had trouble getting long DNA from the extractions and so the rapid kit didn't work for us. It has a built in fragmentation step that also applies the adapters/barcodes. The ligation kit you don't have to fragment the DNA to attach the adapters or barcodes so that's what we have been using.

If you're getting high molecular weight good quality DNA from your extractions it's my understanding that there's no reason that the rapid kit wouldn't work for you.