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URL: https://dx.doi.org/10.17504/protocols.io.7ejhjcn

Authors: John Tyson

Summary: Old-School Phenol/Chloroform Genomic HMW DNA PreparationIn order to mitigate damage/shearing of genomic DNA we have avoided kits etc. that employ beads or a matrix that your DNA must associate with or sieve through (blend :o)). We have not gone the whole hog at this point and used nuclei preps, dialysis or plug extractions etc. as we have found that material produced from a simple and rapid phenol/chloroform prep is more than adequate and high yielding. We have done some limited salting out experiments as a substitute for the phenol/chloroform approach but have some remaining questions around size and stability in the fridge for extended periods that need resolving. We will be revisiting this.The jumping off point for us was using methods detailed in “Molecular Cloning: A laboratory Manual” by Sambrooke and Russell. If you are at a large institution there will probably be copies around on people’s shelves or in the library collecting dust. It’s time to dust those off, they have been patiently waiting for their day in the sun again :o)). Chapter 6 is a good place to start. This approach produces DNA that is more than large enough for any nanopore sequencing currently.

Affiliations: Snutch Lab, UBC, Vancouver, BC, Canada

External URL: https://www.longreadclub.org/mountain-protocol/

Version: 1

Publication Date: 2020

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