Year: 2025
Type: preprint
Abstract: This protocol is for the automated extraction and SPRI of HMW DNA from cryoprepped tissue samples from plants and fungi intended for long-read sequencing using the Qiagen MagAttract HMW DNA extraction kit and the Thermo Fisher KingFisher™ Apex. This process is effective for a wide variety of plant species covered by the Tree of Life Programme. The output of this protocol is HMW DNA, which – depending upon yield and genome size of the species – can be directed towards either HMW DNA Pooling, HMW DNA Fragmentation: Opentrons® OT-2 for PacBio LI, HMW DNA Fragmentation: Diagenode Megaruptor® 3 for LI PacBio or HMW DNA Fragmentation: Covaris g-Tube for ULI PacBio. This protocol was adapted from Sanger Tree of Life HMW DNA Extraction: Automated Plant MagAttract v.4 to include an overnight elution in order to maximise the yield of DNA, as well as an updated pre-shear SPRI to improve recoveries of HMW DNA. Acronyms HMW: high molecular weight SPRI: solid-phase reversible immobilisation LI: low input ULI: ultra-low input
Authors: Graeme Oatley, Amy Denton, Anna Kovalevskaia, Caroline Howard
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
OpenAlex ID: W7084112752
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