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Method Article
Erwan Denis
Wincker Lab
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Long read sequencing (Pacific Bioscience, Oxford Nanopore Technologies) and synthetic long read sequencing (chromium , 10x genomics) as well as technologies like next-generation mapping (Bionano Genomics) or Dovetail Chicago method need DNA molecules as long as possible. Even if traditional DNA extraction methods using commercials kits can lead to a maximum of 50-100kb long High Molecular Weight DNA (called HMW DNA), such fragment lengths may not be sufficient for some technologies. Isolation of high quality ultra-High Molecular Weight (uHMW) DNA, defined as a fragment of more than 50-100kb, can be a real challenge. The classical uHMW DNA isolation method is based on the isolation of cells or nuclei in agarose gel plugs. Nowadays, despite the fact that this is time consuming and requires handling expertise, this method is the only one recommended for DNA preparation for optical mapping. The impact of DNA molecule length for long read sequencing, optical mapping and Chicago method appears to be crucial. Here we present the development of a quick and inexpensive method to purify and isolate high yield of high quality uHMW from Saccharomyces cerevisiae.
Keywords
High molecular weight DNA, DNA extraction, nanopore, Saccharomyces cerevisiae, Yeast
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The authors declare no competing financial interests. Genoscope Institute was part of the Oxford Nanopore Technologies MinION Access Programme (MAP).
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Method Article
Erwan Denis
Wincker Lab
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 12 Jun, 2018
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Long read sequencing (Pacific Bioscience, Oxford Nanopore Technologies) and synthetic long read sequencing (chromium , 10x genomics) as well as technologies like next-generation mapping (Bionano Genomics) or Dovetail Chicago method need DNA molecules as long as possible. Even if traditional DNA extraction methods using commercials kits can lead to a maximum of 50-100kb long High Molecular Weight DNA (called HMW DNA), such fragment lengths may not be sufficient for some technologies. Isolation of high quality ultra-High Molecular Weight (uHMW) DNA, defined as a fragment of more than 50-100kb, can be a real challenge. The classical uHMW DNA isolation method is based on the isolation of cells or nuclei in agarose gel plugs. Nowadays, despite the fact that this is time consuming and requires handling expertise, this method is the only one recommended for DNA preparation for optical mapping. The impact of DNA molecule length for long read sequencing, optical mapping and Chicago method appears to be crucial. Here we present the development of a quick and inexpensive method to purify and isolate high yield of high quality uHMW from Saccharomyces cerevisiae.
Figure 1
Figure 2
Figure 3
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