Rapid isolation of yeast genomic DNA: Bust n' Grab

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dc.contributor.author Susanna Harju en_US
dc.contributor.author Halyna Fedosyuk en_US
dc.contributor.author Kenneth Peterson en_US
dc.date.accessioned 2009-05-05T16:13:40Z
dc.date.available 2004 - en_US
dc.date.available 2009-05-05T16:13:40Z
dc.date.issued 2004-04-21 en_US
dc.identifier.citation Susanna Harju;Halyna Fedosyuk;Kenneth Peterson: Rapid isolation of yeast genomic DNA: Bust n' Grab. BMC Biotechnol 2004, 4(1):8. en_US
dc.identifier.uri http://www.biomedcentral.com/1472-6750/4/8 en_US
dc.identifier.uri http://hdl.handle.net/2271/590
dc.description.abstract BACKGROUND:Mutagenesis of yeast artificial chromosomes (YACs) often requires analysis of large numbers of yeast clones to obtain correctly targeted mutants. Conventional ways to isolate yeast genomic DNA utilize either glass beads or enzymatic digestion to disrupt yeast cell wall. Using small glass beads is messy, whereas enzymatic digestion of the cells is expensive when many samples need to be analyzed. We sought to develop an easier and faster protocol than the existing methods for obtaining yeast genomic DNA from liquid cultures or colonies on plates.RESULTS:Repeated freeze-thawing of cells in a lysis buffer was used to disrupt the cells and release genomic DNA. Cell lysis was followed by extraction with chloroform and ethanol precipitation of DNA. Two hundred ng – 3 µg of genomic DNA could be isolated from a 1.5 ml overnight liquid culture or from a large colony. Samples were either resuspended directly in a restriction enzyme/RNase coctail mixture for Southern blot hybridization or used for several PCR reactions. We demonstrated the utility of this method by showing an analysis of yeast clones containing a mutagenized human ß-globin locus YAC.CONCLUSION:An efficient, inexpensive method for obtaining yeast genomic DNA from liquid cultures or directly from colonies was developed. This protocol circumvents the use of enzymes or glass beads, and therefore is cheaper and easier to perform when processing large numbers of samples. en_US
dc.format.extent 639801 bytes
dc.format.extent 2910 bytes
dc.format.extent 12055 bytes
dc.format.extent 0 bytes
dc.format.extent 43 bytes
dc.format.mimetype application/pdf
dc.format.mimetype text/plain
dc.format.mimetype application/octet-stream
dc.format.mimetype application/octet-stream
dc.format.mimetype application/octet-stream
dc.language en en_US
dc.language.iso en_US
dc.publisher BioMedCentral en_US
dc.relation.ispartof 1472-6750 en_US
dc.relation.hasversion http://www.biomedcentral.com/content/pdf/1472-6750-4-8.pdf en_US
dc.rights This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. en_US
dc.rights.uri http://creativecommons.org/licenses/by/2.0 en_US
dc.subject.mesh Blotting, Southern en_US
dc.subject.mesh Chromosomes, Artificial, Yeast en_US
dc.subject.mesh DNA, Fungal/ isolation & purification en_US
dc.subject.mesh Genome, Fungal en_US
dc.subject.mesh Globins/genetics en_US
dc.subject.mesh Mycology/ methods en_US
dc.subject.mesh Polymerase Chain Reaction en_US
dc.subject.mesh Saccharomyces cerevisiae/ genetics en_US
dc.title Rapid isolation of yeast genomic DNA: Bust n' Grab en_US
dc.type article en_US
dc.date.captured 2009-04-27 en_US
dc.identifier.doi doi:10.1186/1472-6750-4-8 en_US
dc.identifier.pmid 15102338 en_US

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This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Except where otherwise noted, this item's license is described as This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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