ATTENTION: The software behind KU ScholarWorks is going to be upgraded to a new version toward the end of July. Starting July 15th, users will not be able to log in to the system until the new version is in place. Searching for articles and opening files will continue to work while the system is being updated. If you have any questions, please contact Marianne Reed at mreed@ku.edu .

Show simple item record

dc.contributor.authorSusanna Harjuen_US
dc.contributor.authorHalyna Fedosyuken_US
dc.contributor.authorKenneth Petersonen_US
dc.date.accessioned2009-05-05T16:13:40Z
dc.date.available2009-05-05T16:13:40Z
dc.date.issued2004-04-21en_US
dc.identifier.citationSusanna Harju;Halyna Fedosyuk;Kenneth Peterson: Rapid isolation of yeast genomic DNA: Bust n' Grab. BMC Biotechnol 2004, 4(1):8.en_US
dc.identifier.urihttp://hdl.handle.net/2271/590en_US
dc.description.abstractBACKGROUND: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.languageenen_US
dc.language.isoen_USen_US
dc.publisherBioMedCentralen_US
dc.relation.isversionofhttp://www.biomedcentral.com/1472-6750/4/8en_US
dc.relation.hasversionhttp://www.biomedcentral.com/content/pdf/1472-6750-4-8.pdfen_US
dc.rightsThis 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.urihttp://creativecommons.org/licenses/by/2.0en_US
dc.subject.meshBlotting, Southernen_US
dc.subject.meshChromosomes, Artificial, Yeasten_US
dc.subject.meshDNA, Fungal/ isolation & purificationen_US
dc.subject.meshGenome, Fungalen_US
dc.subject.meshGlobins/geneticsen_US
dc.subject.meshMycology/ methodsen_US
dc.subject.meshPolymerase Chain Reactionen_US
dc.subject.meshSaccharomyces cerevisiae/ geneticsen_US
dc.titleRapid isolation of yeast genomic DNA: Bust n' Graben_US
dc.typeArticleen_US
dc.identifier.doi10.1186/1472-6750-4-8en_US
dc.identifier.pmidPMC15102338en_US
dc.rights.accessrightsopenAccessen_US
dc.date.captured2009-04-27en_US


Files in this item

Thumbnail
Thumbnail
Thumbnail
Thumbnail

This item appears in the following Collection(s)

Show simple item record

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.