Peter Weil
Faculty Member
Last active: 11/4/2015

A rapid technique for the determination of unknown plasmid library insert DNA sequence directly from intact yeast cells.

Klebanow ER, Weil PA
Yeast. 1999 15 (6): 527-31

PMID: 10234790 · DOI:10.1002/(SICI)1097-0061(199904)15:6<527::AID-YEA374>3.0.CO;2-0

A polymerase chain reaction (PCR)-based technique is described which allows for the determination of library plasmid insert DNA sequence directly and rapidly from intact yeast cells. Yeast spheroplasts are used to template a PCR reaction to amplify the insert sequence. This PCR product is then purified and its sequence directly determined using thermal cycle sequencing. Readable sequence can reproducibly be obtained from multiple yeast colonies in just two days. Uses of this technique in yeast two-hybrid screening as well as other types of yeast library screens are discussed.

MeSH Terms (11)

Fungal Proteins Genes, Fungal Genetic Vectors Genomic Library Plasmids Polymerase Chain Reaction Sequence Analysis, DNA Spheroplasts Templates, Genetic Time Factors Yeasts

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