摘要：

This chapter discusses one-step gene disruption in yeast. The method can be used to determine whether a gene is essential by first disrupting the gene in a diploid. The one-step gene disruption technique requires cloning of a selectable gene fragment into the region to be disrupted. Cloning techniques have permitted the isolation of numerous yeast genes. For genes cloned by complementation, it is necessary to demonstrate that a fragment codes for the wild-type gene, not for a phenotypic suppressor. For disruptions of essential genes, the recessive lethal phenotype becomes tightly linked to the insertion that is nonreverting and can be used as a selectable marker for further genetic manipulation. The resultant chromosomal insertion is nonreverting and contains a genetically linked marker.

展开