Send to

Choose Destination
Gene. 2011 Sep 15;484(1-2):86-9. doi: 10.1016/j.gene.2011.06.006. Epub 2011 Jun 22.

Optimized protocols and plasmids for in vivo cloning in yeast.

Author information

Dept. of Biological Sciences, University of Alabama in Huntsville, AL, USA.


Saccharomyces cerevisiae has proven a valuable system for the construction of plasmids via gap repair or in vivo cloning. The method allows cloning with superior accuracy and without the need to use restriction enzymes. However, despite its remarkable efficiency, the process may occasionally require the screening of large number of candidates. We have previously reported that by simply using shuttle plasmids that allow blue/white selection in Escherichia coli, it is possible to pre-select for positive clones. Here, we demonstrate that the same strategy can be used to assemble plasmids from several ectopic DNA fragments, which are all introduced in yeast cells by a simple transformation step. Further, to facilitate the subcloning of the fragment cloned into other targeting or expression vectors, the multi-cloning sites of three shuttle plasmids have been extended to include fifteen new restriction enzyme recognition sites.

[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Elsevier Science
Loading ...
Support Center