𝔖 Bobbio Scriptorium
✦   LIBER   ✦

Heterologous URA3MX cassettes for gene replacement inSaccharomyces cerevisiae

✍ Scribed by Goldstein, Alan L.; Pan, Xuewen; McCusker, John H.


Book ID
101224069
Publisher
John Wiley and Sons
Year
1999
Tongue
English
Weight
162 KB
Volume
15
Category
Article
ISSN
0749-503X

No coin nor oath required. For personal study only.

✦ Synopsis


Heterologous gene replacement cassettes are powerful tools for dissecting gene function in Saccharomyces cerevisiae. Their primary advantages over homologous gene replacement cassettes include reduced gene conversion (leading to efficient site-specific integration of the cassette) and greater independence of strain background. Perhaps the most widely used cassettes are the MX cassettes containing the dominant selectable kanamycin resistance gene (kan r ), which confers resistance to G418 (Wach et al., 1994). One limitation of the kanMX cassettes is that they are not counterselectable and therefore not readily recyclable, which is important when constructing strains with more than one gene deletion. To address this limitation, and to expand the choices of heterologous markers, we have created two new MX cassettes by replacing the kan r ORF from plasmids pFA6-kanMX3 and pFA6-kanMX4 with the Candida albicans URA3 ORF. These plasmids, pAG60 (CaURA3MX4) and pAG61 (CaURA3MX3) are identical to the kanMX cassettes in all other respects but have the added advantage of being counterselectable and therefore readily recyclable in S. cerevisiae.


πŸ“œ SIMILAR VOLUMES