Gene disruption in Candida albicans using a synthetic, codon-optimised Cre-loxP system
Gene disruption in Candida albicans using a synthetic, codon-optimised Cre-loxP system
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DOI：
10.1016/j.fgb.2005.05.006
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发表时间：
2005-09-01
影响因子：
3
通讯作者：
Brown, AJP
中科院分区：
文献类型：
--
作者：
Dennison, PMJ;Ramsdale, M;Brown, AJP
由于缺乏可利用的性周期、二倍体的性质和非规范的遗传密码，白色念珠菌的分子工具箱的开发一直受到阻碍。我们描述了Cre-loxP位点特异性重组系统的适应性，作为一种有效和可控的破坏白色念珠菌基因的工具。我们已经通过干扰两个白色念珠菌基因座：ADE2和MET15来验证这个系统。Ade2和mat15缺失突变体使用loxP侧翼的ArG4和HIS1基断裂盒。然后使用合成的密码子优化的cre重组酶基因从白念珠菌基因组中解析出这些标记，效率接近100%。最后，构建了含有URA3、HIS1和ARG4标记的CIP质粒，用于标记和目的基因在对照菌株中的整合。该系统允许多个和顺序的遗传操作，这将有助于对白色念珠菌多基因家族的功能分析。(C)2005 Elsevier Inc.保留所有权利。
The development of the molecular toolbox for the fungal pathogen Candida albicans has been hampered by its lack of an exploitable sexual cycle, its diploid nature, and its non-canonical genetic code. We describe the adaptation of the Cre-loxP site-specific recombination system as a tool for the efficient and controlled disruption of C albicans genes. We have validated this system by disrupting two C. albicans loci: ADE2 and MET15. Ade2 and mat15 null mutants were made using loxP-flanked ARG4- and HIS1-based disruption cassettes. These markers were then resolved from the C albicans genome using a synthetic codon-optimised cre recombinase gene,with near 100% efficiency. Finally, CIp plasmids containing the URA3, HIS1, and ARG4 markers were generated for the reintegration of markers and target genes in control strains. This system allows multiple and sequential genetic manipulations, which will facilitate the functional analysis of multigene families in C albicans. (c) 2005 Elsevier Inc. All rights reserved.