开发一套用于致病性酵母近平滑念珠菌基因操作的质粒载体。
Development of a set of plasmid vectors for genetic manipulations of the pathogenic yeast Candida parapsilosis.
作者信息
Kosa Peter, Gavenciakova Barbora, Nosek Jozef
机构信息
Department of Biochemistry and Genetics, Faculty of Natural Sciences, Comenius University, Mlynska dolina CH-1, 842 15 Bratislava, Slovak Republic.
出版信息
Gene. 2007 Jul 15;396(2):338-45. doi: 10.1016/j.gene.2007.04.008. Epub 2007 Apr 18.
Abstract
A system for genetic transformation of the yeast Candida parapsilosis, recently developed in our laboratory, opened a venue for investigation of this pathogenic species at the molecular level. In this study we extend the range of available experimental tools by construction of a genomic DNA library suitable for screening and isolation of genes by functional complementation of yeast mutants and a set of replicative plasmid vectors for genetic manipulation of C. parapsilosis cells. The plasmids are based on auxotrophic (CpGAL1, CpURA3, CpMET2, CpLYS4) and dominant (CaIMH3) selection markers. In addition, we constructed plasmid derivatives containing reporter genes yEGFP3 and KlLAC4 coding for enhanced version of the green fluorescent protein and Kluyveromyces lactis beta-galactosidase, respectively. The vectors facilitate propagation and expression of cloned genes in C. parapsilosis cells and allow intracellular localization of gene products and/or monitoring the activity of promoter sequences.
摘要
我们实验室最近开发的一种用于近平滑念珠菌基因转化的系统，为在分子水平上研究这种致病物种开辟了一条途径。在本研究中，我们通过构建一个适合通过酵母突变体功能互补筛选和分离基因的基因组DNA文库以及一套用于近平滑念珠菌细胞基因操作的复制性质粒载体，扩展了可用实验工具的范围。这些质粒基于营养缺陷型（CpGAL1、CpURA3、CpMET2、CpLYS4）和显性（CaIMH3）选择标记。此外，我们构建了含有报告基因yEGFP3和KlLAC4的质粒衍生物，它们分别编码绿色荧光蛋白的增强版本和乳酸克鲁维酵母β-半乳糖苷酶。这些载体有助于克隆基因在近平滑念珠菌细胞中的繁殖和表达，并允许基因产物的细胞内定位和/或监测启动子序列的活性。
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