Department of Genetics, Biology of Microorganisms, Anthropology, Evolution, University of Parma, Viale Usberti 11/A, 43100 Parma, Italy.
J Biotechnol. 2009 Sep 25;143(4):247-54. doi: 10.1016/j.jbiotec.2009.08.007. Epub 2009 Aug 14.
A set of shuttle yeast vectors containing the dominant selectable markers KanMX4 or HphMX4 cassettes, conferring resistance to geneticin and hygromycin B, respectively, was constructed. Dominant selectable markers are useful for genetic manipulation of natural, wine and industrial strains which do not contain any auxotrophic markers as well as of strains which cannot grow on synthetic mineral medium. Vectors were characterized by (i) copy number, (ii) mitotic stability both in selective and non-selective conditions, (iii) the efficiency and frequency of transformations, (iv) optimal adaptation times in non-selective media, (v) optimum conditions for transformation of various laboratory, commercial and wine strains, and (vi) expression level of an inserted gene. Furthermore we produced GFP-containing vectors that can be used for protein subcellular localization in prototrophic strains.
构建了一套包含显性选择标记物 KanMX4 或 HphMX4 盒的穿梭酵母载体，分别赋予遗传霉素和潮霉素 B 的抗性。显性选择标记物可用于遗传操作天然、葡萄酒和工业菌株，这些菌株不含任何营养缺陷型标记物，也可用于不能在合成矿物培养基中生长的菌株。载体的特征在于：（i）拷贝数，（ii）有丝分裂稳定性，无论是在选择性条件下还是非选择性条件下，（iii）转化的效率和频率，（iv）在非选择性培养基中的最佳适应时间，（v）各种实验室、商业和葡萄酒菌株转化的最佳条件，以及（vi）插入基因的表达水平。此外，我们还生产了含有 GFP 的载体，可用于在营养缺陷型菌株中进行蛋白质亚细胞定位。
