利用质粒介导的对抗生素G418的抗性进行酵母突变体文库的快速筛选。
Use of plasmid-mediated resistance to the antibiotic G418 for the rapid screening of a yeast mutant library.
作者信息
Chenevert J M, Naumovski L, Dumas L B, Friedberg E C
机构信息
Department of Pathology, Stanford University, California 94305.
出版信息
J Exp Pathol. 1984 Fall;1(4):307-13.
Both haploid and diploid yeast cells are sensitive to the antibiotic G418. The former develop spontaneous resistance to G418 at a frequency of approximately 2 X 10(-6), however, the frequency of resistance in diploids is less than 1 X 10(-9) and was undetectable in these experiments. Mating of spontaneously resistant haploid cells to G418-sensitive strains yields sensitive diploids, indicating that spontaneous resistance to the antibiotic is a recessive trait. Both haploid and diploid cells can be efficiently transformed to G418 resistance by a plasmid carrying the Escherichia coli kanamycin resistance (kanr) marker. The ability to select for cells transformed with plasmids containing the kanr gene has facilitated the screening of a large series of temperature sensitive yeast mutants to determine whether any of them are allelic to RAD3.
单倍体和二倍体酵母细胞都对抗生素G418敏感。前者以约2×10⁻⁶的频率自发产生对G418的抗性，然而，二倍体中抗性频率小于1×10⁻⁹，在这些实验中无法检测到。对G418敏感的菌株与自发抗性单倍体细胞交配产生敏感二倍体，表明对抗生素的自发抗性是隐性性状。携带大肠杆菌卡那霉素抗性（kanr）标记的质粒可有效地将单倍体和二倍体细胞转化为对G418有抗性。选择用含有kanr基因的质粒转化的细胞的能力有助于筛选大量温度敏感酵母突变体，以确定它们中是否有任何一个与RAD3等位。
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