McKnight G L, McConaughy B L
Proc Natl Acad Sci U S A. 1983 Jul;80(14):4412-6. doi: 10.1073/pnas.80.14.4412.
Yeast cDNA was prepared in a yeast expression plasmid to generate a cDNA plasmid pool composed of approximately 40,000 members. Several yeast mutants were transformed with the cDNA plasmid pool, and the cDNAs for ADC1, HIS3, URA3, and ASP5 were isolated by functional complementation. Restriction enzyme analysis confirmed the genetic identity of the ADC1, HIS3, and URA3 cDNAs and demonstrated that the URA3 cDNA contains 5' noncoding sequences. The relative abundance of the various cDNAs in the cDNA plasmid pool paralleled the abundance of the mRNAs in total poly(A)+ RNA, which ranged from approximately 0.01% to 1%. The utility of this approach to isolate rare cDNAs from higher eukaryotes is discussed.
酵母cDNA在酵母表达质粒中制备，以生成一个由约40,000个成员组成的cDNA质粒库。用该cDNA质粒库转化了几个酵母突变体，并通过功能互补分离出了ADC1、HIS3、URA3和ASP5的cDNA。限制性酶切分析证实了ADC1、HIS3和URA3 cDNA的基因同一性，并表明URA3 cDNA包含5'非编码序列。cDNA质粒库中各种cDNA的相对丰度与总聚腺苷酸加尾RNA中mRNA的丰度平行，其范围约为0.01%至1%。讨论了这种从高等真核生物中分离稀有cDNA的方法的实用性。
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