Merryweather A, Bernander R, Nordström K
Department of Microbiology, University of Uppsala, Sweden.
Mol Gen Genet. 1987 Dec;210(3):381-4. doi: 10.1007/BF00327186.
Recombination is extensively used in order to move alleles between replicons. The exchange of wild-type chromosomal and mutant plasmid-borne alleles is a two-step process entailing the formation of a cointegrate between the entire plasmid and the chromosome, followed by resolution of such cointegrates to give a mutant chromosome and a plasmid carrying the wild-type chromosomal sequence. Often the cointegrate and the resolved forms cannot be distinguished phenotypically. To enable the direct isolation of the resolved products we have developed a positive selection technique. Cells containing a cointegrated plasmid R1 were constructed by transduction using a P1 lysate prepared from cells harbouring a plasmid comprising a mutant chromosomal allele and the so-called omega fragment which carries an aad (aminoglycoside adenylyltransferase) gene. P1 transduction from the cointegrate strain into an SmD recipient allowed direct selection for the resolved complex, since transduction of the aad gene is lethal to an SmD strain.
为了在复制子之间转移等位基因，重组被广泛应用。野生型染色体等位基因与突变体质粒携带的等位基因的交换是一个两步过程，需要在整个质粒和染色体之间形成共整合体，随后这种共整合体进行拆分，产生一条突变染色体和一个携带野生型染色体序列的质粒。通常，共整合体和拆分后的形式在表型上无法区分。为了能够直接分离拆分后的产物，我们开发了一种正向选择技术。通过使用从含有包含突变染色体等位基因的质粒和携带aad（氨基糖苷腺苷酸转移酶）基因的所谓ω片段的细胞制备的P1裂解物进行转导，构建了含有共整合质粒R1的细胞。从共整合菌株到SmD受体的P1转导允许直接选择拆分后的复合物，因为aad基因的转导对SmD菌株是致死的。
