通过共转化介导的同源重组将P1插入片段转移到酵母-细菌穿梭载体中。
Transfer of P1 inserts into a yeast-bacteria shuttle vector by co-transformation mediated homologous recombination.
作者信息
Criswell T L, Bradshaw S
机构信息
Department of Biological Sciences, PO Box 210006, University of Cincinnati, Cincinnati, OH 45221-0006, USA.
出版信息
Nucleic Acids Res. 1998 Aug 1;26(15):3611-3. doi: 10.1093/nar/26.15.3611.
Manipulation of genomic inserts cloned into the bacteriophage P1 vector is hindered by the large size of the inserts. We have used co-transformation mediated recombination between the yeast-bacteria shuttle vector, pClasper, and various P1 clones to transfer the entire insert from the P1 into pClasper. This results in the insert being stably maintained in yeast, facilitating mutagenesis by homologous recombination. The recombinant plasmid can subsequently be transferred to and stably maintained in bacteria for efficient plasmid preparation. This method can also be applied to inserts from P1 artificial chromosome or bacterial artificial chromosome vectors.
克隆到噬菌体P1载体中的基因组插入片段由于其较大的尺寸而难以操作。我们利用酵母-细菌穿梭载体pClasper与各种P1克隆之间的共转化介导重组，将整个插入片段从P1转移到pClasper中。这使得插入片段能够在酵母中稳定维持，便于通过同源重组进行诱变。随后，重组质粒可以转移到细菌中并稳定维持，以高效制备质粒。该方法也可应用于来自P1人工染色体或细菌人工染色体载体的插入片段。
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