Danchin A, Sezer O, Glaser P, Chalon P, Caput D
Régulation de l'Expression Génétique, Institut Pasteur, Paris, France.
Gene. 1989 Aug 1;80(1):145-9. doi: 10.1016/0378-1119(89)90259-x.
Cloning of higher eukaryotic genes has seldom been performed by complementation of a defective prokaryotic function. This is especially true in the case of functions that are normally absent from the prokaryotic host. We demonstrate here that it is possible to identify by complementation the cDNA from mouse brain, which encodes calmodulin (CaM) synthesis, in spite of the fact that the recipient strain, Escherichia coli, does not normally harbour a CaM function. A three-component cloning procedure in which a gene product requiring CaM for activity, adenylate cyclase from the pathogen Bordetella pertussis, was used to screen a cDNA library for cAMP synthesis in E. coli. The nucleotide sequence of the corresponding cDNA is also reported.
高等真核生物基因的克隆很少通过缺陷原核功能的互补来进行。对于原核宿主中通常不存在的功能而言尤其如此。我们在此证明，尽管受体菌株大肠杆菌通常不具有钙调蛋白（CaM）功能，但仍有可能通过互补鉴定出小鼠脑中编码CaM合成的cDNA。采用了一种三组分克隆程序，其中利用一种需要CaM激活的基因产物——病原体百日咳博德特氏菌的腺苷酸环化酶，来筛选大肠杆菌中用于cAMP合成的cDNA文库。还报道了相应cDNA的核苷酸序列。
