Tandeau de Marsac N, Borrias W E, Kuhlemeier C J, Castets A M, van Arkel G A, van den Hondel C A
Gene. 1982 Nov;20(1):111-9. doi: 10.1016/0378-1119(82)90092-0.
A new strategy for molecular cloning in the cyanobacterium Anacystis nidulans R-2 is described. This strategy involved the use of a transposon and was developed for the cloning of a gene encoding methionine biosynthesis. A met::Tn901 mutant was isolated. Chromosomal DNA fragments were cloned in the Escherichia coli plasmid vector pACYC184. A recombinant plasmid carrying the inactivated met::Tn901 gene was selected after transformation to E. coli. The cloned met::Tn901 DNA fragment was used as a probe to select the corresponding A. nidulans R-2 wild-type met gene from a gene library prepared in E. coli, using the newly constructed shuttle cosmid vector pPUC29. When transformed into A. nidulans Met- mutants, this cloned gene allowed the mutants to grow prototrophically.
本文描述了一种在蓝藻念珠藻（Anacystis nidulans）R-2中进行分子克隆的新策略。该策略涉及使用转座子，并且是为克隆编码甲硫氨酸生物合成的基因而开发的。分离出了一个met::Tn901突变体。将染色体DNA片段克隆到大肠杆菌质粒载体pACYC184中。转化到大肠杆菌后，选择携带失活的met::Tn901基因的重组质粒。使用新构建的穿梭粘粒载体pPUC29，将克隆的met::Tn901 DNA片段用作探针，从在大肠杆菌中制备的基因文库中筛选相应的念珠藻R-2野生型met基因。当将该克隆基因转化到念珠藻Met-突变体中时，它能使突变体以原养型生长。
