双酶切法克隆肺炎支原体DNA的研究

STUDY ON CLONING OF MYCOPLASMA PNEUMONIAE DNA DIGESTED WITH DOUBLE RESTRICTION ENZYMES

用PstⅠ、EcoRⅠ两种限制性内切酶切割pUR222质粒DNA.经离心沉淀除去两酶切位点之间的小分子DNA片段,与肺炎支原体DNA重组,转化受体茵。用X-gal平板筛选出236株耐氨苄青霉素呈白色的菌落。经酶切图谱分析及DNA斑点杂交实验确证,这些菌株为含有肺炎支原体DNA片段的重组株。

Two restriction endonucleases, EcoR Ⅰ and Pst Ⅰ were used to digest pUR 222 plasmids and the small fragments produced by digestion were removed by centrifugation. Vectors were recombined with M. pneumonia cleaved with Pst Ⅰ and EcoR Ⅰ and the recombinated plasmids were transformed into recipient bacteria E. coli K12 BMH 71-18. 236 white ampicillin-resistant colonies were obtained with X-gal plate. It was confirmed that those colonies were recombinants containing M. pneumoniae DNA fragments by means of restriction enzymes digestion analysis and DNA dot blot hybridizatin.