限制性内切酶组合对阪崎克罗诺杆菌的脉冲场凝胶电泳基因分型效果的影响

Effect of multi-restriction enzymes on the genotyping effect of the pulsed-field gel electrophoresis scheme for Cronobacter sakazakii

目的利用多种限制性内切酶提高脉冲场凝胶电泳(pulsed-field gel electrophoresis,PFGE)技术对阪崎克罗诺杆菌(Cronobacter Sakazakii)的分型效果,找到更多的以PFGE为基础的阪崎克罗诺杆菌分子分型信息。方法本研究选用XbaⅠ、SpeⅠ、NheⅠ和BlnⅠ分别酶切34株克罗诺杆菌分离株的DNA,并经PFGE得到相应的基因图谱,通过束平均数、束菌株百分比、结与菌株比值与Simpson多样性指数对各限制内切酶以及结合酶的相对分辨率进行评价。结果 NheⅠ是PFGE单酶切分型中效果最好的一种限制性内切酶;4种限制性内切酶相结合的分型方法可提高PFGE阪崎克罗诺杆菌基因分型效果。结论运用PFGE技术对菌株进行亚分型时,选用的酶种类越多,则结果越可靠,使用XbaⅠ、SpeⅠ、NheⅠ、BlnⅠ4种内切酶结合分析,可以得到理想的菌株亚分型结果。

Objective To improve the typing effect of pulsed-field gel electrophoresis （PFGE） technology on Cronobacter Sakazakii, and find more information about the molecular typing of the PFGE-based Cronobacter Sakazakii by using a variety of restriction enzymes. Methods Xba Ⅰ, Spe Ⅰ, NheⅠ and BlnⅠ were selected for the restriction enzyme digestion of DNA of 34 strains of isolated Cronobacter sakazakii, and the corresponding genetic map was obtained by PFGE. The relative resolution of each restriction enzyme and the binding enzyme were evaluated by means of average number of bundles, the percentage of the bundles of the strains, the ratio of the strains and the ratio of the nodes to the strains and the Simpson diversity index. Results Nhe Ⅰ was one of the best in the restriction enzymes in PFGE type single enzyme segmentation. Four types of restriction enzymes binding couldimprove the genotyping effect of the PFGE. Conclusion When using PFGE technology to subdivide the strain, the more we select the variety of enzymes, the more the results are reliable. Using Xba Ⅰ, Spe Ⅰ, Nhe Ⅰ, Bln Ⅰ 4 kinds of enzyme combined with the analysis can get ideal strains and the classification results.