中文摘要：

随机扩增多态性DNA （RAPD）作为一种分子标记技术,可用于多种病原菌的基因分型.为建立适合禽波氏杆菌RAPD分析的扩增体系,本试验以禽波氏杆菌基因组DNA为模板,对20条随机引物进行了筛选,并对RAPD-PCR反应体系中的重要参数设置梯度进行单因素试验分析,以确定最佳反应条件.经大量重复试验筛选到6条随机引物,确定25 μL的反应体系中模板DNA的量为50 ng,随机引物浓度为0.2 μmol/L,dNTPs的量为0.2 mmol/L,Mg2＋浓度为1 mmol/L,Taq酶为1U.该优化体系对22株禽波氏杆菌均能扩增出清晰稳定且多态性好的DNA指纹图谱,为后续的禽波氏杆菌RAPD基因分型奠定了良好的基础.

英文摘要：

As a molecular marker technique, random amplified polymorphie DNA （RAPD） can be used for the genotyping of a variety of pathogens. To establish a suitable RAPD reaction system for Bordetella avium, genomic DNAs of Bordetella avium were ex- tracted as templates and 20 random primers were screened. Several significant parameters of RAPD-PCR reaction system were stud- ied and optimized by setting gradient for single-factor analysis. Through a large number of repeated trials, 6 suitable random primers were found and a stable reaction system was established： 25 IxL reaction system containing 50 ng DNA template, 0.2 p.mol/L random primer, 0.2 mmol/L dNTPs, lmmol/L Mg^2＋ and 1U Taq DNA polymerase. It can generate clear, steady and polymorphie DNA frag- ment patterns for 22 Bordetella avium isolates which laid a good foundation for the subsequent RAPD genotyping.