Roberts M A, Crawford D L
Department of Microbiology, Molecular Biology, and Biochemistry, University of Idaho, Moscow, Idaho 83844-3052, USA.
Appl Environ Microbiol. 2000 Jun;66(6):2555-64. doi: 10.1128/AEM.66.6.2555-2564.2000.
We have analyzed 20 randomly amplified polymorphic DNA (RAPD) primers against 36 Streptomyces strains, including 17 taxonomically undefined strains, 25 nonstreptomycete actinomycetes, and 12 outgroups consisting of gram-positive and -negative species. Most of the primers were useful in identifying unique DNA polymorphisms of all strains tested. We have used RAPD techniques to develop a genus-specific probe, one not necessarily targeting the ribosomal gene, for Streptomyces, and a strain-specific probe for the biological control agent Streptomyces lydicus WYEC108. In the course of these investigations, small-scale DNA isolations were also developed for efficiently isolating actinomycete DNA. Various modifications of isolation procedures for soil DNA were compared, and the reliability and specificity of the RAPD methodology were tested by specifically detecting the S. lydicus WYEC108 in DNA isolated from soil.
我们针对36株链霉菌菌株分析了20种随机扩增多态性DNA(RAPD)引物,这些菌株包括17株分类学上未定义的菌株、25株非链霉菌放线菌以及12株由革兰氏阳性和阴性物种组成的外群。大多数引物可用于鉴定所有测试菌株独特的DNA多态性。我们利用RAPD技术开发了一种针对链霉菌的属特异性探针(不一定靶向核糖体基因)以及针对生防菌利迪链霉菌WYEC108的菌株特异性探针。在这些研究过程中,还开发了小规模DNA分离方法以高效分离放线菌DNA。比较了土壤DNA分离程序的各种改进方法,并通过特异性检测从土壤中分离出的DNA中的利迪链霉菌WYEC108来测试RAPD方法的可靠性和特异性。