从奥尔平黏菌属菌株PC-2中分离富含AT的基因组DNA并分析编码碳水化合物降解酶的基因。
Isolation of extremely AT-rich genomic DNA and analysis of genes encoding carbohydrate-degrading enzymes from Orpinomyces sp. strain PC-2.
作者信息
Chen Huizhong, Hopper Sherryll L, Li Xin-Liang, Ljungdahl Lars G, Cerniglia Carl E
机构信息
Division of Microbiology, National Center for Toxicological Research, U.S. FDA, Jefferson, AR, 72079, USA.
出版信息
Curr Microbiol. 2006 Nov;53(5):396-400. doi: 10.1007/s00284-006-0098-2. Epub 2006 Oct 3.
Abstract
An effective method for extraction of intact genomic DNA from the extremely AT-rich polycentric anaerobic fungus Orpinomyces sp. strain PC-2 has been developed. This procedure involves removal of glycogen-like storage polysaccharides using hexadecyltrimethylammonium bromide (CTAB) and high salt washes. The DNA was digested with various restriction enzymes and was suitable for use as a PCR template, for Southern blotting, and for genomic library construction. Genomic DNA analysis of three representative genes (celE, bgl1, and xynA) encoding (hemi-) cellulolytic enzymes of the fungus revealed multiplicity of family 5 endocellulase genes (celE-like), and family 1 beta-glucosidase genes (bgl1-like), but only a single copy of family 11 xylanase gene (xynA).
摘要
已开发出一种从富含AT的多中心厌氧真菌奥皮诺霉菌属PC-2菌株中提取完整基因组DNA的有效方法。该方法包括使用十六烷基三甲基溴化铵（CTAB）和高盐洗涤去除糖原样储存多糖。DNA用各种限制性内切酶消化，适用于作为PCR模板、Southern印迹和基因组文库构建。对该真菌编码（半）纤维素分解酶的三个代表性基因（celE、bgl1和xynA）的基因组DNA分析表明，5家族内切纤维素酶基因（celE样）和1家族β-葡萄糖苷酶基因（bgl1样）具有多样性，但11家族木聚糖酶基因（xynA）只有单拷贝。
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