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Screening and characterization of a novel ruminal cellulase gene(Umcel-1) from a metagenomic library of gayal(Bos frontalis)

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Journal of Integrative Agriculture 2016年第4期15卷 855-861页

作者： LI Bi-feng ZHU Ya-xin GU Zhao-bing CHEN Yuan LENG Jing GOU Xiao FENG Li LI Qing XI Dong-mei MAO Hua-ming YANG Shu-Li Yunnan Provincial Key Laboratory of Animal Nutrition and Feed Science Yunnan Agricultural University Institute of Microbiology Chinese Academy of Sciences Department of Biological Science Huizhou University

gayal is a rare semi-wild bovine species found in the Indo-China. They can graze grasses, including bamboo leaves, as well as reeds and other plant species, and grow to higher mature live weights than Yunnan Yellow cattle maintained in similar harsh environments. The aim of this study was to identify specific cellulase in the gayal rumen. A metagenomic fosmid library was constructed using genomic DNA isolated from the ruminal contents of four adult gayals. This library contained 38400 clones with an average insert size of 35.5 kb. The Umcel-1 gene was isolated from this library. Investigation of the cellulase activity of 24 random clones led to the identification of the Umcel-1 gene, which exhibited the most potent cellulase activity. Sequencing the Umcel-1 gene revealed that it contained an open reading frame of 942 base pairs that encoded a product of 313 amino acids. The putative gene Umcel-1 product belonged to the glycosyl hydrolase family 5 and showed the highest homology to the cellulase （GenBank accession no. YP_004310852.1 ） from Clostridium lentocellum DSM 5427, with 44% identity and 62% similarity. The Umcel-1 gene was heterologously expressed in Escherichia coil BL21, and recombinant Umcel-1 was purified. The activity of purified recombinant Umcel-1 was assessed, and the results revealed that it hydrolyzed carboxymethyl cellulose with optimal activity at pH 5.5 and 45~C. To our knowledge, this study provides the first evidence for a cellulase produced by bacteria in gayal rumen.

关键词： gayal ruminal microorganism fosmid library Umcel-I gene gene expression cellulase activity

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Phylogenetic Relationships and Status Quo of Colonies for gayal Based on Analysis of Cytochrome b Gene Partial Sequences

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Journal of Genetics and Genomics 2007年第5期34卷 413-419页

作者：马国龙常洪李世平陈宏宇冀德君耿荣庆常春芳李永红扬州大学动物科学与技术学院扬州225009

Thirty-three mutations and four different haplotypes were found when cytochrome b（Cytb） gene partial sequences of 12 gayals were analyzed. Together with sequences of Bos indicus, Bos taurus, Bos grunniens, and Bos gaurus with Bubalus bubalis as the out group, the partial sequences of Cytb gene of gayals were aligned and base composition and nucleotide variation of Cytb gene were analyzed. The phylogenetic trees were constructed by the NJ method and the MP method respectively, both supporting almost the same topology. gayal is an independent species of Bos from Bos indicus, Bos taurus, and Bos gaurus. The results also indicate that a great proportion of gayal bloodline was invaded by other species, and the protection of gayal is facing a formidable situation.

关键词： gayal cytochrome b gene phylogenetic relationship status quo of colony

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