Study On The Microbial Diversity In Tianshan Permafrost Cold-adapted Strain And The Phylogenetic Analysis Of Cold-adapted Lipase Producing Strain

Lipases is one of the most important enzyme for industry, which is widely used in several vocation like food industry, detergent industry, medicine industry et al. Cold-adapted lipases is having extensive application prospect in the directions of biological engineering and energy source saving because of its high catalytic activity and temperature sensitive. Generally, cold-adapted lipases is product by the cold-adapted microorganisms, consequently, the screening of cold-adapted lipases-producing strains is the most important and fundamental factor to solve the source of cold-adapted lipases. Frozen soil which called the resource-treasure house of cold-adapted microorganisms, is useful for us to study cold-adapted microorganisms and their cold active enzymes. Thus, isolation and purification of cold-adapted microorganisms from alpine frozen soils in the Tianshan Mountains is having great significance for screening the high-quality and high-activity cold-adapted enzymes-producing strains.Based on the unique geographical environment in Xinjiang, isolated by pure culture at low temperature frozen soil microbial diversity and sequencing of their system, and using technology to build metagenomics microbial library Tianshan frozen. Isolated in pure culture from the low-temperature micro-organisms were screened, purified lipase producing strain of low temperature to resolve the problem of lipase enzyme source, the screening of strain identification and analysis of lipase phylogenetic diversity of bacteria for the low-temperature lipase Construction and Engineering establish some industrial application of theory and practice.1 By dilution plate method can be a total of 76 strains isolated cultures, (65.2)% mainly Gram-positive bacteria, and most Nengchan pigment (68.4)%, colony color was pink, lemon yellow, white, etc., isolated low-temperature optimum growth temperature of microorganisms 20-24℃, the vast majority are resistant to psychrophilic bacteria. The 16S rDNA sequence analysis, the selected representative strains of 38 strains of the phylogenetic analysis, these strains in the development of low temperature cultivation belong to the treeα-proteobacteria subclass,β-proteobacteria subclass,γ-proteobacteria subclass, high G+C Gram-positive bacteria (Actinobacteria), low G+C Gram-positive bacteria (Firmicute), CFB (Cytophaga-Flavobacterium-Bacteroides) 6 phylogenetic groups, includingα-,β,γ-proteobacteria was the dominant flora. Pseudomonas dominant species is the largest proportion of total bacteria, the middle of frozen low-temperature microbial diversity can contribute to develop the most, followed by Rhodococcus and Brevundimonas. This study shows that permafrost is the Tianshan Mountains in the natural products of the culture collections, which can nurture the rich diversity of microorganisms at low temperature.2 Using technology to build metagenomics microbial library of Tianshan permafrost temperature, and the positive clones by the bacterium PCR, restriction enzyme digestion for detecting gene sequence obtained 367 sequences, whereα-proteobacteria subclass a total of 95,β-proteobacteria subclass, y-proteobacteria subclass 85 amended the high G+C Gram-positive bacteria (Actinobacteria) 102 amended the low G+C Gram-positive bacteria (Firmicute), CFB (Cytophaga-Flavobacterium-Bacteroides) 45 bar. Library is the largest gates in the deformation of the evolution of bacteria clusters, and a-proteobacteria subclass, y-proteobacteria subclass larger proportion,25.9% and 23.1%, followed by actinomycetes Mun Oasis, sub-high G+C Gram-positive bacteria (Actinobacteria) and low G+C Gram-positive bacteria (Firmicute), the largest subclass of the total number of sequences 27.8%. CFB (Cytophaga-Flavobacterium-Bacteroides) cluster of the 16S rDNA sequence evolution of 45, accounting for 12.3% of the total, is the smallest of the development of gene library spectrum. Studies have shown that molecular methods have gained more than the artificial culture of genetic information, diversity is more abundant, and can more accurately reflect the permafrost in the Tianshan Mountains, The distribution of microorganisms at low temperature.3 Of the totally 17 bacterial stains exhibiting cold-adapted lipase activity, we found that only 8 stains were able to hydrolyze olive oil. Enzyme producing strains from most of the optimal growth temperature around 24℃, growth is basically in the temperature range 4-37℃, bacteria are resistant to cold areas, most of the salt is slightly better low-temperature bacterial. Lipase produced 17 strains belong to a-proteobacteria subclass,β-proteobacteria subclass,γ-proteobacteria, high G+C Gram-positive bacteria (Actinobacteria), CFB (Cytophaga-Flavobacterium-Bacteroides) 5 phylogenetic groups, including high G+C Gram-positive bacteria (Actinobacteria) in the genus most, Aeromonas genus(Pseudomonas spp.) Largest share of the dominant genus of permafrost temperature lipase diversity of the middle class contributed most, followed by Chryseobacterium, Rhodococcus and Brevundimonas. Thus, permafrost in the Tianshan Mountains, bears a very rich and diverse microbial lipase production of low temperature resources. Tianshan Mountains is the ideal enzyme source.