Analysis Of Microbial Diversity Of Sayram Milk And Yogurt And Screening Of High-yield ?-Aminobutyric Acid Strains

Sayram yogurt is a traditional and popular handcraft set yogurt produced by Uighur peasant woman in baicheng county Xinjiang province of China.In the early time of the research group,it was found that Sayram yogurt contains a higher content of functional factor?-aminobutyric acid(GABA)than industrial yogurt products.It is speculated that the?-aminobutyric acid was metabolized by microorganisms.In this thesis,we used high-throughput sequencing to conduct metagenomic detection in sixty samples(thirty raw milk and thirty traditional fermented milk samples)from the Sayram town.Then,the microorganisms in the samples were cultured,and the dominant strains were selected,separated and purified by traditional culture-dependent method.The transformation solution of each dominant strains was prepared.Using thin-layer chromatography(TLC)to detect whether these strains can produce?-aminobutyric acid and high performance liquid chromatography(HPLC)to get the contents.Combining morphological,physiology,biochemistry experiments and sequence identification techniques for strain identification.The results were as follows:(1)Through high-throughput sequencing obtained total of 4,092,5761 high quality bacterial16 Sr RNA gene sequences and 4,056,820 fungal ITS high quality reads from the all samples,The results of experiments revealed the presence of 15 bacterial and 18 fungal phyla,comprising 218 bacterial and 495 fungal genera respectively,among all samples.Firmicutes and Proteobacteria were the predominant phyla in Sayram Ketteki and raw milk,Ascomycota was the dominant fungal phylum.Macrococcus and Lactobacillus were the dominated genera in raw milk and fermented milk respectively,Candida was the dominated fungal genus.The results showed that there were rich and diverse microbial species in Sayram milk and yogurt.The community structure and abundance of the two types of dairy products were quite different.(2)187 strains were isolated from the sixty samples using de Man,Rogosa and Sharpe(MRS),M17 and Yeast Extract Peptone Dextrose(YPD)medium.By the method of Thin Layer Chromatography,preliminary screening 8 strains had the ability to synthesized GABA,they were S8-42?S13-32?X26-42?S30-52?S33-32?X10-31?S20-31?X30-21.(3)Based on morphological characteristics experiment,the eight strains were identified as three yeasts and five lactic acid bacteria.Through the quantitative analysis by high performance liquid chromatography,three strains named S20-31?S33-32?and S30-52 and yields of 302.72,397.68 and 331.80 ?g/m L were selected.Combining traditional identification results and phylogenetic trees,the strain of S20-31,S30-52 and S33-32 were identified as Kluyveromyces marxianus,Enterococcus faecalis and Lactobacillus delbrueckii subspecies.Bulgaria.After screening and identification,L.derbergii subsp.Bulgaria S33-32 was obtained in this experiment with high yield GABA which provided a foundation for the next step to optimize GABA production and prepare a good starter.