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ID:26798961
大小:1.07 MB
页数:48页
时间:2018-11-29
《产CLA乳酸菌的筛选鉴定及LAI酶动力学分析》由会员上传分享,免费在线阅读,更多相关内容在学术论文-天天文库。
1、-摘要本试验围绕如何开发有保健功能的乳酸菌,以自家酿制的酸菜汁为试验原料,以传统分离细菌的方法,通过革兰氏染色初步鉴定后,通过PCR扩增16SrDNA片段并将之与T-载体连接转化后序列分析,确定菌种,结果表明该菌种为棒状乳杆菌,通过紫外法分析可知其产生的共轭亚油酸的含量0.0107mg/ml,共轭亚油酸转化率为2.51%。首先通过50%饱和硫酸铵对棒状乳杆菌亚油酸异构酶进行分离纯化,其回收率为65.57%,纯化倍数为1.15,再经80%饱和硫酸铵盐析其回收率为44.62%,纯化倍数为3.73,最后经过SephadexG-100凝胶过滤层析亚油酸异构酶的回收率为10
2、.32%,纯化倍数为8.36。对产共轭亚油酸棒状乳杆菌从培养温度,培养时间,pH值和接种量四个因素方面进行单因素实验,确定棒状乳杆菌产共轭亚油酸最佳培养温度为36℃,最佳培养时间为36h,最佳pH值为5,最佳接种量为4%;设计四因素三水平正交试验确定棒状乳杆菌产共轭亚油酸的最优组合为培养时间42h、起始pH值5.0、培养温度40℃、接种量4%,在该优化条件下共轭亚油酸含量为0.0375mg/mL,共轭亚油酸的转化率为8.8%,进一步提高了底物的转化率和产物得率,该结果为乳酸菌株在食品,医药,工业上的进一步应用奠定基础。关键词:乳酸菌;共轭亚油酸;分离纯化;培养条件
3、---StudiesonScreeningandidentificationoflacticacidbacteriaproducingCLAandtheenzymekineticsofLAIAbstractTheteststudiesonhowtodevelopaLacticacidbacteriawithhealthcarefunction fromfermentedChinesecabbageJuice.Thestrainwasisolatedbytraditionallaboratory methodsandwaspreliminarilyidentifie
4、dbytheGramstain.The16SrDNAdomain sequencewasamplifiedbyPCR,thenPCRprocuctswereligatedintotheTvector.After transformation,sequenceanalysissandstrainSpeciesIdentificationwerecarriedout.The resultsshowthatthestrainisLactobacilluscoryniformissubsp.torquensstrain30.The productionofconjugat
5、edlinoleicacidis0.0107mg/mlbyUVanalysis,andtheconversion rateofconjugatedlinoleicacidis2.51%.Firstly,LinoleateisomeraseofLactobacilluscoryniformiswereisolatedandpurified by50%saturatedammoniumsulfate,therecoveryrateis65.57%andthepurificationfold is1.15,thenitwasisolatedandpurifiedby80
6、%saturatedammoniumsulfate,therecovery rateis44.62%andThepurificationfoldis3.73.Finally,therecoveryrateis10.32%and thepurificationfoldis8.36bySephadexG-100gelfiltrationchromatography.Thefactorsofculturetemperature,culturetime,pH,andinoculumof Lactobacilluscoryniformiswereresearchedbysi
7、nglefactorexperiment.Theoptimum parametersforproductionofCLAwasdetermined:temperature36℃,time36h,pH5.0, inoculum4%.ByFourfactorsandthreelevelOrthogonaloptimaldesign,theoptimal parametersforproductionofCLAbyLactobacilluscoryniformiswasconfirmed: incubationtime42h,temperature40℃,initial
8、pH5.0
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