摘要
用玻璃珠法、溶菌酶法、冻融法、氯化苄(phCH2Cl)法提取啤酒发酵液、生产用啤酒酵母(Saccharomyces cerevisiae)、K氏酵母、啤酒生产污染细菌(T)、G-细菌(E.coli.DH5α)和G+细菌(BacillusYK-1R)的基因组DNA。溶菌酶法和冻融法对细菌的裂解率为99%～100%,对酵母的裂解率只有15%～28%;玻璃珠法对酵母的裂解率为92%～94%;phCH2Cl法对细菌和酵母的裂解率均为100%。对提取的啤酒发酵液基因组DNA浓度进行测定,结果为:玻璃珠法15.4g/ml、溶菌酶法18.0g/ml、冻融法13.8g/ml、phCH2Cl法40.7g/ml。电泳检测结果表明,除冻融法外,其余3种方法得到的混合菌DNA片段均大于23kb,无拖尾、无蛋白污染。phCH2Cl法图谱更清晰,DNA得率108μg/g湿菌体,RAPD-PCR图谱重复性好。为用基因诊断技术在线检测啤酒发酵过程中微生物污染,提供了一种可靠的DNA提取方法。
Genome DNA was respectively extracted from fermented beer liquid, beer yeast （S.cerevisiae）, yeast（K）, beercontaminating bacteria（T）, E.coli.DH5 α and Bacillus YK- 1R by various methods to produce different cell-disruption efficacy.Glass bead-beating method has the yeast cells lysis efficacy beyond 90%. Although the lysozyme or freeze-thaw method shows that bacteria lysis efficacy can reach 99%- 100%, yet can only reach 15%-28% for yeast cells lysis efficacy. However the use of phCH2Cl has lysis efficacy for both bacterial and yeast cells reaching 100%. The genome DNA extracted by either glass beads, or lysozyme or phCH2Cl reaches beyond 23kb in DNA size and good integrity when assayed in gel electrophoresis. The genome DNA extracted through phCH2Cl is the most clear and sharp figure, DNA yield can achieve 108μg/g of wet-organism and the RAPD- PCR is reproducible. This protocol has potential for detection of beer-spoilage microorganisms.
出处
《食品科学》
EI
CAS
CSCD
北大核心
2007年第3期191-194,共4页
Food Science
基金
山西省科技攻关计划项目(041147)
