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基于16s rDNA的PCR快速鉴定啤酒腐败菌的研究 总被引:5,自引:0,他引:5
从Genebank中下载啤酒腐败菌所在属的63种细菌的16s rDNA序列,用DNAstar软件进行序列对比分析,找出这63种细菌16s rDNA序列所共有的5条同源序列,以这5条同源序列为基础设计出一对适合所有腐败菌的通用引物:上游:5’-GAACAGGATTAGATACCC-3’,下游:5’-GACTFAACCCAACATCTCAC-3’。利用该对引物,可以检测出啤酒厂出现的8种腐败菌,而对啤酒厂出现的13种非腐败菌则不能检出,保证了该引物对啤酒腐败菌的特异性。 相似文献
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根据乳酸菌16SrDNA序列的特征,设计合成了针对啤酒有害乳酸菌的通用引物,在16SrDNA基因水平上采用聚合酶链式反应(PCR)技术鉴定了啤酒中59株有害乳酸菌的种类。同时根据鉴定结果设计了8种乳酸菌的特异引物,PCR技术验证结果表明该8种特异引物能够准确鉴定乳酸菌。 相似文献
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传统筛种方法具有随机性、盲目性,而PCR筛菌,增加了目的性和效率.在基因序列数据库中查找益生双歧杆菌的16S rDNA共有序列,找出它们共有的特异性序列,设计了PCR引物.平板培养分离,初步筛选双歧杆菌菌落.根据细菌16S rDNA和23S rDNA两侧高度保守区域设计通用引物,提取菌落基因组DNA,扩增16S-23S rDNA间区序列,并送测序.通过序列同源性比对分析,鉴定、筛选到人体长双歧杆菌. 相似文献
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啤酒有害菌是一些能在啤酒中存活并使啤酒的外观和风味发生改变的细菌,对其进行快速检测和定量是啤酒生产急待解决的问题。我们从华润雪花啤酒(中国)有限公司各工厂提供的样品中分离到28株啤酒有害菌,16S rDNA序列的系统进化分析表明,其中26个菌株属于乳杆菌属(Lactobacillus spp.)、1个菌株为明串珠菌属(Leuconostoc spp.),1个菌株为片球菌属(Pediococcu sp.)。根据酒花(hop)抗性基因horA、horB和horC的保守序列设计了扩增这3个基因的PCR引物,用这些引物对28株啤酒有害菌进行了常规PCR检测,检出率分别为89%、79%和75%,用hor A—horC双引物进行检测,检出率为100%。用SYBR Green实时定量PCR技术,以horA基因为靶序列,建立了对啤酒有害菌的细胞数进行快速定量的新方法,用该方法测定的污染啤酒样品中有害菌的浓度与平板培养法相近。 相似文献
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通过筛选16S rDNA序列库,设计了一对双歧杆菌属特异性引物P175和P874在试验确定的反应条件下,该引物对能正确区分双歧杆菌和非双歧杆菌。RAPD技术用于种及菌株的鉴定。选用10种引物对9株标准菌株基因组DNA进行扩增,对其指纹图谱分析表明S 256引物扩增的图谱可于双歧杆菌菌种及菌株鉴定的依据。通过对图谱相似性进行聚类分析,揭示了双歧杆菌属基因组的多态性及其遗传发育关系。 相似文献
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利用分子生物学手段,以R.solanacearum染色体上的16S rDNA ITS以及毒性质粒携带的致病性相关基因fliC为靶点,分别设计5对序列特异性引物,筛选得到了特异性扩增16SrDNA ITS保守序列的引物对RaITS-1/2以及特异性扩增病菌fliC基因片段的引物对RalfliC-F/R.比较这两对引物的扩增灵敏度、稳定性和特异性可以发现,它们均能够稳定、快速、灵敏地检测青枯菌DNA,检测灵敏度可以达到10 fg DNA/μL.在此基础上成功构建了直接检测土壤青枯菌DNA的PCR检测技术体系. 相似文献
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Monitoring the lactic acid bacterial diversity during shochu fermentation by PCR-denaturing gradient gel electrophoresis 总被引:1,自引:0,他引:1
The presence of lactic acid bacteria (LAB) during shochu fermentation was monitored by PCR-denaturing gradient gel electrophoresis (DGGE) and by bacteriological culturing. No LAB were detected from fermented mashes by PCR-DGGE using a universal bacterial PCR primer set. However, PCR-DGGE using a new primer specific for the 16S rDNA of Lactococcus, Streptococcus, Tetragenococcus, Enterococcus, and Vagococcus and two primers specific for the 16S rDNA of Lactobacillus, Pediococcus, Leuconostoc, and Weissella revealed that Enterococcus faecium, Lactobacillus casei, Lactobacillus fermentum, Lactobacillus nagelii, Lactobacillus plantarum, Lactococcus lactis, Leuconostoc citreum, Leuconostoc mesenteroides, and Weissella cibaria inhabited in shochu mashes. It was also found that the LAB community composition during shochu fermentation changed after the main ingredient and water were added during the fermentation process. Therefore, we confirmed that PCR-DGGE using all three primers specific for groups of LAB together was well suited to the study of the LAB diversity in shochu mashes. The results of DGGE profiles were similar to the results of bacteriological culturing. In conclusion, LAB are present during shochu fermentation but not dominant. 相似文献
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为准确鉴定分离自我国四川鲜牦牛奶和曲拉中9株乳酸菌到种水平,作者运用16S rDNA序列分析、recA基因特异扩增和hsp60-RFLP等多种分子技术对分离自我国四川地区鲜牦牛奶和曲拉中的9株乳酸菌进行分类鉴定。结果证实,16S rDNA序列分析法可将9株乳酸菌初步归类为植物乳杆菌群(4株),肠膜明串珠菌(4株),瑞士乳杆菌(1株)。由于16S rDNA序列分析法不能区分植物乳杆菌和戊糖乳杆菌,为了进一步鉴定植物乳杆菌群中的4株菌,继续采用recA基因特异扩增和hsp60-RFLP技术对其细分,结果表明recA基因特异扩增和hsp60-RFLP的方法均能很好地把植物乳杆菌群中的4株菌鉴定到种水平,且均为植物乳杆菌。 相似文献
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为了解中华开菲尔微生物菌群的结构特征,本论文运用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术对开菲尔菌株发酵过程中微生物菌群的结构变化进行了实验分析,结果表明:细菌菌群DGGE图谱上出现有三种不同迁移位置的斑带,而酵母菌群DGGE图谱上只有一条斑带;经过DNA序列的对比分析可知:细菌菌群分别为肠膜明串珠菌(Leuconostoc mesenteroides)、马乳样乳杆菌(Lactobacillus kefiranofaciens)和开菲尔乳杆菌(Lactobacillus kefir),它们的序列同源性都达到100%;酵母菌群为德尔布有孢圆酵母(Torulaspora delbrueckii),其序列同源性为99%。本论文首次报道了德尔布有孢圆酵母在开菲尔菌落中的存在。 相似文献
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Gala E Landi S Solieri L Nocetti M Pulvirenti A Giudici P 《International journal of food microbiology》2008,125(3):347-351
The diversity of dominant lactic acid bacteria population in 12 months ripened Parmigiano Reggiano cheeses was investigated by a polyphasic approach including culture-dependent and independent methods. Traditional plating, isolation of LAB and identification by 16S rDNA analysis showed that strains belonging to Lactobacillus casei group were the most frequently isolated. Lactobacillus helveticus, Lactobacillus delbrueckii subsp. lactis, Lactobacillus parabuchneri, and Lactobacillus buchneri species were detected with lower frequency. PCR-denaturing gradient gel electrophoresis (DGGE) applied to DNA extracted directly from cheese samples and sequencing of rDNA amplicons confirmed the complex microbiological pattern of LAB in ripened Parmigiano Reggiano cheeses, with the significant exception of the Lactobacillus fermentum species, which dominated in several samples, but was not detected by cultivation. The present combination of different approaches can effectively describe the lactic acid bacteria population of Parmigiano Reggiano cheese in advanced stages of ripening, giving useful information for elucidating the role of LAB in determining the final cheese quality. 相似文献
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Identification and molecular characterisation of lactobacilli isolated from traditional Koopeh cheese 下载免费PDF全文
Hassan Hassanzadazar Karim Mardani Mohammad Yousefi Ali Ehsani 《International Journal of Dairy Technology》2017,70(4):556-561
This study was undertaken in order to phenotype and genotype wild‐type lactic acid bacteria isolated from Koopeh cheese of West Azerbaijan. Lactic acid bacteria (LAB) isolates were identified by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) and confirmed by phylogenetic analysis. Genotyping based on phylogenetic analysis of 16s rDNA gene revealed that LAB isolated from Koopeh cheese were mostly Lactobacillus plantarum as well as other species including Lactobacillus brevis, Entrococcus faecium and Enterococcus durans. It was concluded that a combined phenotypic and genotypic method could be used as a reliable technique for the identification and differentiation of LAB from dairy products. 相似文献
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目的构建珠江口水产养殖动物源的乳酸菌资源库,开发一套较为完善的高通量筛选优良乳酸菌的方法。方法采集珠江口水域多种水产动物的肠道,研磨,稀释涂布于含溴甲酚紫及碳酸钙的MRS平板上,挑取有溶钙圈的单菌落,进行过氧化氢酶实验以排除产气菌。提取细菌的基因组作模板, PCR扩增16S rDNA基因,根据测序结果进行同源性比对分析确定乳酸菌的种属,建立乳酸菌库。进一步借助96孔板培养技术,将乳酸菌与水产致病菌(指示菌)混合培养,根据对指示菌的抑菌效果,确定具有优良抑菌活性的乳酸菌。结果从水产动物肠道中筛选出乳酸菌14属71种1638株,获得了15株对常见水产养殖致病菌(嗜水气单胞菌、创伤弧菌、副溶血弧菌等)具有高效广谱抑制效果的菌株。结论本研究建立的乳酸菌菌种资源库,弥补了国内目前水产养殖来源乳酸菌菌株储备缺乏的现状,有望提供各类性能优良菌株用于渔业、畜牧业和食品生产。 相似文献
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为研究从驼骆瘤胃内容物分离、筛选的乳酸菌,其体外降解吲哚的功能。利用MRS培养基筛选骆驼瘤胃乳酸菌,提取细菌总DNA,扩增16S rDNA序列,构建系统发育树。在以吲哚(500 mg/L)为唯一碳源、氮源的无机盐培养基中胁迫培养乳酸菌,紫外分光光度计测定吸光度值(600 nm),测定乳酸菌在高浓度吲哚环境中的生长曲线;使用GC-MS动态检测吲哚残留量,测定乳酸菌对吲哚的降解能力。结果显示:筛选获得4株乳酸菌,GU366019、GU366027、GU366021、GU366034均归属为乳杆菌属(Lactobacillus),其中GU366019、GU366027、GU366021为德氏乳杆菌亚种(Lactobacillus delbrueckii subsp.),GU366034为驯马乳杆菌(Lactobacillus equicursoris)。四株菌在添加吲哚的无机盐培养基中生长缓慢,但对吲哚降解能力强。GU366027、GU366019、GU366021在144 h内,GU366034在156 h可将吲哚100%降解,证实来源于骆驼瘤胃的乳酸菌具有降解吲哚的功能。本实验在补充肠道毒素降解微生物资源的同时,也为乳酸菌降解、清除肠道毒素提供了研究依据。 相似文献