牛乳中嗜冷性杆菌的分离鉴定及PCR快速检测

论文编号:SW124  论文字数:9763,页数:18

摘　要：随着生活的改善，消费者对乳制品质量要求的日渐提高，乳制品中微生物的污染程度成为影响市场竞争的关键因素。牛乳在贮存过程中，大多数嗜冷菌，主要是假单胞菌属和一些杆菌属等，会产生耐受巴氏消毒甚至UHT处理的蛋白酶和脂肪酶，分解牛乳中的蛋白质和脂肪。分解产物将导致乳制品风味改变，品质降低，保质期缩短，对产品的形象和消费者的健康都造成了最大的危害。不同地区的嗜冷菌微生物种类和数量及分布规律有所差异，这种差异主要来自环境、设备及人员等。因此，要减少嗜冷菌的危害，早期对牛奶原料中嗜冷菌的检测和控制是关键。本论文通过牛肉膏蛋白胨培养基及CVT培养基分离纯化牛乳中的嗜冷菌，通过形态学观察和生理生化试验，初步确定其所在的菌属，根据菌属特异性基因序列设计PCR引物，提取基因组DNA进行PCR快速检测，最后进行测序比对确定具体菌属。
关键词：牛乳；嗜冷菌；分离纯化；生理生化试验；PCR

Abstract:　As living standards improved, consumers are much more concerned about the quality of dairy products. Microorganism contamination of dairy products has been a key factor in intense market competition. When lac vaccinum is in reserve, most psychrophilic bacteria, mainly Pseudomonas and Xanthomonas, produce proteinase and lipase which tolerate pasteurization even UHT curing, reducing protein and grease. Cleavage products will change special flavor, deteriorate quality, shorten deadline, heavily influence the quality of dairy products, which will damage the products image and consumer health. The species, quantity and distribution of psychrophilic bacteria are different in each area, and the difference lies in environment, equipment and staff. The control and detection of lac vaccinum in early stage is pivotal to reduce the hazard of psychrophilic bacteria. The text discusses the isolation and decontamination, physiological reaction and PCR detection of psychrophilic bacteria,and then according to the bacteria specificity gene sequence, we designed PCR primer,collected genome DNA for PCR rapid test, tested and contrasted sequence,then confirmed the bacterium.
Keywords:　Milk; Psychrophile;Isolating; Microbiological characteristics; PCR 

目录
1  绪论 4
 1.1  嗜冷菌概述 4
 1.2 分子生物学研究进展 6
2  实验部分 7
 2.1　嗜冷菌的分离鉴定 7
 2.2　生理生化试验 8
 2.3  嗜冷菌的DNA快速提取 10
 2.4　牛乳中嗜冷菌的PCR快速检测 11
⑶ PCR产物回收: 13
 3.1 菌种分离纯化 13
 3.2 生理生化实验结果 14
 3.3  提取DNA实验现象 14
 3.4  PCR快速检测、DNA回收及测序比对结果 15
3.5  实验结果与分析 16
4  总结与展望 17
致  谢 18

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