變性高效液相色譜技術(shù)在快速檢測(cè)食品微生物中的應(yīng)用研究.pdf

1、安徽農(nóng)業(yè)大學(xué)碩士學(xué)位論文變性高效液相色譜技術(shù)在快速檢測(cè)食品微生物中的應(yīng)用研究姓名：楊大偉申請(qǐng)學(xué)位級(jí)別：碩士專業(yè)：農(nóng)產(chǎn)品貯藏與加工指導(dǎo)教師：周裔彬;劉云國2011-06Ⅱ Abstract In this paper, based on the molecular biology technology, The flatoxigenic bacteria of Aspergillus, candida albicans, Clostrid

2、ium botulinum-A ， Bacillus coagulans, Lactobacillus fermenti were selected to establish a rapid detection method by denaturing high performance liquid chromatography (DHPLC) and polymerase chain reaction (PCR). Our aim

3、is to offer theoretical basis and scientific support for quick testing microorganisms in food. The main results are as follows after investigation: 1. The technologies of PCR and DHPLC were applied to provide a method fo

4、r detection flatoxigenic bacteria in food rapidly. AflR gene from a coded aflatoxin biosynthesis was taken as the target design primer. The PCR system causing aflatoxin could rapidly be recognized by the method establis

5、hed and optimized. The amplification segment was 184 bp. The result showed that specification and sensitivity were better. The minimum detection limit could reach 100 cfu/ml. The 75 samples from different food were analy

6、zed and identified two positive samples, those results were consistent with those of old methods and PCR applied. 2. The technologies of PCR and DHPLC were applied to provide a method for candida albicans detected in

7、food rapidly. The unique target sequence acid protease gene SAP6 from candida albicans was selected as the specific design primer, that is, the primer were respectively F-5`- CTGGGTCTTCTGATTTGTGG -3` and R-5`- CTGG

8、TAGCTTCGTTGGTTTG -3`, and the amplification segment was 307 bp. PCR system was established and optimized. Our results indicated that the specification was better, and the limit of sensitivity reached 1.0×103 cfu/ml.

9、 The candida albicans in milk powder could be rapidly and truly determined. 3. The technologies of PCR and DHPLC were applied to provide a method for clostridium botulinum-A determined in food rapidly. The A type-botuli

10、num neurotoxin gene from clostridium botulinum-A was selected as the specific primer. Twenty-three strains from non- clostridium botulinum-A and clostridium perfringens were considered as a control for specific analysis,

11、 and the DNA template was diluted into different gradient to analyze the sensitivity. The result showed that the primer had a better specification and sensitivity, and lowest detectable limit could achieve for110 ng/tube