应用DGGE和Real-time PCR分析酸菜发酵液中乳酸菌的动态变化

Dynamic changes of lactic acid bacteria in fermentation of Chinese sauerkraut by PCR-DGGE and Real-time PCR

目的使用DGGE和实时荧光定量PCR分析酸菜发酵液中乳酸菌的动态改变。方法采集传统天然发酵1~8周的酸菜发酵液50mL,抽提基因组DNA,使用DGGE对乳酸菌菌群进行多样性、相似性研究和优势菌的鉴定,实时荧光定量PCR测定乳酸菌含量。结果发酵周期加长,乳酸菌的含量随之也逐渐增大。清酒乳杆菌是酸菜自然发酵过程中的优势菌型,鼠李糖乳杆菌主要存在于发酵初期,发酵的中期(3~5周)、后期(6~8周)明显减少,同时发酵后期植物乳杆菌成为优势菌并完成全部发酵过程。发酵周期延长后,香农多样性指数和丰富度较高,出现先下降后上升的趋势,在第7周达到最大值。结论在DNA水平上,DGGE和实时定量PCR检测了酸菜发酵液中乳酸菌的含量,进行菌群结构的动态分析,明确优势菌型,为实现酸菜标准化生产提供理论依据。

Abstract： Objective To study the dynamic changes of lactic acid bacteria in the fermentation of Chinese sauerkraut by using Real-time PCR and DGGE. Methods Chinese sauerkraut fermentation broths which fermented for 1 to 8 weeks were collected. Quantitation analysis of lactic acid bacteria flora and analyses of diversity, similarity and sequence were carried out by using RT-PCR and DGGE respectively after DNA ex- traction. Results The content of lactic acid bacteria increased as the extension of fermentation. Lactobacil- lus sakei was the dominant bacterium. Lactobacillus rharnnosus existed in early fermentation period and re- duced in later fermentation period. Lactobacillus plantarurn became dominant bacterium in the later fermen- tation period. Shannon-Weaver indexes showed a trend of first increasing and then decreasing, which reached the maximum value at 7 weeks. Conclusion The content and dynamic changes of lactic acid bacteria were detected with DGGE and Real-time PCR at DNA level, and the dominant bacterium was verified. The result can provide the theoretical basis for Chinese sauerkraut fermentation process and standardization production.