Wang Shishan, Levin Robert E
Department of Food Science, Massachusetts Agricultural Experiment Station, University of Massachusetts, Amherst, 01003, USA.
Food Microbiol. 2006 Dec;23(8):757-61. doi: 10.1016/j.fm.2006.01.007. Epub 2006 Mar 29.
We used a rapid DNA extraction and purification method to obtain the DNA from Vibrio vulnificus seeded into clam tissue homogenates for real-time PCR quantification of the organism. Without enrichment, the limit of detection was 1 x 10(2) cfu/g of tissue with a linear detection range of 1 x 10(2) to 1 x 10(8) cfu/g. With a 5 h non-selective enrichment, the limit of detection was 1 cfu/g of tissue with a linear detection range of 1 to 1 x 10(6) cfu/g of tissue. We found a 10-fold higher detection limit with seeded clam tissue homogenates compared to pure culture in TSB(+). The detection limits with pure broth culture and seeded tissue homogenates were identical, 1 cfu/ml and 1 cfu/ml, respectively, following 5 h non-selective enrichment. However, the Ct value with tissue homogenates was about 3 threshold cycles higher than with pure culture.
我们采用一种快速DNA提取和纯化方法，从接种到蛤组织匀浆中的创伤弧菌中获取DNA，用于该菌的实时PCR定量分析。未经富集时，检测限为每克组织1×10² cfu，线性检测范围为每克组织1×10²至1×10⁸ cfu。经过5小时的非选择性富集后，检测限为每克组织1 cfu，线性检测范围为每克组织1至1×10⁶ cfu。我们发现，与TSB(+)中的纯培养物相比，接种蛤组织匀浆的检测限高10倍。经过5小时的非选择性富集后，纯肉汤培养物和接种组织匀浆的检测限相同，分别为1 cfu/ml和1 cfu/ml。然而，组织匀浆的Ct值比纯培养物高约3个阈值循环。
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