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首页> 外文期刊>Journal of Veterinary Science >A novel low-cost method for Mycobacterium avium subsp paratubercuiosis DNA extraction from an automated broth culture system for real-time PCR analysis
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A novel low-cost method for Mycobacterium avium subsp paratubercuiosis DNA extraction from an automated broth culture system for real-time PCR analysis

机译：一种新的低成本方法，用于自动肉汤培养系统进行实时PCR分析的自动肉汤培养系统促进分枝杆菌分枝杆菌抑制症DNA提取

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摘要

PCR is a highly accurate technique for confirming the presence of Mycobacterium avium subsp. paratuberculosis (Map) in broth culture. In this study, a simple, efficient, and low-cost method of harvesting DNA from Map cultured in liquid medium was developed. The proposed protocol (Universidad Austral de Chile [UACH]) was evaluated by comparing its performance to that of two traditional techniques (a QIAamp DNA Stool Mini Kit and cethyltrimethylammonium bromide [CTAB] method). The results were statistically assessed by agreement analysis for which differences in the number of cycles to positive (CP) were compared by Student's t-test for paired samples and regression analysis. Twelve out of 104 fecal pools cultured were positive. The final PCR results for 11 samples analyzed with the QIAamp and UACH methods or ones examined with the QIAamp and CTAB methods were in agreement. Complete (100%) agreement was observed between data from the CTAB and UACH methods. CP values for the UACH and CTAB techniques were not significantly different, while the UACH method yielded significantly lower CP values compared to the QIAamp kit. The proposed extraction method combines reliability and efficiency with simplicity and lower cost.
机译：PCR是一种高度准确的技术，用于确认厌氧亚副潜能子的存在。肉汤培养中的副植物抑制（地图）。在该研究中，开发了一种简单，有效的和从液体培养的地图中收获DNA的简单，有效和低成本的方法。通过将其性能与两种传统技术的性能与两种传统技术的性能进行比较来评估拟议的议定书（Universidad Austral De Chile [Uach]）。通过协议分析统计评估结果，循环循环次数差异的差异是通过学生的T检验进行比较进行配对样本和回归分析。培养的104个粪便池中的十二个是阳性的。与QIAAMP和UACH方法分析的11个样本的最终PCR结果或与QIAAMP和CTAB方法检查的样品进行了一致。从CTAB和UACH方法之间的数据之间观察到完整（100％）协议。 UACH和CTAB技术的CP值没有显着差异，而UACH方法与QIAAMP套件相比，CP值显着降低。所提出的提取方法以简单性和更低的成本结合了可靠性和效率。

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