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首页> 外文OA文献 >Improved Repetitive-Element PCR Fingerprinting of Salmonella enterica with the Use of Extremely Elevated Annealing Temperatures
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Improved Repetitive-Element PCR Fingerprinting of Salmonella enterica with the Use of Extremely Elevated Annealing Temperatures

机译：使用极高的退火温度改进了肠炎沙门氏菌的重复元素PCR指纹图谱

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Modified thermal cycling conditions were explored in an effort to improve the reproducibility and resolving power of repetitive-element PCR (rep-PCR) fingerprinting. Assay performance was rigorously evaluated under standard and modified cycling conditions, using as a test set 12 strains putatively representing 12 serovars of Salmonella enterica. For all three fingerprint types (ERIC2, BOXA1R, and composite fingerprints), the use of extremely elevated annealing temperatures plus an initial “touchdown” cycling routine yielded significant improvements in day-to-day reproducibility and discriminating power despite the somewhat sparser appearance of the fingerprints. Modified cycling conditions markedly reduced the variability of fingerprints between cyclers, allowing fingerprints from different cyclers to be analyzed together without the degradation of assay performance that occurred with between-cycler analyses under standard cycling conditions. With modified cycling, composite fingerprints exhibited the lowest reproducibility but the highest net discriminating power of the three fingerprint types. rep-PCR fingerprints led to the discovery of a serotyping error involving one of the 12 test strains. These data demonstrate that modified cycling regimens that incorporate elevated annealing temperatures (with or without an initial touchdown routine) may markedly improve the performance of rep-PCR fingerprinting as a bacterial typing tool.
机译：为了改善重复元素PCR（rep-PCR）指纹图谱的可重复性和分辨能力，对改良的热循环条件进行了研究。在标准和改良的循环条件下，使用假定代表十二种肠炎沙门氏菌的12个菌株作为测试集，严格评估了测定性能。对于所有三种类型的指纹（ERIC2，BOXA1R和复合指纹），尽管外观略显稀疏，但使用极高的退火温度加上初始的“触地得分”循环程序仍可显着改善日常重现性和辨别力。指纹。修改后的循环条件显着降低了循环仪之间指纹的变异性，可以将来自不同循环仪的指纹一起进行分析，而不会降低标准循环条件下循环之间的分析性能。经过改进的循环，复合指纹在三种指纹类型中显示出最低的重现性，但最高的净辨别力。 rep-PCR指纹图谱导致发现涉及12个测试菌株之一的血清分型错误。这些数据表明，结合了较高的退火温度（有或没有初始接地程序）的改良循环方案可以显着改善rep-PCR指纹图谱作为细菌分型工具的性能。

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