基因组大小和rrn基因拷贝数对从细菌物种混合物中PCR扩增16S rRNA基因的影响。
Effect of genome size and rrn gene copy number on PCR amplification of 16S rRNA genes from a mixture of bacterial species.
作者信息
Farrelly V, Rainey F A, Stackebrandt E
机构信息
DSM-German Collection of Microorganisms and Cell Cultures GmbH, Braunschweig.
出版信息
Appl Environ Microbiol. 1995 Jul;61(7):2798-801. doi: 10.1128/aem.61.7.2798-2801.1995.
Abstract
In order to assess the effect of genome size and number of 16S rRNA genes (rDNAs) on the quantities of PCR-generated partial 16S rDNA fragments, equimolar amounts of DNA from pairs of different species for which these parameters are known were subjected to gene amplification. The experimentally determined ratio of PCR products obtained, as determined by image analysis of SYBR-Green I-stained amplification products, corresponded well with the predicted ratio calculated from the number of rrn genes per equimolar amounts of DNA in mixtures of Escherichia coli and "Thermus thermophilus" and of Pseudomonas aeruginosa and "T. thermophilus." The values for the pair of Bacillus subtilis and "T. thermophilus" showed greater deviations from the predicted value. The dependence of the amount of 16S rDNA amplification product on these two parameters makes it impossible to quantify the number of species represented in 16S rDNA clone libraries of environmental samples as long as these two parameters are unknown for the species present.
摘要
为了评估基因组大小和16S rRNA基因（rDNA）数量对PCR产生的部分16S rDNA片段数量的影响，对已知这些参数的不同物种对的等摩尔量DNA进行基因扩增。通过对SYBR - Green I染色的扩增产物进行图像分析确定的实验获得的PCR产物比例，与根据大肠杆菌和“嗜热栖热菌”混合物以及铜绿假单胞菌和“嗜热栖热菌”混合物中等摩尔量DNA的rrn基因数量计算出的预测比例非常吻合。枯草芽孢杆菌和“嗜热栖热菌”这一对的值与预测值的偏差更大。只要环境样品中存在的物种的这两个参数未知，16S rDNA扩增产物的量对这两个参数的依赖性就使得无法对16S rDNA克隆文库中所代表的物种数量进行定量。
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