Kelley S T, Harris J K, Pace N R
Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder 80309, USA.
RNA. 2001 Sep;7(9):1310-6. doi: 10.1017/s1355838201010573.
DNA harvested directly from complex natural microbial communities by PCR has been successfully used to predict RNase P RNA structure, and can potentially provide an abundant source of information for structural predictions of other RNAs. In this study, we utilized genetic variation in natural communities to test and refine the secondary and tertiary structural model for the bacterial tmRNA. The variability of proposed tmRNA secondary structures in different organisms and the lack of any predicted tertiary structure suggested that further refinement of the tmRNA could be useful. To increase the phylogenetic representation of tmRNA sequences, and thereby provide additional data for statistical comparative analysis, we amplified, sequenced, and compared tmRNA sequences from natural microbial communities. Using primers designed from gamma proteobacterial sequences, we determined 44 new tmRNA sequences from a variety of environmental DNA samples. Covariation analyses of these sequences, along with sequences from cultured organisms, confirmed most of the proposed tmRNA model but also provided evidence for a new tertiary interaction. This approach of gathering sequence information from natural microbial communities seems generally applicable in RNA structural analysis.
通过聚合酶链式反应（PCR）直接从复杂的天然微生物群落中获取的DNA已成功用于预测核糖核酸酶P RNA的结构，并有可能为其他RNA的结构预测提供丰富的信息来源。在本研究中，我们利用天然群落中的遗传变异来测试和完善细菌转移信使核糖核酸（tmRNA）的二级和三级结构模型。不同生物体中tmRNA二级结构的变异性以及缺乏任何预测的三级结构表明，进一步完善tmRNA可能会有所帮助。为了增加tmRNA序列的系统发育代表性，从而为统计比较分析提供更多数据，我们对天然微生物群落中的tmRNA序列进行了扩增、测序和比较。使用从γ-变形菌序列设计的引物，我们从各种环境DNA样本中确定了44条新的tmRNA序列。对这些序列以及来自培养生物体的序列进行共变分析，证实了大多数提出的tmRNA模型，但也为一种新的三级相互作用提供了证据。这种从天然微生物群落收集序列信息的方法似乎普遍适用于RNA结构分析。
