Computational discovery and RT-PCR validation of novel Burkholderia conserved and Burkholderia pseudomallei unique sRNAs.
Computational discovery and RT-PCR validation of novel Burkholderia conserved and Burkholderia pseudomallei unique sRNAs.
复制标题
DOI：
10.1186/1471-2164-13-s7-s13
复制
发表时间：
2012
期刊：
影响因子：
4.4
通讯作者：
Firdaus-Raih M
中科院分区：
文献类型：
--
作者：
Khoo JS;Chai SF;Mohamed R;Nathan S;Firdaus-Raih M
已知细菌病原体的sRNAs参与各种细胞作用，包括环境适应以及毒力和致病性的调节。预计srna对假马利氏伯克霍尔德菌也可能具有类似的功能，伯克霍尔德菌是一种能够适应多种环境条件的土壤细菌，引起类鼻疽病，也能够感染多种宿主。通过将几个已证实的sRNA预测程序整合到计算管道中，筛选可用的伯克霍尔德氏菌基因组以确定sRNA候选基因。然后通过比较分析确定同源sRNA候选物。从总预测中，发现21个候选候选物具有Rfam同源物。RT-PCR和未知功能的候选sRNA基因测序显示，在正常培养条件下转录组中存在6个在伯克霍尔德氏菌中高度保守的sRNA基因，以及2个为假假杆菌所特有的sRNA基因。已验证的sRNA包括与蛋氨酸代谢调节相关的潜在顺式作用元件，以及一个预计与Hfq蛋白结合的假b杆菌特异性sRNA。本研究中开发的管道的使用和随后的比较分析已经成功地帮助发现和筛选sRNA候选基因进行验证。该方法鉴定了29个假假芽孢杆菌sRNA基因，其中21个具有Rfam同源基因，8个是新基因。
The sRNAs of bacterial pathogens are known to be involved in various cellular roles including environmental adaptation as well as regulation of virulence and pathogenicity. It is expected that sRNAs may also have similar functions for Burkholderia pseudomallei, a soil bacterium that can adapt to diverse environmental conditions, which causes the disease melioidosis and is also able to infect a wide variety of hosts. By integrating several proven sRNA prediction programs into a computational pipeline, available Burkholderia spp. genomes were screened to identify sRNA gene candidates. Orthologous sRNA candidates were then identified via comparative analysis. From the total prediction, 21 candidates were found to have Rfam homologs. RT-PCR and sequencing of candidate sRNA genes of unknown functions revealed six putative sRNAs which were highly conserved in Burkholderia spp. and two that were unique to B. pseudomallei present in a normal culture conditions transcriptome. The validated sRNAs include potential cis-acting elements associated with the modulation of methionine metabolism and one B. pseudomallei-specific sRNA that is expected to bind to the Hfq protein. The use of the pipeline developed in this study and subsequent comparative analysis have successfully aided in the discovery and shortlisting of sRNA gene candidates for validation. This integrated approach identified 29 B. pseudomallei sRNA genes - of which 21 have Rfam homologs and 8 are novel.