Transcriptome profile of a bovine respiratory disease pathogen: Mannheimia haemolytica PHL213.
Transcriptome profile of a bovine respiratory disease pathogen: Mannheimia haemolytica PHL213.
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DOI：
10.1186/1471-2105-13-s15-s4
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发表时间：
2012
影响因子：
3
通讯作者：
Nanduri B
中科院分区：
文献类型：
--
作者：
Reddy JS;Kumar R;Watt JM;Lawrence ML;Burgess SC;Nanduri B
用于结构基因注释的计算方法已经推动了基因发现，但是在原核基因组注释方面面临某些缺点。使用这些方法识别转录起始位点、划分重叠基因边界和识别调控元件（例如小RNA）是不准确的。在这项研究中，我们重新访问的结构注释溶血性曼氏菌PHL 213，牛呼吸道疾病病原体。M.溶血性病原体是牛呼吸道疾病的病原体之一，该疾病每年给养牛业造成约30亿美元的损失。我们使用RNA-Seq，并使用免费提供的计算方法和资源分析数据。其目的是使用基于RNA-Seq的表达谱来识别基因组中以前未注释的区域，以补充该病原体的现有注释。使用Illumina基因组分析仪，我们生成了9，055，826个读段（平均长度约76 bp），并使用Bowtie将它们与参考基因组进行比对。使用SAMTOOLS和定制Perl脚本结合BLAST搜索和可用的基因注释信息分析转录区域。单核苷酸分辨率图谱能够鉴定14个新的蛋白质编码区以及44个潜在的新sRNA。基础转录谱显示，2，837个注释区域中的2，506个在体外表达，覆盖率为95.25%，代表了基因组中所有广泛的功能基因类别。表达谱还帮助鉴定了518个潜在的操纵子结构，涉及1，086个共表达对。我们还鉴定了11种具有突变/交替起始密码子的蛋白质。基于RNA-Seq的转录组分析在结构基因注释中的应用有助于纠正现有的注释错误，并识别潜在的新蛋白质编码区和sRNA。我们使用计算工具来预测与新表达区域相关的调控元件，例如启动子和终止子，以进一步表征这些新的功能元件。我们的研究基于实验证据补充了溶血曼海姆菌PHL 213的现有结构注释。鉴于sRNA在毒力基因调控和应激反应中的作用，本研究中描述的潜在新sRNA可以为未来的研究提供框架，以确定sRNA在M.溶血性致病
Computational methods for structural gene annotation have propelled gene discovery but face certain drawbacks with regards to prokaryotic genome annotation. Identification of transcriptional start sites, demarcating overlapping gene boundaries, and identifying regulatory elements such as small RNA are not accurate using these approaches. In this study, we re-visit the structural annotation of Mannheimia haemolytica PHL213, a bovine respiratory disease pathogen. M. haemolytica is one of the causative agents of bovine respiratory disease that results in about $3 billion annual losses to the cattle industry. We used RNA-Seq and analyzed the data using freely-available computational methods and resources. The aim was to identify previously unannotated regions of the genome using RNA-Seq based expression profile to complement the existing annotation of this pathogen. Using the Illumina Genome Analyzer, we generated 9,055,826 reads (average length ~76 bp) and aligned them to the reference genome using Bowtie. The transcribed regions were analyzed using SAMTOOLS and custom Perl scripts in conjunction with BLAST searches and available gene annotation information. The single nucleotide resolution map enabled the identification of 14 novel protein coding regions as well as 44 potential novel sRNA. The basal transcription profile revealed that 2,506 of the 2,837 annotated regions were expressed in vitro, at 95.25% coverage, representing all broad functional gene categories in the genome. The expression profile also helped identify 518 potential operon structures involving 1,086 co-expressed pairs. We also identified 11 proteins with mutated/alternate start codons. The application of RNA-Seq based transcriptome profiling to structural gene annotation helped correct existing annotation errors and identify potential novel protein coding regions and sRNA. We used computational tools to predict regulatory elements such as promoters and terminators associated with the novel expressed regions for further characterization of these novel functional elements. Our study complements the existing structural annotation of Mannheimia haemolytica PHL213 based on experimental evidence. Given the role of sRNA in virulence gene regulation and stress response, potential novel sRNA described in this study can form the framework for future studies to determine the role of sRNA, if any, in M. haemolytica pathogenesis.