dRNA-seq-based analysis of the Thermococcus kodakarensis KOD1 transcriptional landscape to identify growth-regulated small regulatory RNAs

其中，高通量测序技术的出现彻底改变了从微生物到人类的RNA种群的特征；然而，还没有分析过高温Eurya chaeota的成员。这项拟议的研究项目旨在更详细地了解柯达热球菌KOD1的(S)核糖核酸种群。一种新近开发的高通量cDNA文库差异测序方法(dRNA-seq)将被应用于分析kodadarensis转录组对生长阶段的响应。除了转录组分析外，这些实验还将重点研究选定的sRNA的遗传、生物计算和生化特性，以揭示它们的生物学功能并确定潜在的相互作用(M)RNA。对此，将建立一个独特的报告系统来验证体内潜在的相互作用。进一步的实验将集中在RNA伴侣LSM的功能作用上。为此，标记LSM蛋白以进行免疫共沉淀，或标记稳定的RNA，将用于识别体外和体内潜在的相互作用。总之，这将导致更好地理解调控sRNA在古生菌领域中的作用。

Amongst others, the advent of high throughput sequencing technologies revolutionized the characterization of RNA populations from microbe to humans; however, no members of the hyperthermophilic euryarchaeota have yet been analyzed. The proposed research project aims to gain a more detailed insight into the (s)RNA population of Thermococcus kodadarensis KOD1. A recently developed differential method for high throughput sequencing of cDNA libraries (dRNA-seq) will be applied to analyse the T. kodadarensis transcriptome in response to the growth phase. Besides the transcriptome-analysis, the experiments will focus on the genetic, biocomputational and biochemical characterization of selected sRNAs in order to unravel their biological function and identify potential interacting (m)RNAs. With regard to this, a unique reporter system will be established to verify potential interactions in vivo. Further experiments will concentrate on the functional role of the RNA-chaperone Lsm. For this purpose, tagging of the Lsm protein for co-immunoprecipitation, or of stable RNAs, will be used to identify potential interactions in vitro and in vivo. Together, this will lead to a better understanding of the role of regulatory sRNAs in the archaeal domain.