Genetics of long non-coding RNAs in zebrafish

描述（由申请人提供）：拟议研究的长期目标是确定长链非编码RNA在脊椎动物发育中的功能。在过去的十年里，全基因组研究出现了爆炸式的增长，这些研究发现了大量以前未注释的转录本。在这些新的转录本中，长链非编码RNA（lncRNA）引起了特别的关注，因为它们代表了基因组中广泛的、大部分未被探索的组成部分：基因组的“暗物质”。该研究旨在进行反向遗传筛选并分析斑马鱼中100种lncRNA的功能。基因组学和计算方法已经鉴定了斑马鱼发育过程中表达的数百种lncRNA。通过保守、基因组定位、表达和染色质谱选择的100个这些lncRNA将被破坏。反义寡核苷酸将用于直接干扰lncRNA，并且位点特异性核酸内切酶（TALEN和CRISPR）将用于破坏lncRNA基因。然后，胚胎和幼虫将进行系统的表型表征，以揭示lncRNA的功能。拟议的反向遗传筛选将提供脊椎动物lncRNA在体内功能的第一次全面分析，并为该领域提供资源，以揭示这类神秘分子的作用。

DESCRIPTION (provided by applicant): The long-range goal of the proposed studies is to determine the functions of long non-coding RNAs in vertebrate development. The past decade has seen an explosion of genome-wide studies that have identified a plethora of previously unannotated transcripts. Among these novel transcripts, the class of long non-coding RNAs (lncRNAs) has attracted particular attention since they represent an extensive, largely unexplored component of the genome: the "dark matter" of the genome. The proposed study aims to perform a reverse genetic screen and analyze the function of 100 lncRNAs in zebrafish. Genomics and computational approaches have led to the identification of several hundred lncRNAs expressed during zebrafish development. 100 of these lncRNAs, selected by conservation, genomic location, expression, and chromatin profile, will be disrupted. Antisense oligonucleotides will be used to directly interfere with lncRNAs, and site-specific endonucleases (TALENs and CRISPRs) will be employed to disrupt lncRNA genes. Embryos and larvae will then undergo systematic phenotypic characterization to reveal the functions of lncRNAs. The proposed reverse genetic screen will provide the first comprehensive analysis of vertebrate lncRNA function in vivo and generate the resources for the field to reveal the roles of this mysterious class of molecules.