Digital Gene Expression Analysis by 3’ End Sequencing

通过 3™ 末端测序进行数字基因表达分析

• 批准号: 9048237
• 负责人: BIN TIAN
• 金额: $ 27万
• 依托单位: BIOO SCIENTIFIC CORPORATION
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9048237
• 关键词: Address; Back; Biology; Cells; Charge; DNA analysis; Data; Data Analyses; Eukaryota; Gene Expression; Gene Expression Profiling; Genes; Genetic Transcription; Genetic Translation; Genome; Goals; Grant; Informatics; Lead; Legal patent; Length; Licensing; Maps; Medicine; Messenger RNA; Metabolism; Methods; Molecular; Oligonucleotides; Plague; Poly A; Poly(A) Tail; Poly(A)+ RNA; Polyadenylation; Polymerase; Polynucleotide Adenylyltransferase; Protein Isoforms; RNA; RNA Sequences; Reading; Research; Sequence Analysis; Site; Small Business Technology Transfer Research; Stretching; Techniques; Technology; Transcript; Translations; Untranslated RNA; base; commercialization; cost effective; deep sequencing; digital; experience; indexing; locked nucleic acid; mRNA Stability; next generation sequencing; novel; public health relevance; sequencing platform; serial analysis of gene expression; success; tool; transcriptome sequencing; usability

 DESCRIPTION (provided by applicant): A key application of deep sequencing is gene expression analysis at the RNA level, commonly known as RNA-seq. Reads from RNA-seq cover all regions of an mRNA sequence. However, sequences derived from the 3' end region are sufficient to indicate gene expression levels. All 3' end sequencing methods have revealed that most eukaryotic genes display alternative cleavage and polyadenylation (APA), where RNA isoforms using different cleavage and polyadenylation sites (pAs) can be generated from a gene. Importantly, different APA isoforms have been shown to have different metabolisms, including stability, localization and translation. APA clearly poses a challenge for gene expression analysis. Current RNA-seq methods cannot capture this information; second, false pA identification due to internal priming leads to inaccurate APA information; third, complete analysis of gene expression needs to take into consideration all APA isoforms. We have proposed a new method that completely addresses false priming, incomplete 3' end annotation and significantly propels research on single cell RNA expression and poly(A) tail length, which have so far been elusive. The final goal of this proposal is to develop a commercial kit, allowing non-specialized labs the ability to examine 3'ends in an accurate and sensitive manner.

 描述（通过应用程序提供）：深度测序的关键应用是RNA水平的基因表达分析，通常称为RNA-Seq。从RNA-Seq读取mRNA序列的所有区域。但是，从3'末端区域得出的序列足以指示基因表达水平。所有3'末端测序方法都表明，大多数真核基因都显示出替代性裂解和聚腺苷酸化（APA），其中使用不同的裂解和聚腺苷酸化位点（PAS）的RNA同工型可以从基因中产生。重要的是，已经证明不同的APA同工型具有不同的代谢，包括稳定性，定位和翻译。 APA显然对基因表达分析提出了挑战。当前的RNA-seq方法无法捕获此信息；其次，由于内部启动而引起的错误PA识别导致APA信息不准确；第三，对基因表达的完整分析需要考虑所有APA同工型。我们提出了一种新方法，该方法完全解决了错误的启动，不完整的3'末端注释，并显着推动了对单细胞RNA表达和poly（a）尾巴长度的研究，而迄今为止难以捉摸。该提案的最终目标是开发商业套件，允许非专业实验室以准确和敏感的方式检查3端的能力。