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Single cell tagging of localized RNA from whole populations

来自整个群体的局部 RNA 的单细胞标记

基本信息

批准号：
10456941
负责人：
Michael Aaron White
金额：
$ 33.86万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-20 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10456941
关键词：
Affinity Animal Model Animals Bar Codes Binding Binding Proteins Biotechnology CRISPR interference CRISPR library Cataloging Cell Culture Techniques Cell Separation Cells Clustered Regularly Interspaced Short Palindromic Repeats Code Complex Custom Cytoplasmic Granules DNA DNA Sequence Data Data Pooling Development Elements Gene Expression Gene Expression Regulation Genes Genetic Genetic Diseases Genetic Screening Genetic Variation Genetic study Genome Goals Guide RNA Individual Invertebrates Label Lentivirus Vector Ligation Link Location Mammalian Cell Mammalian Genetics Measures Messenger RNA Methods Mitochondria Poly U Poly(A) Tail Population Proteins RNA RNA analysis RNA-Binding Proteins RNA-Directed DNA Polymerase Regulatory Element Reporter Genes Resolution Ribonucleases Ribosomal Proteins Ribosomes Sampling Specificity Technology Tissues Transcript Untranslated RNA Viral Vector Work aptamer base design disease phenotype experimental study gene function gene repression genetic variant genome editing human disease in vivo knock-down mRNA tagging mammalian genome new technology single cell analysis single cell sequencing single cell technology small hairpin RNA transcriptome

项目摘要

Project Summary/Abstract The objective of this proposal is to develop a broadly applicable technology, called SCALoP (Single Cell Analysis of Localized RNA on whole Populations), to measure gene expression in single cells, without the need to perform technically challenging manipulations on individual cells. Currently, large-scale genetic screens based on new technologies are leading to important advances in our understanding of mammalian genomes and genetic variation linked to human disease. However, the power of these screens is limited by the lack of methods to measure gene expression in these large screens. Our proposed technology would fill this major unmet need, and thus have a broad impact on mammalian genetics. Our goal is to develop a method to attach single-cell-specific sequence barcodes to transcripts, using RNA proximity ligation in pooled samples. We propose to do this by designing barcoded “tagRNAs” which are expressed in cells and targeted to specific RNA binding proteins. These tagRNAs are attached to transcripts derived from the same single cell by proximity ligation. To develop this method, our aims are 1) optimize and quantify the efficiency of methods to link tagRNAs and cellular RNA, 2) optimize in vivo specificity and single cell resolution of SCALoP, and 3) develop RNA aptamers to natural proteins and alternate location targets.

项目总结/文摘