Generation of a pooled shRNA library for functional genomics screens

STAR Protocols

卷:3

Carsten Patrick Ade¹  Dimitrios Papadopoulos²  Martin Eilers² 

[1] Corresponding author;

[2] Theodor Boveri Institute, Department of Biochemistry and Molecular Biology, Biocenter, University of Würzburg, Am Hubland, 97074 Würzburg, Germany;

关键词: Bioinformatics;    Sequence analysis;    Genomics;    Sequencing;    High Throughput Screening;    Molecular Biology;   

DOI  :  

来源: DOAJ

【 摘 要 】

Summary: Here, we detail a protocol for the generation of pooled short hairpin RNA (shRNA) libraries. We cover the design of optimized miR-E backbone shRNAs, cloning into a Tet-on vector system, and transformation of competent bacteria. We also describe library quality check by next-generation sequencing, and finally the production of lentiviruses. This protocol will generate high-quality inducible libraries suitable for both genome-wide and targeted functional genomics screens, allowing the high-throughput interrogation of protein depletion effects in the cell system of choice.For complete details on the use and execution of this protocol, please refer to Papadopoulos et al. (2022).

【 授权许可】

Unknown