Chapter title |
Application of TALE-Based Approach for Dissecting Functional MicroRNA-302/367 in Cellular Reprogramming
|
---|---|
Chapter number | 21 |
Book title |
MicroRNA Protocols
|
Published in |
Methods in molecular biology, January 2018
|
DOI | 10.1007/978-1-4939-7601-0_21 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7600-3, 978-1-4939-7601-0
|
Authors |
Zhonghui Zhang, Wen-Shu Wu, Zhang, Zhonghui, Wu, Wen-Shu |
Abstract |
MicroRNAs are small 18-24 nt single-stranded noncoding RNA molecules involved in many biological processes, including stemness maintenance and cellular reprogramming. Current methods used in loss-of-function studies of microRNAs have several limitations. Here, we describe a new approach for dissecting miR-302/367 functions by transcription activator-like effectors (TALEs), which are natural effector proteins secreted by Xanthomonas and Ralstonia bacteria. Knockdown of the miR-302/367 cluster uses the Kruppel-associated box repressor domain fused with specific TALEs designed to bind the miR-302/367 cluster promoter. Knockout of the miR-302/367 cluster uses two pairs of TALE nucleases (TALENs) to delete the miR-302/367 cluster in human primary cells. Together, both TALE-based transcriptional repressor and TALENs are two promising approaches for loss-of-function studies of microRNA cluster in human primary cells. |
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