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Application of TALE-Based Approach for Dissecting Functional MicroRNA-302/367 in Cellular Reprogramming

  • Zhonghui Zhang
  • Wen-Shu Wu
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1733)

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.

Key words

TALE TALEN Transcriptional repressor MicroRNA Cellular reprogramming Human cells 

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Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  1. 1.School of Life SciencesShanghai UniversityShanghaiChina
  2. 2.Division of Hematology/Oncology, Department of Medicine and Cancer CenterUniversity of Illinois at ChicagoChicagoUSA

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