Construction and Characterization of a Synthetic MicroRNA Cluster for Multiplex RNA Interference in Mammalian Cells.
作者： Tingting Wang , Yue Xie , Aidi Tan , Shao Li , Zhen Xie
DOI: 10.1021/ACSSYNBIO.5B00180
关键词: Genetics 、 microRNA 、 RNA interference 、 RNA 、 Synthetic biology 、 RNA polymerase II 、 PiggyBac Transposon System 、 Biology 、 Computational biology 、 Multiplex 、 CRISPR
摘要: RNA interference (RNAi) technology is widely used in basic and translational research. By mimicking a natural primary microRNA (pri-miRNA) cluster, multiple engineered hairpins can be transcribed as single transcript from the same Pol II promoter, enabling formation of multiplex RNAi mammalian cells. However, constructing synthetic miRNA cluster still time-consuming, processing function are incompletely understood. Here, we identified precursor architecture that allows precise maturation. We established hierarchical cloning method for efficient construction harboring up to 18 precursors. demonstrated maturation individual precursors were independent their positions cluster. then analyzed integration efficiency clusters having varied number by using CRISPR/Cas9-mediated integration, piggyBac transposon system, lentiviral system. This system provides an important tool
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