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Multiplexed sgRNA Expression Allows Versatile Single Non-repetitive DNA Labeling and Endogenous Gene Regulation

Shipeng Shao, Lei Chang, Yuao Sun, Yingping Hou, Xiaoying Fan, Yujie Sun
doi: https://doi.org/10.1101/121905
Shipeng Shao
1State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, China.
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Lei Chang
1State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, China.
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Yuao Sun
1State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, China.
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Yingping Hou
1State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, China.
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Xiaoying Fan
1State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, China.
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Yujie Sun
1State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, China.
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ABSTRACT

The CRISPR/Cas9 system has made significant contribution to genome editing, gene regulation and chromatin studies in recent years. High-throughput and systematic investigations into the multiplexed biological systems and disease conditions require simultaneous expression and coordinated functioning of multiple sgRNAs. However, current co-transfection based sgRNA co-expression systems remain poorly efficient and virus-based transfection approaches are relatively costly and labor intensive. Here we established a vector-independent method allowing multiple sgRNA expression cassettes to be assembled in series into a single plasmid. This synthetic biology-based strategy excels in its efficiency, controllability and scalability. Taking the flexibility advantage of this all-in-one sgRNA expressing system, we further explored its applications in single non-repetitive genomic locus imaging as well as coordinated gene regulation in live cells. With its strong potency, our method will greatly facilitate the understandings in genome structure, function and dynamics, and will contribute to the systemic investigations into complex physiological and pathological conditions.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY 4.0 International license.
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Posted March 29, 2017.
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Multiplexed sgRNA Expression Allows Versatile Single Non-repetitive DNA Labeling and Endogenous Gene Regulation
Shipeng Shao, Lei Chang, Yuao Sun, Yingping Hou, Xiaoying Fan, Yujie Sun
bioRxiv 121905; doi: https://doi.org/10.1101/121905
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Multiplexed sgRNA Expression Allows Versatile Single Non-repetitive DNA Labeling and Endogenous Gene Regulation
Shipeng Shao, Lei Chang, Yuao Sun, Yingping Hou, Xiaoying Fan, Yujie Sun
bioRxiv 121905; doi: https://doi.org/10.1101/121905

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