PT  - JOURNAL ARTICLE
AU  - Shipeng Shao
AU  - Lei Chang
AU  - Yuao Sun
AU  - Yingping Hou
AU  - Xiaoying Fan
AU  - Yujie Sun
TI  - Multiplexed sgRNA Expression Allows Versatile Single Non-repetitive DNA Labeling and Endogenous Gene Regulation
AID  - 10.1101/121905
DP  - 2017 Jan 01
TA  - bioRxiv
PG  - 121905
4099  - http://biorxiv.org/content/early/2017/03/29/121905.short
4100  - http://biorxiv.org/content/early/2017/03/29/121905.full
AB  - 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.