Robust genome and RNA editing via CRISPR nucleases in PiggyBac systems

作     者：Yuqian Jiang Rachel Catherine Hoenisch Yun Chang Xiaoping Bao Craig E.Cameron Xiaojun Lance Lian 

作者机构：Department of Biomedical EngineeringPennsylvania State UniversityUniversity ParkPA16802USA Huck Institutes of the Life SciencesPennsylvania State UniversityUniversity ParkPA16802USA Department of BiologyPennsylvania State UniversityUniversity ParkPA16802USA Davidson School of Chemical EngineeringPurdue UniversityWest LafayetteIN47907USA Department of Microbiology and ImmunologyUniversity of North Carolina School of MedicineChapel HillNC27599USA 

出 版 物：《Bioactive Materials》 (生物活性材料（英文）)

年 卷 期：2022年第7卷第8期

页      面：313-320页

核心收录：

学科分类：0710[理学-生物学] 0831[工学-生物医学工程（可授工学、理学、医学学位）] 07[理学] 08[工学] 071007[理学-遗传学] 0805[工学-材料科学与工程（可授工学、理学学位）] 

基　　金：supported by NIH R21EB026035(X.L.L.) NIH R21AI149312(C.E.C.,and X.L.L.) NSF CBET-1943696(X.L.L.) Penn State startup funding to X.L.L. 

主　　题：CRISPR-Cas9 Genome editing Cas13d RNA editing PiggyBac transposon Human pluripotent stem cells 

摘      要：CRISPR/Cas-mediated genome editing in human pluripotent stem cells(hPSCs)offers unprecedented opportunities for developing in vitro disease modeling,drug screening and cell-based therapies.To efficiently deliver the CRISPR components,here we developed two all-in-one vectors containing Cas9/gRNA and inducible Cas13d/gRNA cassettes for robust genome editing and RNA interference respectively.These vectors utilized the PiggyBac transposon system,which allows stable expression of CRISPR components in hPSCs.The Cas9 vector PB-CRISPR exhibited high efficiency(up to 99%)of inducing gene knockout in both protein-coding genes and long non-coding RNAs.The other inducible Cas13d vector achieved extremely high efficiency in RNA knockdown(98%knockdown for CD90)with optimized gRNA designs.Taken together,our PiggyBac CRISPR vectors can serve as powerful toolkits for studying gene functions in hPSCs.