Boosting CRISPR/Cas9 multiplex editing capability with the endogenous tRNA-processing system

Proceedings of the National Academy of Sciences of the United States of America
Kabin XieYinong Yang

Abstract

The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 nuclease (Cas9) system is being harnessed as a powerful tool for genome engineering in basic research, molecular therapy, and crop improvement. This system uses a small guide RNA (gRNA) to direct Cas9 endonuclease to a specific DNA site; thus, its targeting capability is largely constrained by the gRNA-expressing device. In this study, we developed a general strategy to produce numerous gRNAs from a single polycistronic gene. The endogenous tRNA-processing system, which precisely cleaves both ends of the tRNA precursor, was engineered as a simple and robust platform to boost the targeting and multiplex editing capability of the CRISPR/Cas9 system. We demonstrated that synthetic genes with tandemly arrayed tRNA-gRNA architecture were efficiently and precisely processed into gRNAs with desired 5' targeting sequences in vivo, which directed Cas9 to edit multiple chromosomal targets. Using this strategy, multiplex genome editing and chromosomal-fragment deletion were readily achieved in stable transgenic rice plants with a high efficiency (up to 100%). Because tRNA and its processing system are virtually conserved in all living organis...Continue Reading

Associated Protocols

References

Feb 16, 2016·Nature Reviews. Genetics·Jackson ChamperOmar S Akbari
Feb 14, 2016·Current Opinion in Biotechnology·Raissa Estrela, Jamie Harrison Doudna Cate
Sep 24, 2015·Wiley Interdisciplinary Reviews. Systems Biology and Medicine·Zehua BaoHuimin Zhao
Nov 15, 2015·Plant Science : an International Journal of Experimental Plant Biology·Francis Quétier
Dec 29, 2015·Current Opinion in Biotechnology·Oleg Raitskin, Nicola J Patron
Dec 27, 2015·Plant Science : an International Journal of Experimental Plant Biology·James R Ketudat CairnsJong-Seong Jeon
Sep 16, 2015·Plant Biotechnology Journal·Junwon LeeHyongbum Kim
Aug 12, 2015·Plant Biotechnology Journal·Donald P WeeksBing Yang
Jan 30, 2016·Plant Methods·Simon Schiml, Holger Puchta
Jan 18, 2016·Trends in Biotechnology·Sachin TeotiaGuiliang Tang
Nov 19, 2015·The Plant Journal : for Cell and Molecular Biology·Jeannette SteinertHolger Puchta
Jan 9, 2016·Journal of Genetics and Genomics = Yi Chuan Xue Bao·Chun WangKejian Wang
Jun 25, 2016·Phytochemistry Reviews : Proceedings of the Phytochemical Society of Europe·Matthew B Kilgore, Toni M Kutchan
Dec 17, 2015·Scientific Reports·Shin YoshiokaKunihiko Naito
Sep 12, 2015·GM Crops & Food·Chung-Jui Tsai, Liang-Jiao Xue
Oct 16, 2015·Biotechnology and Bioengineering·Michelle L LuoChase L Beisel
May 12, 2016·Journal of Genetics and Genomics = Yi Chuan Xue Bao·Dandan ZhangJian-Feng Li
Oct 18, 2015·Plant Science : an International Journal of Experimental Plant Biology·Scott M Schaeffer, Paul A Nakata
Mar 31, 2016·Plant Cell Reports·Teodoro Cardi, C Neal Stewart
Feb 16, 2016·PeerJ. Computer Science·Elisha D O Roberson
May 6, 2016·Frontiers in Plant Science·Zafar IqbalMuhammad Shafiq
Jun 10, 2015·Molecular Plant·Liang-Jiao Xue, Chung-Jui Tsai
May 6, 2016·Frontiers in Plant Science·Surender KhatodiaNarendra Tuteja
Apr 27, 2016·Plant Cell Reports·Joseph W Paul, Yiping Qi
May 18, 2016·Progress in Retinal and Eye Research·Sandy S C HungAlex W Hewitt
May 20, 2016·Plant Cell Reports·Fabien NoguéJosep M Casacuberta
Jun 3, 2016·Frontiers in Plant Science·Yuduan DingKabin Xie
Apr 25, 2016·Molecular Plant·Xingliang MaYao-Guang Liu
Aug 21, 2016·Journal of Genetics and Genomics = Yi Chuan Xue Bao·Rongfang XuJianbo Yang
Aug 4, 2016·Scientific Reports·Roberto GaliziAndrea Crisanti
Dec 19, 2015·The Plant Journal : for Cell and Molecular Biology·Holger Puchta
Mar 8, 2016·Journal of Integrative Plant Biology·Hyeran KimSang-Gyu Kim
Jul 9, 2016·Phytopathology·Vincent N FondongSavithramma P Dinesh-Kumar
Sep 16, 2016·Frontiers in Plant Science·Longxiang WangDeqiang Duanmu
Sep 7, 2016·Nature Methods·Fillip Port, Simon L Bullock
Aug 12, 2016·Plant Biotechnology Journal·Si Nian CharBing Yang
Sep 14, 2016·Scientific Reports·Zhongping XuXianlong Zhang

Citations

Aug 17, 1990·Science·A C Forster, S Altman
Oct 24, 1995·Proceedings of the National Academy of Sciences of the United States of America·S Yokobori, S Pääbo
May 29, 2002·The EMBO Journal·Steffen SchifferAnita Marchfelder
Nov 6, 2007·Trends in Genetics : TIG·Giorgio DieciAldo Pagano
Nov 6, 2008·Nucleic Acids Research·Patricia P Chan, Todd M Lowe
Nov 6, 2008·PloS One·Carola EnglerSylvestre Marillonnet
May 5, 2009·Plant Physiology·Giusy CaninoAnita Marchfelder
Sep 3, 2010·Genes & Development·Eric M Phizicky, Anita K Hopper
May 5, 2011·Nature Reviews. Genetics·Robert J White
May 3, 2012·Genes & Development·Bernard GutmannPhilippe Giegé
Jun 30, 2012·Science·Martin JinekEmmanuelle Charpentier
Jan 5, 2013·Science·Le CongFeng Zhang
Jan 5, 2013·Science·Prashant MaliGeorge M Church
Feb 7, 2013·ELife·Martin JinekJennifer A Doudna
Aug 10, 2013·Nature Biotechnology·Qiwei ShanCaixia Gao
Aug 10, 2013·Nature Biotechnology·Vladimir NekrasovSophien Kamoun
Aug 21, 2013·Molecular Plant·Kabin Xie, Yinong Yang
Oct 1, 2013·Nature Methods·Prashant MaliGeorge M Church
Feb 20, 2014·Proceedings of the National Academy of Sciences of the United States of America·Zhengyan FengJian-Kang Zhu
Mar 4, 2014·Nature Biotechnology·Jeffry D Sander, J Keith Joung
Apr 29, 2014·Nature Biotechnology·Shengdar Q TsaiJ Keith Joung

Related Concepts

Study
In Vivo
Basic Research
Small Nuclear RNA
Genes, Plant
Rice (Dietary)
CRISPR-Cas Systems
Endonuclease
MT-TA gene
Gene Products, Protein

Related Feeds

CRISPR Genome Editing & Therapy

CRISPR-Cas system enables the editing of genes to create or correct mutations. This feed focuses on the application of this system for gene editing and therapy in human diseases.

CRISPR (general)

Clustered regularly interspaced short palindromic repeats (CRISPR) are DNA sequences in the genome that are recognized and cleaved by CRISPR-associated proteins (Cas). CRISPR-Cas system enables the editing of genes to create or correct mutations. Discover the latest research on CRISPR here.

CRISPR Ribonucleases Deactivation

CRISPR-Cas system enables the editing of genes to create or correct mutations. This feed focuses on mechanisms that underlie deactivation of CRISPR ribonucleases. Here is the latest research.

CRISPR for Genome Editing

Genome editing technologies enable the editing of genes to create or correct mutations. Clustered regularly interspaced short palindromic repeats (CRISPR) are DNA sequences in the genome that are recognized and cleaved by CRISPR-associated proteins (Cas). Here is the latest research on the use of CRISPR-Cas system in gene editing.

Biophysics of CRISPR

This feed focuses on broad characteristics of the CRISPR system and the proteins associated with it.