左二偉課題組

Zuo Erwei Lab

左二偉,，中國農(nóng)業(yè)科學(xué)院深圳農(nóng)業(yè)基因組研究所研究員、博士生導(dǎo)師,。近年來,，致力于利用分子生物學(xué)、計算生物學(xué)等手段開展新型基因編輯技術(shù)的研發(fā)與應(yīng)用研究,，推動基因編輯技術(shù)醫(yī)學(xué)應(yīng)用安全性評價體系建設(shè),。提出脫靶效應(yīng)產(chǎn)生的分子基礎(chǔ),，研發(fā)多種檢測技術(shù),，證實了單堿基編輯器會導(dǎo)致基因組范圍內(nèi)完全隨機無法預(yù)測的脫靶效應(yīng),；發(fā)現(xiàn)了脫氨酶產(chǎn)生脫靶效應(yīng)的直接證據(jù)，理清了脫氨酶的核酸結(jié)合結(jié)構(gòu)域與脫靶效應(yīng)之間的關(guān)系,，研發(fā)了效率,、精度具佳的單堿基編輯工具，在一定程度上解決了醫(yī)學(xué)應(yīng)用重大問題,；實現(xiàn)了對多種人類疾病小鼠模型的治療研究，取得了良好效果,，為開展臨床實驗提供依據(jù),，有望治療相關(guān)疾病。構(gòu)建了C-CRISPR技術(shù),，顯著提高了基因編輯效率,，研制了F0代基因完全敲除靈長類動物,，為快速建立靈長類動物疾病模型做出貢獻(xiàn)。其中單堿基基因編輯造成大量脫靶效應(yīng)及其優(yōu)化解決方法入選“2019中國生命科學(xué)十大進展”,。合理設(shè)計胞嘧啶堿基編輯器可在保持高靶向活性同時降低DNA和RNA的非靶向效應(yīng)入選中國農(nóng)業(yè)科學(xué)院“2020重大科學(xué)發(fā)現(xiàn)”,，獲得2021基因組研究所“重大成果獎”。一步法生產(chǎn)基因完全敲除動物研究成果在2017年國際干細(xì)胞研究學(xué)會年會（2017 ISSCR）獲得“Merit Award”,。近年來,，基因編輯領(lǐng)域在Science、Nature等期刊發(fā)表多篇研究論文,。

Erwei Zuo is a researcher of Institute of Agricultural Genome Research, Chinese Academy of Agricultural Sciences. As a molecular biologist, Zuo focused on the development of genome editing tools, and the construction of a safety evaluation system for gene-editing applications. He and his team have proposed the molecular basis of the off-target effect, developed various detection technologies, and confirmed that single-base editors can cause completely random and unpredictable off-target effects within the genome. He revealed direct evidence of off-target effects of the deaminase, and clarified the relationship between the nucleic acid-binding domain of deaminase and off-target effects, developed a single-base editing tool with high efficiency and accuracy, which overcame the major challenge in medical applications to a certain extent. Moreover, his achievements in therapeutic studies using mouse models of human diseases provide support for therapeutic applications. Additionally, C-CRISPR technology significantly improved the efficiency of gene editing, and developed F0 generation of primates bearing gene knockout, which made important contributions to the rapid establishment of primate disease models.

Zuo is a recipient of many awards. Single-base gene editing caused substantial off-target effects, and its optimization was selected as the "Top Ten Advances in Life Sciences in China 2019". The rational engineering of cytosine base editor that enables to minimize of the off-target effect of DNA and RNA and maintains high targeting activity was selected as one of the "2020 Major Scientific Discoveries" by the Chinese Academy of Agricultural Sciences and received the "Major Achievement Award" by the Institute of Genomic Research 2021. The one-step production of complete knockout animals received the "Merit Award" at the 2017 ISSCR annual meeting. Recently, he has published a large number of articles in gene editing top journals including  Science ,  Nature, etc.

團隊研究方向:

團隊主要開展新型基因編輯技術(shù)的研發(fā)與應(yīng)用研究,。通過提高基因編輯工具的效率和精準(zhǔn)度，減少其脫靶效應(yīng),，擴大其應(yīng)用范圍,。推動基因編輯技術(shù)在醫(yī)學(xué)應(yīng)用的安全性評價體系建設(shè)，不斷探索基因編輯技術(shù)在生豬育種領(lǐng)域的創(chuàng)新應(yīng)用,，創(chuàng)制育種新材料,。

The team focuses on the development and application of novel gene editing technologies. The improved efficiency and accuracy of gene editing tools reduce their off-target effects and expand their application scope. The achievements promoted the construction of a safety assessment system of gene editing technology in both medical and biological breeding fields, enhanced the application of gene editing technology in the pig breeding field, and created new breeding materials.

研究進展：

（1）建立了“一步法”獲得基因完全敲除動物的技術(shù)體系，有效縮短基因編輯動物的創(chuàng)制時間,。(Zuo et. al., Cell Research, 2017)

Established a "one-step" technology system to generate complete knockout animals and effectively shorten the time of producing genetic animals.

（2）建立了全染色體敲除技術(shù),，為開發(fā)染色體缺失的動物模型提供了一種新方法，也為涉及人類非整倍體染色體疾病提供了潛在的治療策略,。(Zuo et. al., Genome Biology, 2017)

Established a whole chromosome knockout technique that offers a new approach to developing animal models with chromosome deletions, and a potential therapeutic strategy for human aneuploidy diseases involving additional chromosomes.

（3）建立了GOTI的新型脫靶檢測技術(shù),，可檢測之前手段無法發(fā)現(xiàn)的完全隨機的脫靶位點，顯著提高基因編輯技術(shù)的脫靶檢測靈敏性,，為基因編輯工具的安全性評估帶來了突破性的新工具,，有望成為新的行業(yè)檢測標(biāo)準(zhǔn)。(Zuo et.al., Science, 2019; Zuo et.al., Nature Protocols, 2020)

Developed “GOTI”, a novel off-target detection technology that can detect completely random off-target which is undetectable by previous means. Moreover, this strategy significantly improved the sensitivity of off-target detection for gene editing technology, brought a breakthrough new tool for safety assessment of gene editing tools, and potentially became a new industry standard for detection.

（4）開發(fā)新一代高保真基因編輯技術(shù),，可進行高效率和高保真度的C-G單堿基編輯,，有效降低脫靶效率。(Zuo et. al., Nature methods. 2020,；Yuan et. al., Nature Communications,2021)

Designed a new version of high-fidelity gene editing technology that allows for high-efficiency and high-fidelity C-G single-base editing, effectively reducing off-target efficiency.

研究成果：

1. Wei,Y.*, Li,Z.*, Xu,K.*, Feng,H.*, Xie,L.*, Li,D., Zuo,Z., Zhang,M.#, Xu,C.#, Yang,H.#, Zuo,E.#. Mitochondrial base editor DdCBE causes substantial DNA off-target editing in nuclear genome of embryos[J]. Cell Discovery, 2022; 8(27): 1-4.

2. Wei,Y.*, Xu,C.*, Feng,H.*, Xu,K.*, Li,Z.*, Hu,J., Zhou,L., Wei,Y., Zuo,Z., Zuo, E., Li,W.#, Yang,H.#, Zhang,M.#. Human cleaving embryos enable efficient mitochondrial base-editing with DdCBE[J]. Cell Discovery, 2022; 8(7): 1-4

3. Gu,X.*, Hu,X.*, Wang,D.*, Xu,Z.,  Wang,F., Li,D.,  Li, G., Yang,H.,Li,H.#, Zuo,E.# & Shu,Y.#, （2022）.Treatment of autosomal recessive hearing loss via in vivo CRISPR/Cas9-mediated optimized homology-directed repair in mice.Cell Research, volume 32, pages 699–702

4. Sameh A.Abdelnour,Xie,L., Abdallah A. Hassanin, Zuo,E.#and  Lu,Y#.(2021)The Potential of CRISPR/Cas9 Gene Editing as a Treatment Strategy for Inherited. Diseases.Frontiers in Cell and Developmental Biology.2021,9

5. Yuan,T.*, Yan,N.*, Fei,T.*, Zheng,J.*, Meng,J.*, Li,N.*, Liu,J.*, Zhang,H., Xie,L., Ying,W., Li,D., Shi,L., Sun,Y.,Li,Y., Li,Y., Sun,Y.# and Zuo,E.#.(2021). Optimization of C-to-G base editors with sequence context preference predictable by machine learning methods.Nature Communications: 12:4902

6. Xue,Y.*, Hu,X.*,Wang,D.*, Li,D.*,Li,Y.,Wang,F.,Huang,M., Gu,X., Xu,Z., Zhou,J., Wang,J., Chai,R., Shen,J., Chen,Z., Li,G., Yang,H., Li,H.#, Zuo,E.# and Shu,Y.#. (2021).Gene editing in a Myo6 semi-dominant mouse model rescues auditory function Author links open overlay panel .Molecular Therapy. DOI: 10.1016/j.ymthe.2021.06.015

7. Zhang,J.*, E,M.Khazalwa.*, H, M.Abkallo., Zhou,Y., Nie,X., Ruan,J., Zhao,C., Wang,J., Xu,J., Li.X., Zhao,S., Zuo,E.#, L,Steinaa#.and Xie,S.#.(2021).The Advancements, Challenges and Future Implications of the CRISPR/Cas9 System in Swine Research.Journal of Genetics and Genomics:48(5).347-360.

8. Wei,Y.*, Zhou, Y.*, Liu,Y.*, Ying,W., Lv,R., Zhao,Q.,  Zhou,H., Zuo,E.#,  Sun,Y.#, Yang,H.# and Zhou,C,#. (2021). Indiscriminate ssDNA cleavage activity of CRISPR-Cas12a induces no detectable off-target effects in mouse embryos.Protein & Cell .12,741–745.  DOI: 10.1007/s13238-021-00824-z

9. Huang, X.*, Lv, J.,*, Li, Y.*, Mao, S., Li, Z., Jing, Z., Sun, Y., Zhang,X., Shen, S., Wang, X., Di, M., Ge, J., Huang, X., Zuo,E.# and Chi, T.#.（2020）. Programmable C-to-U RNA editing using the human APOBEC3A deaminase .The EMBO Journal:39(22):e104741.

10. Zuo,E.*, Sun,Y. *, Wei,Wu.*, Yuan ,T.*, Ying,W., Sun,H., Yuan,L., Lars M. Steinmetz # , Li,Y.# and Yang,H.#.(2020).GOTI, a method to identify genome-wide off-target effects of genome editing in mouse embryos.Natrue Protocols:15.3009–3029.

11. Lee,S.*, Ding,N.*, Sun,Y., Yuan,T., Li,J., Yuan,Q., Liu,L.,Yang,J.,Wang,Q., Anatoly B. Kolomeisky, Isaac B. Hilton, Zuo,E.# and Gao,X.#. (2020).Single C-to-T substitution using engineered  APOBEC3G-nCas9 base editors with minimum  genome- and transcriptome-wide off-target effects.Science Advance:6（29）.

12. Zuo,E.*#, Sun,Y.*, Yuan ,T.*?, He,B.*?, Zhou,C.*?, Ying,W., Liu,L., Wei?,W., Zeng,R., Li,Y.# and Yang,H.#.(2020).A rationally engineered cytosine base editor retains high on-target activity while reducing both DNA and RNA off-target effects.Nature Methods:17(6):600–604.

13.  Lin X.*, Chen H.*, Lu Y.*, Hong S.*, Hu X.*, Gao Y., Lai L., Li J., Wang Z., Ying W., Ma L., Wang N., Zuo,E.#, Yang H.# and Chen W. #.(2020).Base editing-mediated splicing correction therapy for spinal muscular atrophy. Cell Research:0:1-3. 30(6).

14. Deng K.*, Feng W.*, Liu X.*, Su X., Zuo,E., Du S., Huang Y., Shi D. # and Lu,F. # .(2020).Anti-silencing Factor 1A is Associated with Genome Stability Maintenance of Mouse Preimplantation Embryos.Biology of Reproduction:102(4),817-827.

15. Li J. *, Lin X. *, Tang C. *, Lu Y. *, Hu X. *, Zuo,E., Li H., Ying W., Sun Y. , Lai L., Chen H., Guo X., Zhang Q., Wu S., Zhou C., Shen X., Wang Q., Lin ., Ma L., Wang N.,Ad.Krainer, Shi L.#, Yang H.# and Chen W.#.(2020) .Disruption of splicing-regulatory elements using CRISPR/Cas9 to rescue spinal muscular atrophy in human iPSCs and mice.National Science Review:7(1),92-101.

16. Zhao, X., Wei, W., Pan,H., Nie,J., Chen,D., Zhang,P., Chen, F., Fu, Q,. Zuo, E.#, Lu, Y.# and Zhang, M.#. (2019). Identification of the Sex of Pre-implantation Mouse Embryos Using a Marked Y Chromosome and CRISPR/Cas9. SCIENTIFIC REPORTS:9: 14315.

17. Li,J.* , Hong,S.*, Chen,W.#, Zuo,E.# and Yang,H.#.(2019) . Advances in detecting and reducing off-target effects generated by CRISPR-mediated genome editing.Journal of Genetics and Genomics:46(11),513-521.

18. Wang, X.*, Kang, J.*, Wei, L.*, Yang, X.*, Sun, H., Yang, S., Lu, L., Yang, M., Bai, M., Chen, Y., Long, J., Li, N., Li, D., Huang, J., Lei, M., Shao, Z., Yuan, W.#, Zuo, E.#, Lu, K. #, Liu, M.# and Li, J#. (2019). PHF7 is a novel histone H2A E3 ligase prior to histone-to-protamine exchange during spermiogenesis. Development:146（13） dev.175547.

19. Zhou, C.*, Sun, Y.*, Yan, R.*, Liu, Y.*, Zuo, E.*, Gu, C., Han, L., Wei, Y., Hu, X., Zeng, R., Li, Y.#, Zhou, H.#, Guo, F.# and Yang, H.# .(2019).Off-target RNA mutation induced by DNA base editing and its elimination by mutagenesis. Nature:571, 275–278.

20. Zuo, E.*, Sun, Y.*, Wei, W.*, Yuan, T.*, Ying, W., Sun, H., Yuan, L., Steinmetz, L. M.#, Li, Y.# and Yang, H#.(2019). Cytosine base editor generates substantial off-target single-nucleotide variants in mouse embryos. Science:364（6437）, 289-292.

21. Zuo, E.*, Huo, X.*, Yao, X.*, Hu, X.*, Sun, Y.*, Yin, J*., He, B., Wang, X., Shi, L., Ping, J., Wei, Y., Ying, W., Wei, W., Liu, W., Tang, C., Li, Y., Hu, J.# and Yang, H.#. (2017).CRISPR/Cas9-mediated targeted chromosome elimination. Genome Biology:18:224.

22. Zuo, E.*, Cai, Y. J.*, Li, K.*, Wei, Y.*, Wang, B. A.*, Sun, Y.*, Liu, Z., Liu, J., Hu, X., Wei, W., Huo, X., Shi, L., Tang, C., Liang, D., Wang, Y., Nie, Y. H., Zhang, C. C., Yao, X., Wang, X., Zhou, C., Ying, W., Wang, Q., Chen, R. C., Shen, Q., Xu, G. L., Li, J., Sun, Q., Xiong#, Z. Q.# and Yang, H#.(2017).One-step generation of complete gene knockout mice and monkeys by CRISPR/Cas9-mediated gene editing with multiple sgRNAs. Cell Research:27, 933-945.

23. Wang,L. *, Li,M. *, Qu, C.,Miao,W., Yin,Q., Liao,J., Cao,H., Huang,M., Wang,K. Zuo,E. , Peng,G., Zhang,S., Chen,G., Li,Q., Tang,K., Yu, Q., Li,Z., Catherine CL Wong, Xu,G., Jing, N.,Yu,X.,# and Li,J.#.(2017).CRISPR-Cas9-mediated genome editing in one blastomere of two-cell embryos reveals a novel Tet3 function in regulating neocortical development.Cell Research:1-15.

24. Zuo, E. *, Yang, X.#, Lu, Y., Xie, L., Shang, J., Li, D., Yang, H., Hu, L., Zhao, H., Lu, S., Lu, K.#.(2015). ZPAC is required for normal spermatogenesis in mice. Mol. Reprod：Dev. 82(10): 747-755.

25. Zhang, M.*, Zhou, H., Zheng, C. ,Xiao, J. , Zuo, E., Liu, W., Xie, D.,Shi, Y.,Wu, C., Wang, H. , Li, D. & Li, J .#.(2014).The Roles of Testicular C-kit Positive Cells in De novo Morphogenesis of Testis. SCIENTIFIC REPORTS：4 : 5936.

26. Liu,H.*, Lv,P., Zhu,X., Wang,X., Yang,X., Zuo,E., Lu,Y., Lu, S., Lu,K.#.(2014).In vitro development of porcine transgenic nuclear-transferred embryos derived from newborn Guangxi Bama mini-pig kidney fibroblasts.In Vitro Cell.Dev.Biol.—Animal：50:811–821.