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gene targeting using Agrobacterium tumefaciens-mediated CRISPR-Cas system in crop plants

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Gene targeting using Agrobacterium tumefaciens-mediated CRIS...

Application of Artificially Induced Double-strand Breaks (DSB) and Triplex-forming Oligonucleotides (TFO) in the Improvement of gene targeting Efficiency

生态文明建设中的植物学：现在与未来——中国植物学会第十...

作者： Rong-Fang XU Hao LI Rui-Ying QIN Lu WANG Li LI Peng-Cheng WEI Jian-Bo YANG Key Laboratory of Rice genetics Breeding of Anhui Province Rice Research InstituteAnhui Academy of Agricultural Sciences

The type Ⅱ clustered,regularly interspaced,short palindromic repeat(CRISPR)/CRISPR-associated protein 9(Cas9) system is a novel molecular tool of site-specific genome modification.Very recently,the CRISPR-Cas9 system was reported to be successfully performed by transient or biolistic-mediated stable transformation in Arabidopsis,Nicotiana benthamiana and rice.Here,we reported the gene targeting and genome editing in rice via Agrobacterium tumefaciens-mediated CRISPR-Cas system.Three 20nt guide RNA were designed to pair with the distinct sites followed by the protospacer adjacent motif(PAM) of a rice herbicide-resistance gene BEL.After integrating the sgRNA and Cas9 cassettes in one binary vector,the transgenic rice plants

关键词： gene targeting CRISPR/Cas9 Rice Agrobacterium Stable transformation

来源： cnki会议评论

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Agricultural Biotechnology 2013年第1期2卷 1-6,12页

作者： Hegang LI Wenke CHENG Ke JIANG Xiaoli REN Yongping JIANG Lele HOU Xiaojing HAO Jinshan ZHAO Qingdao Institute of Animal Science and Veterinary Medicine Animal Husbandry Technology Extension Station of Hainan Province Laixi Husbandry and Veterinary Bureau of Qingdao City Rizhuang Agricultural Service Center of Laixi City

gene targeting technology is an important means to investigate gene functions, but its efficiency of gene targeting is very low, especially for somatic cell targeting. Artificially induced double-strand breaks （DSB） and triplex forming oligonucleotide （TFO） are currently developed methods to improve the targeting efficiency. This paper summarized the basic principles, design ideas and application in gene targeting efficiency improvement of these two methods, analyzed and com- pared their characteristics, and finally proposed prospects for their future development.

关键词： gene targeting Double-strand breaks Zinc finger nuclease Homing endonuclease Triplex-forming oligonucleotides

Advances in Drosophila gene targeting and related techniques

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Frontiers in Biology 2010年第3期5卷 238-245页

作者： Zhongsheng YU Renjie JIAO State Key Laboratory of Brain and Cognitive Science Institute of BiophysicsChinese Academy of SciencesBeijing 100101China Graduate School of Chinese Academy of Sciences Beijing 100080China

Functional biological research has benefited tremendously by analyses of the phenotypes of mutant organisms which can be generated through targeted mutation of genes.In Drosophila,compared with random mutagenesis methods gene targeting has gained its popularity because it can introduce any desired mutation into a gene of interest.However,applications of gene targeting have been limited because the targeting efficiency varies with different genes,and the time and labor of targeting procedure are intensive.Nevertheless,improvement of gene targeting and development of its variant technologies have received much attention of scientists.Here we review recent progress that has been made in expanding the applications of gene targeting,which include the ϕC31 integration system and zinc-finger nucleases induced gene targeting,and new strategies that generate more efficient and reliable gene targeting.

关键词： gene targeting ends-in ends-out ϕC31 integration system zinc-finger nucleases(ZFNs) homologous recombination(HR) Drosophila melanogaster

gene targeting in Marine Embryonic Stem Cells, a New Method to Manipulate the Mouse Genome

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Progress in Natural Science:Materials International 1994年第1期 22-33页

作者：邓初夏 200 Longwood Avenue Boston Dupartment of genetics Harvard Medical School MA 02115 USA

In eukaryotic cells, homologous recombination between DNA sequences residing in the chromosome and newly introduced DNA is commonly referred to as gene targeting.This emerging technology has proven to be a powerful means for precisely manipulating the mammalian genome. gene targeting in totipotent murine embryonic stem (ES) cells can, theoretically, be used to generate mice of any desired genotype, thus aiding in the study of gene structure and function, as well as mammalian developmental biology. In addition, gene targeting also provides a

关键词： gene targeting ES cells mouse mutants isogenic DNA

gene editing in plants:progress and challenges

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National Science Review 2019年第3期6卷 421-437页

作者： Yanfei Mao Jose Ramon Botella Yaoguang Liu Jian-Kang Zhu Shanghai Center for Plant Stress Biology CAS Center of Excellence in Molecular Plant Sciences Chinese Academy of Sciences School of Agriculture and Food Sciences University of Queensland State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources College of Life Sciences South China Agricultural University Department of Horticulture and Landscape Architecture Purdue University

The clustered regularly interspaced short palindromic repeat(CRISPR)-associated protein 9(Cas9)genome editing system is a powerful tool for targeted gene modifications in a wide range of species,including plants. Over the last few years, this system has revolutionized the way scientists perform genetic studies and crop breeding, due to its simplicity, flexibility, consistency and high efficiency. Considerable progress has been made in optimizing CRISPR/Cas9 systems in plants, particularly for targeted gene mutagenesis. However, there are still a number of important challenges ahead, including methods for the efficient delivery of CRISPR and other editing tools to most plants, and more effective strategies for sequence knock-ins and replacements. We provide our viewpoint on the goals, potential concerns and future challenges for the development and application of plant genome editing tools.

关键词： CRISPR Cas9 genome editing base editing gene targeting crop breeding

Inhibiting effects on HeLa cell growth by HER-2 gene targeting

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Progress in Natural Science:Materials International 1999年第6期 33-40页

作者：王向东姜琨任庆虎陈培利汪维童坦君高建刚 Academy of Sciences Beijing 100083 Beijing Medical University China Chinese Department of Biochemistry & Molecular Biology institute of Developmental Biology

A plasmid (promoter pPNT 1305) for HER-2 gene targeting was constructed and introduced me HeLa cells by electroporation . After positive selection by G418 and negative selection by Gancyclovir, the targeted cell line (HER-2-1 cells) containing only one copy of HER-2 allies was selected. Northern blot assay showed that the mR-NA expression of HER-2 was significantly less than that of its parental cells. The results of Western blot also showed that HER-2 protein of HER-2 - cells was obviously less than that of HeLa cells. The dala of FA.CS assay demenstrated that after one of the two HER-2 copies in HeLa cells was knocked out, the G1 phase percentage of HER-2 cells was more and the M plus G2 phase percentage of HER-2 -1 cells was less than those of HeLa ceils respectively These reaults show that gene targeting of HER-2 has exhibited the inhibiting effects on the proliferational capability of HeLa cells

关键词： gene targeting HER-2 proto-oncogene.

Precision Breeding Made Real with CRISPR: Illustration through genetic Resistance to Pathogens

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Plant Communications 2020年第5期1卷 1-20页

作者： Florian Veillet Mickael Durand Thomas Kroj Stella Cesari Jean-Luc Gallois IGEPP INRAEInstitut AgroUniv RennesPloudaniel 29260France Germicopa Breeding KerguivarchChateauneuf Du Faou 29520France Institut Jean-Pierre Bourgin INRAEAgroParisTechUniversitéParis-SaclayVersailles 78000France INRAE BGPIBiology and Genetics of Plant-Pathogen InteractionsCampus International de BaillarguetMontpellierFrance INRAE GAFLMontfavet F-84143France

Since its discovery as a bacterial adaptive immune system and its development for genome editing in eukaryotes,the CRISPR technology has revolutionized plant research and precision crop breeding.The CRISPR toolbox holds great promise in the production of crops with genetic disease resistance to increase agriculture resilience and reduce chemical crop protection with a strong impact on the environment and public health.In this review,we provide an extensive overviewon recent breakthroughs in CRISPR technology,including the newly developed prime editing system that allows precision gene editing in plants.We present how each CRISPR tool can be selected for optimal use in accordance with its specific strengths and limitations,and illustrate how the CRISPR toolbox can foster the development of genetically pathogen-resistant crops for sustainable agriculture.

关键词： CRISPR-Cas9 gene targeting base editing prime editing plant/pathogen interactions precision crop breeding

Genome engineering technologies in rabbits

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The Journal of Biomedical Research 2021年第2期35卷 135-147页

作者： Jun Song Jifeng Zhang Jie Xu Minerva Garcia-Barrio Y.Eugene Chen Dongshan Yang Center for Advanced Models for Translational Sciences and Therapeutics University of Michigan Medical CenterAnn ArborMI 48109USA

The rabbit has been recognized as a valuable model in various biomedical and biological research fields because of its intermediate size and phylogenetic proximity to primates.However,the technology for precise genome manipulations in rabbit has been stalled for decades,severely limiting its applications in biomedical research.Novel genome editing technologies,especially CRISPR/Cas9,have remarkably enhanced precise genome manipulation in rabbits,and shown their superiority and promise for generating rabbit models of human genetic diseases.In this review,we summarize the brief history of transgenic rabbit technology and the development of novel genome editing technologies in rabbits.

关键词： rabbit transgenic animal gene targeting genome editing base editing

维普期刊数据库

An Open-Source System for In Planta gene Stacking by Bxbl and Cre Recombinases

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Molecular Plant 2014年第12期7卷 1756-1765页

作者： Lili Hou Yuan-Yeu Yau Junjie Wei Zhiguo Han Zhicheng Dong David W. Ow Plant gene Engineering Center South China Agricultural Plant Molecular Analysis and Genetic Improvement Key Laboratory SouthChina Botanical Garden Chinese Academy of Sciences 723 Xingke Road Guangzhou 510650 China University of Chinese Academy of Sciences 19 Yuquan Road Beijing 100049 China Former Affiliation Plant Gene Expression Center USDA-ARS & Plant & Microbial Biology University of California-Berkeley 800Buchanan St. Albany CA 94710 USA

The rapid development of crops with multiple transgenic traits arouses the need for an efficient system for creating stacked cultivars. Most major crops rely on classical breeding to introgress the transgene from a laboratory variety to the numerous cultivars adapted to different growing regions. Even with vegetative propagated crops, genetic crosses are conducted during varietal improvement prior to vegetative cloning. The probability to assort the ＇x＇ number of transgenic loci into a single genome may seem trivial, （~）x for a diploid species, but given the ＇y＇ number of other nontransgenic traits that breeders also need to assemble into the same genome, the （~）~＊y probability for a ＇breeding stack＇ could quickly make the line conversion process unmanageable. Adding new transgenes onto existing transgenic varieties without creating a new segregating locus would require site-specific integration of new DNA at the existing transgenic locus. Here, we tested a recombinase-mediated gene-stacking scheme in tobacco. Sequential site-specific inte- gration was mediated by the mycobacteriophage Bxbl integrase-catalyzed recombination between attP and attB sites. Transgenic DNA no longer needed after integration was excised by Cre recombinase-mediated recombination of Iox sites. Site-specific integration occurred in -10% of the integration events, with half of those events usable as substrates for a next round of gene stacking. Among the site-specific integrants, however, a third experienced gene silencing. Overall, precise structure and reproducible expression of the sequentially added triple traits were obtained at an overall rate of -3% of the transformed clones--a workable frequency for the development of commercial cultivars. Moreover, since nei- ther the Bxbl-att nor the Cre-lox system is under patent, there is freedom to operate,

关键词： transgenesis site-specific recombination integrase gene targeting transgene expression.