Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated 9(CRISPR/Cas9)system has recently become one popular technology due to its efficiency,precision,and simplicity compared with other genome edi...Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated 9(CRISPR/Cas9)system has recently become one popular technology due to its efficiency,precision,and simplicity compared with other genome editing tools such as Zinc Finger Nucleases(ZFNs)and Transcription Activator Like Effector Nucleases(TALENs).Horticultural crops provide energy and health-keeping nutrients to humankind.Genome-editing technology has become widely adopted in horticultural breeding with the increasing demand for high yield and better-quality horticultural crops.Here,we describe the CRISPR/Cas9 system construction,its optimization,including sgRNA promoter,sgRNA design,Cas9 protein promoter,SpCas9 variants and orthologs,and vector delivery methods.We also summarized the application of this technology in horticultural plants for stress responses enhancement,fruit quality improvement,and cultivation traits modification.This detailed review was compiled to help establish comprehensive understanding of the CRISPR/Cas9 systems and provide a reference for further developing this technology to manipulate horticultural plant traits effectively.展开更多
Targeted genome editing is a continually evolving technology employing programmable nucleases to specifically change,insert,or remove a genomic sequence of interest.These advanced molecular tools include meganucleases...Targeted genome editing is a continually evolving technology employing programmable nucleases to specifically change,insert,or remove a genomic sequence of interest.These advanced molecular tools include meganucleases,zinc finger nucleases,transcription activator-like effector nucleases and RNA-guided engineered nucleases(RGENs),which create double-strand breaks at specific target sites in the genome,and repair DNA either by homologous recombination in the presence of donor DNA or via the error-prone non-homologous end-joining mechanism.A recently discovered group of RGENs known as CRISPR/Cas9 gene-editing systems allowed precise genome manipulation revealing a causal association between disease genotype and phenotype,without the need for the reengineering of the specific enzyme when targeting different sequences.CRISPR/Cas9 has been successfully employed as an ex vivo gene-editing tool in embryonic stem cells and patient-derived stem cells to understand pancreatic beta-cell development and function.RNA-guided nucleases also open the way for the generation of novel animal models for diabetes and allow testing the efficiency of various therapeutic approaches in diabetes,as summarized and exemplified in this manuscript.展开更多
Gene editing technology provides important technical basics for the research in plant functional genes and crop genetic improvement.CRISPR/Cas9-mediated gene editing is an effective experimental tool for crop genome d...Gene editing technology provides important technical basics for the research in plant functional genes and crop genetic improvement.CRISPR/Cas9-mediated gene editing is an effective experimental tool for crop genome directed editing in recent years,which has been widely used in many crops as rice,wheat and other crops.CRISPR/Cas9 system was expected to be a powerful experimental tool in genetic improvement and molecular design breeding of rapeseed.This paper,which based on the development history and the latest research of CRISPR/Cas9-mediated gene editing technology in rapeseed,summarized the progress of CRISPR/Cas9 including plant type improvement,yield traits,quality improvement,disease and stress resistance improvement,yellow seed creation and other utilizes at present.The application scope,development direction and target analysis method of this technology in rape were focused.The problems of CRISPR/cas9 system in rapeseed breeding were analyzed and the improvement strategies were discussed.Finally,views on direction of rapeseed breeding by gene editing were emphasized.展开更多
The recently developed RNA-guided clustered regularly interspaced short palindromic repeat （CRISPR）/CRISPR-associated 9 （Cas9） nuclease system has progressed to be an invaluable technology for genome manipulation ...The recently developed RNA-guided clustered regularly interspaced short palindromic repeat （CRISPR）/CRISPR-associated 9 （Cas9） nuclease system has progressed to be an invaluable technology for genome manipulation in somatic cell types and germline model organisms. While the unprecedented advance in human embryo gene editing research has great potential in next-generation therapeutics, it raises various ethical concerns that need to be addressed before being translated for clinical use. Here, we discuss the current and potential applications of CRISPR/Cas9 technology and its limitations in clinical applications, as well as ethical and legal considerations in the treatment, disease prevention or disability in somatic cells or human embryo via gene editing.展开更多
Dear Editor,Despite tremendous effort devoted to the development of antiretroviral therapies to combat HIV over the past decades, AIDS remains one of the most important global infectious diseases. According to UNAIDS ...Dear Editor,Despite tremendous effort devoted to the development of antiretroviral therapies to combat HIV over the past decades, AIDS remains one of the most important global infectious diseases. According to UNAIDS report on the global AIDS epidemic in 2016, the estimated number of people living with HIV rose from 7.5 million in 2010 to 36.7 million in 2015. Furthermore, drug-resistance HIV strains have recently been reported （Wensing et al., 2017）. Therefore, it is important to develop new therapies to eliminate HIV in the patients. Immortalized cell lines representing the major targets of HIV in human are important for HIV research and therapeutic development.展开更多
The clustered regularly interspaced short palindromic repeats （CRISPR）/Cas9 system, a simple and efficient tool for genome editing, has experienced rapid progress in its technology and applicability in the past two ...The clustered regularly interspaced short palindromic repeats （CRISPR）/Cas9 system, a simple and efficient tool for genome editing, has experienced rapid progress in its technology and applicability in the past two years. Here, we review the recent advances in CRISPR/Cas9 technology and the ways that have been adopted to expand our capacity for precise genome manipulation. First, we introduce the mechanism of CRISPR/Cas9, including its biochemical and structural implications. Second, we highlight the latest improvements in the CRISPR/Cas9 system, especially Cas9 protein modifications for customization. Third, we review its current applications, in which the versatile CRISPR/Cas9 system was employed to edit the genome, epigenome, or RNA of various organisms. Although CRISPR/Cas9 allows convenient genome editing accompanied by many benefits, we should not ignore the significant ethical and biosafety concerns that it raises. Finally, we discuss the prospective applications and challenges of several promising techniques adapted from CRISPR/Cas9.展开更多
Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal ...Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal of the CRISPR/Cas9 transgenes by genetic segregation and by backcross is laborious and time consuming. We previously reported the development of the transgene killer CRISPR(TKC) technology that uses a pair of suicide genes to trigger self-elimination of the transgenes without compromising gene editing efficiency. The TKC technology enables isolation of transgene-free CRISPR-edited plants within a single generation, greatly accelerating crop improvements. Here, we presented two new TKC vectors that show great efficiency in both editing the target gene and in undergoing self-elimination of the transgenes. The new vectors replaced the CaMV35 S promoter used in our previous TKC vector with two rice promoters to drive one of the suicide genes, providing advantages over our previous TKC vector under certain conditions. The vectors reported here offered more options and flexibility to conduct gene editing experiments in rice.展开更多
基金supported by grants from the National Key R&D Program of China (Grant No. 2018YFD1000100)National Natural Science Foundation of China (Grant No. 31972378)Agricultural Variety Improvement Project of Shandong Province (Grant No. 2019LZGC007)
文摘Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated 9(CRISPR/Cas9)system has recently become one popular technology due to its efficiency,precision,and simplicity compared with other genome editing tools such as Zinc Finger Nucleases(ZFNs)and Transcription Activator Like Effector Nucleases(TALENs).Horticultural crops provide energy and health-keeping nutrients to humankind.Genome-editing technology has become widely adopted in horticultural breeding with the increasing demand for high yield and better-quality horticultural crops.Here,we describe the CRISPR/Cas9 system construction,its optimization,including sgRNA promoter,sgRNA design,Cas9 protein promoter,SpCas9 variants and orthologs,and vector delivery methods.We also summarized the application of this technology in horticultural plants for stress responses enhancement,fruit quality improvement,and cultivation traits modification.This detailed review was compiled to help establish comprehensive understanding of the CRISPR/Cas9 systems and provide a reference for further developing this technology to manipulate horticultural plant traits effectively.
基金the Akdeniz University Scientific Research Commission and the Scientific and Technological Research Council of Turkey,No.TUBITAK-215S820.
文摘Targeted genome editing is a continually evolving technology employing programmable nucleases to specifically change,insert,or remove a genomic sequence of interest.These advanced molecular tools include meganucleases,zinc finger nucleases,transcription activator-like effector nucleases and RNA-guided engineered nucleases(RGENs),which create double-strand breaks at specific target sites in the genome,and repair DNA either by homologous recombination in the presence of donor DNA or via the error-prone non-homologous end-joining mechanism.A recently discovered group of RGENs known as CRISPR/Cas9 gene-editing systems allowed precise genome manipulation revealing a causal association between disease genotype and phenotype,without the need for the reengineering of the specific enzyme when targeting different sequences.CRISPR/Cas9 has been successfully employed as an ex vivo gene-editing tool in embryonic stem cells and patient-derived stem cells to understand pancreatic beta-cell development and function.RNA-guided nucleases also open the way for the generation of novel animal models for diabetes and allow testing the efficiency of various therapeutic approaches in diabetes,as summarized and exemplified in this manuscript.
基金This work was supported by the National Major Project for Developing New GM Crops(2018ZX08020001).
文摘Gene editing technology provides important technical basics for the research in plant functional genes and crop genetic improvement.CRISPR/Cas9-mediated gene editing is an effective experimental tool for crop genome directed editing in recent years,which has been widely used in many crops as rice,wheat and other crops.CRISPR/Cas9 system was expected to be a powerful experimental tool in genetic improvement and molecular design breeding of rapeseed.This paper,which based on the development history and the latest research of CRISPR/Cas9-mediated gene editing technology in rapeseed,summarized the progress of CRISPR/Cas9 including plant type improvement,yield traits,quality improvement,disease and stress resistance improvement,yellow seed creation and other utilizes at present.The application scope,development direction and target analysis method of this technology in rape were focused.The problems of CRISPR/cas9 system in rapeseed breeding were analyzed and the improvement strategies were discussed.Finally,views on direction of rapeseed breeding by gene editing were emphasized.
文摘The recently developed RNA-guided clustered regularly interspaced short palindromic repeat （CRISPR）/CRISPR-associated 9 （Cas9） nuclease system has progressed to be an invaluable technology for genome manipulation in somatic cell types and germline model organisms. While the unprecedented advance in human embryo gene editing research has great potential in next-generation therapeutics, it raises various ethical concerns that need to be addressed before being translated for clinical use. Here, we discuss the current and potential applications of CRISPR/Cas9 technology and its limitations in clinical applications, as well as ethical and legal considerations in the treatment, disease prevention or disability in somatic cells or human embryo via gene editing.
文摘Dear Editor,Despite tremendous effort devoted to the development of antiretroviral therapies to combat HIV over the past decades, AIDS remains one of the most important global infectious diseases. According to UNAIDS report on the global AIDS epidemic in 2016, the estimated number of people living with HIV rose from 7.5 million in 2010 to 36.7 million in 2015. Furthermore, drug-resistance HIV strains have recently been reported （Wensing et al., 2017）. Therefore, it is important to develop new therapies to eliminate HIV in the patients. Immortalized cell lines representing the major targets of HIV in human are important for HIV research and therapeutic development.
基金supported by the grant from the Major State Basic Research Development Program of China(Nos.2012CB517902 and 2012CB517904)the Fundamental Research Funds for the Central Universities(No.14QNJJ042)Special Research Program of National Health and Family Planning Commission of China(No.201302002)
文摘The clustered regularly interspaced short palindromic repeats （CRISPR）/Cas9 system, a simple and efficient tool for genome editing, has experienced rapid progress in its technology and applicability in the past two years. Here, we review the recent advances in CRISPR/Cas9 technology and the ways that have been adopted to expand our capacity for precise genome manipulation. First, we introduce the mechanism of CRISPR/Cas9, including its biochemical and structural implications. Second, we highlight the latest improvements in the CRISPR/Cas9 system, especially Cas9 protein modifications for customization. Third, we review its current applications, in which the versatile CRISPR/Cas9 system was employed to edit the genome, epigenome, or RNA of various organisms. Although CRISPR/Cas9 allows convenient genome editing accompanied by many benefits, we should not ignore the significant ethical and biosafety concerns that it raises. Finally, we discuss the prospective applications and challenges of several promising techniques adapted from CRISPR/Cas9.
基金supported by Chinese Ministry of Agriculture and Rural Affairs (Grant No. 2018ZX0801003B)the National Transgenic Science and Technology Program (Grant No. 2016ZX08010002)
文摘Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal of the CRISPR/Cas9 transgenes by genetic segregation and by backcross is laborious and time consuming. We previously reported the development of the transgene killer CRISPR(TKC) technology that uses a pair of suicide genes to trigger self-elimination of the transgenes without compromising gene editing efficiency. The TKC technology enables isolation of transgene-free CRISPR-edited plants within a single generation, greatly accelerating crop improvements. Here, we presented two new TKC vectors that show great efficiency in both editing the target gene and in undergoing self-elimination of the transgenes. The new vectors replaced the CaMV35 S promoter used in our previous TKC vector with two rice promoters to drive one of the suicide genes, providing advantages over our previous TKC vector under certain conditions. The vectors reported here offered more options and flexibility to conduct gene editing experiments in rice.
