With the ever-increasing human population and deteriorating environmental conditions,there is an urgent need to breed environmentally friendly and resource-conserving rice cultivars to achieve sustainable agricultural...With the ever-increasing human population and deteriorating environmental conditions,there is an urgent need to breed environmentally friendly and resource-conserving rice cultivars to achieve sustainable agricultural development and food security.However,conventional rice improvement strategies,such as hybrid breeding,are time-consuming and laborious processes and may not be able to keep pace with increasing food demand in the future.Targeted genome-editing technologies,especially clustered regularly interspaced short palindromic repeat(CRISPR)/CRISPR-associated protein(CRISPR/Cas),permit efficient targeted genome modification and offer great promise for the creation of desired plants with higher yield,improved grain quality,and resistance to herbicides,diseases,and insect pests.There is also great potential for tapping heterosis using the CRISPR/Cas technology.In this review,we focus on the most essential applications of CRISPR/Cas genome editing to rice genetic improvement,considering traits such as yield,quality,and herbicide,disease and insect-pest resistance.We discuss applications of CRISPR/Cas to the exploitation of heterosis.Finally,we outline perspectives for future rice breeding using genome-editing technologies.展开更多
In the past several years, the CRISPR(clustered regularly interspaced short palindromic repeats)/Cas(CRISPR-associated protein) system has been harnessed as an efficient and powerful tool for targeted genome editing i...In the past several years, the CRISPR(clustered regularly interspaced short palindromic repeats)/Cas(CRISPR-associated protein) system has been harnessed as an efficient and powerful tool for targeted genome editing in different prokaryotic and eukaryotic species. Here, we review the development and application of CRISPR/Cas system in rice, emphasizing different varieties of CRISPR/Cas systems have been applied, strategies for multiplex editing, methods for precise gene insertion and replacement, and efficient systems for base editing and site-specific transcriptional regulation. In addition, the biosecurity of CRISPR/Cas system is also discussed, including transgene-free methods and off-target effects of CRISPR/Cas system. Thus, the development and application of CRISPR/Cas system will have significant impact on functional genomic research and variety improvement in rice.展开更多
The identification of new and even more precise technologies for modifying and manipulating the genome has been a challenge since the discovery of the DNA double helix.The ability to modify selectively specific genes ...The identification of new and even more precise technologies for modifying and manipulating the genome has been a challenge since the discovery of the DNA double helix.The ability to modify selectively specific genes provides a powerful tool for characterizing gene functions,performing gene therapy,correcting specific genetic mutations,eradicating diseases,engineering cells and organisms to achieve new and different functions and obtaining transgenic animals as models for studying specific diseases.Clustered regularly interspaced short palindromic repeats(CRISPR)/Cas9 technology has recently revolutionized genome engineering.The application of this new technology to stem cell research allows disease models to be developed to explore new therapeutic tools.The possibility of translating new systems of molecular knowledge to clinical research is particularly appealing for addressing degenerative diseases.In this review,we describe several applications of CRISPR/Cas9 to stem cells related to degenerative diseases.In addition,we address the challenges and future perspectives regarding the use of CRISPR/Cas9 as an important technology in the medical sciences.展开更多
Determination of Cas9 cutting efficiency to the target sites is important for genome editing.However,this determination can only be made via an in vitro method,as the purification of Cas protein and synthesis of gRNA ...Determination of Cas9 cutting efficiency to the target sites is important for genome editing.However,this determination can only be made via an in vitro method,as the purification of Cas protein and synthesis of gRNA are necessary.Here,we developed an in vivo method,called transient CRISPR/Cas editing in plants(TCEP)to determine Cas9 cutting efficiency.The CRISPR/Cas vector for plant transformation mediated by Agrobacterium tumefaciens was constructed as normal.Using the transient transformation method we built,the Cas9 protein and gRNA were transiently expressed and formed a complex to cut its target sites,resulting in dynamic DNA breakage.The broken DNA was quantified using qPCR to measure the efficiencies of Cas9 cutting.We studied the Cas9 cutting efficiencies to different target sites in Betula platyphylla and Populus davidiana×P.bolleana plants using TCEP and an in vitro method.The results of TCEP were consistent with those of the in vitro method,suggesting that the TCEP method is reliable in determining cutting efficiency.Additionally,using the TCEP method,we showed that both heat and sonication treatment significantly improved CRISPR/Cas efficiency.Therefore,the TCEP method has broad application value and can not only be used to analyze the CRISPR/Cas efficiency but also to determine the factors involved in Cas9 cutting.展开更多
基金supported by the National Natural Science Foundation of China(U20A2030)Central Public-interest Scientific Institution Basal Research Fund(Y2020YJ12 and Y2020XK17)+1 种基金Key Research and Development Program of China National Rice Research Institute(CNRRI-2020-01)Foreign Cooperation Project of Ningxia Academy of Agricultural and Forestry Institute(DW-X-2018004)。
文摘With the ever-increasing human population and deteriorating environmental conditions,there is an urgent need to breed environmentally friendly and resource-conserving rice cultivars to achieve sustainable agricultural development and food security.However,conventional rice improvement strategies,such as hybrid breeding,are time-consuming and laborious processes and may not be able to keep pace with increasing food demand in the future.Targeted genome-editing technologies,especially clustered regularly interspaced short palindromic repeat(CRISPR)/CRISPR-associated protein(CRISPR/Cas),permit efficient targeted genome modification and offer great promise for the creation of desired plants with higher yield,improved grain quality,and resistance to herbicides,diseases,and insect pests.There is also great potential for tapping heterosis using the CRISPR/Cas technology.In this review,we focus on the most essential applications of CRISPR/Cas genome editing to rice genetic improvement,considering traits such as yield,quality,and herbicide,disease and insect-pest resistance.We discuss applications of CRISPR/Cas to the exploitation of heterosis.Finally,we outline perspectives for future rice breeding using genome-editing technologies.
基金supported by the Agricultural Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciencesthe National Natural Science Foundation of China (Grant No. 31871703)
文摘In the past several years, the CRISPR(clustered regularly interspaced short palindromic repeats)/Cas(CRISPR-associated protein) system has been harnessed as an efficient and powerful tool for targeted genome editing in different prokaryotic and eukaryotic species. Here, we review the development and application of CRISPR/Cas system in rice, emphasizing different varieties of CRISPR/Cas systems have been applied, strategies for multiplex editing, methods for precise gene insertion and replacement, and efficient systems for base editing and site-specific transcriptional regulation. In addition, the biosecurity of CRISPR/Cas system is also discussed, including transgene-free methods and off-target effects of CRISPR/Cas system. Thus, the development and application of CRISPR/Cas system will have significant impact on functional genomic research and variety improvement in rice.
文摘The identification of new and even more precise technologies for modifying and manipulating the genome has been a challenge since the discovery of the DNA double helix.The ability to modify selectively specific genes provides a powerful tool for characterizing gene functions,performing gene therapy,correcting specific genetic mutations,eradicating diseases,engineering cells and organisms to achieve new and different functions and obtaining transgenic animals as models for studying specific diseases.Clustered regularly interspaced short palindromic repeats(CRISPR)/Cas9 technology has recently revolutionized genome engineering.The application of this new technology to stem cell research allows disease models to be developed to explore new therapeutic tools.The possibility of translating new systems of molecular knowledge to clinical research is particularly appealing for addressing degenerative diseases.In this review,we describe several applications of CRISPR/Cas9 to stem cells related to degenerative diseases.In addition,we address the challenges and future perspectives regarding the use of CRISPR/Cas9 as an important technology in the medical sciences.
基金supported by the National Natural Science Foundation of China(No.31770704)Heilongjiang Touyan Innovation Team Program(Tree Genetics and Breeding Innovation Team)the Overseas Expertise Introduction Project for Discipline Innovation(B16010).
文摘Determination of Cas9 cutting efficiency to the target sites is important for genome editing.However,this determination can only be made via an in vitro method,as the purification of Cas protein and synthesis of gRNA are necessary.Here,we developed an in vivo method,called transient CRISPR/Cas editing in plants(TCEP)to determine Cas9 cutting efficiency.The CRISPR/Cas vector for plant transformation mediated by Agrobacterium tumefaciens was constructed as normal.Using the transient transformation method we built,the Cas9 protein and gRNA were transiently expressed and formed a complex to cut its target sites,resulting in dynamic DNA breakage.The broken DNA was quantified using qPCR to measure the efficiencies of Cas9 cutting.We studied the Cas9 cutting efficiencies to different target sites in Betula platyphylla and Populus davidiana×P.bolleana plants using TCEP and an in vitro method.The results of TCEP were consistent with those of the in vitro method,suggesting that the TCEP method is reliable in determining cutting efficiency.Additionally,using the TCEP method,we showed that both heat and sonication treatment significantly improved CRISPR/Cas efficiency.Therefore,the TCEP method has broad application value and can not only be used to analyze the CRISPR/Cas efficiency but also to determine the factors involved in Cas9 cutting.
