Development of tools for targeted modifications of specific DNA sequences in plants is of great importance to basic plant biology research as well as crop improvement.The ability to cut DNA at specific locations in th...Development of tools for targeted modifications of specific DNA sequences in plants is of great importance to basic plant biology research as well as crop improvement.The ability to cut DNA at specific locations in the genome to generate doublestrand breaks（DSBs）in vivo is a prerequisite for any genome editing efforts.展开更多
Single-guide RNA(sg RNA) is one of the two core components of the CRISPR(clustered regularly interspaced short palindromic repeat)/Cas(CRISPR-associated) genome-editing technology. We established an in vitro Traffic L...Single-guide RNA(sg RNA) is one of the two core components of the CRISPR(clustered regularly interspaced short palindromic repeat)/Cas(CRISPR-associated) genome-editing technology. We established an in vitro Traffic Light Reporter(TLR) system, which is designated as the same colors as traffic lights such as green, red and yellow were produced in cells. The TLR can be readily used in maize mesophyll protoplast for a quick test of promoter activity. The TLR assay indicates the variation in transcription activities of the seven Pol III promoters, from 3.4%(U6-1) to over 21.0%(U6-6). The U6-2 promoter, which was constructed to drive sg RNA expression targeting the Zm Wx1 gene, yielded mutation efficiencies ranging from 48.5% to 97.1%. Based on the reported and unpublished data, the in vitro TLR assay results were confirmed to be a readily system and may be extended to other plant species amenable to efficient genome editing via CRISPR/Cas. Our efforts provide an efficient method of identifying native Pol III-recognized promoters for RNA guide-based genome-editing systems in maize.展开更多
Rice （Oryza sativa） is one of the most important staple food crops for more than half of the world＇s population. The demand is increasing for food security because of population growth and environmental challenges ...Rice （Oryza sativa） is one of the most important staple food crops for more than half of the world＇s population. The demand is increasing for food security because of population growth and environmental challenges triggered by climate changes. This scenario has led to more interest in developing crops with greater productivity and sustainability. The process of genetic transformation, a major tool for crop improvement, utilizes promoters as one of its key elements. Those promoters are generally divided into three types： constitutive, spatiotemporal, and condition-dependent. Tran- scriptional control of a constitutive promoter often leads to reduced plant growth, due to a negative effect of accumu- lated molecules during cellular functions or energy consump- tion. To maximize the effect of a transgene on transgenic plants, it is better to use condition-dependent or tissue- specific promoters. However, until now, those types have not been as widely applied in crop biotechnology. In this review, we introduce and discuss four groups of tissue-specific promoters （5o promoters in total） and six groups of condition-dependent promoters （27 promoters）. These pro- moters can be utilized to fine-tune desirable agronomic traits and develop crops with tolerance to various stresses, enhanced nutritional value, and advanced productivity.展开更多
基金supported by a National Transgenic Science and Technology Program (2016ZX08010002)to R.W.a startup fund from the Huazhong Agricultural University
文摘Development of tools for targeted modifications of specific DNA sequences in plants is of great importance to basic plant biology research as well as crop improvement.The ability to cut DNA at specific locations in the genome to generate doublestrand breaks（DSBs）in vivo is a prerequisite for any genome editing efforts.
基金supported by the National Science Foundation of China(31771808)Ministry of Science and Technology(2015BAD02B0203)+1 种基金National Engineering Laboratory of Crop Molecular Breedingthe Chinese Academy of Agricultural Sciences(Y2017XM03)
文摘Single-guide RNA(sg RNA) is one of the two core components of the CRISPR(clustered regularly interspaced short palindromic repeat)/Cas(CRISPR-associated) genome-editing technology. We established an in vitro Traffic Light Reporter(TLR) system, which is designated as the same colors as traffic lights such as green, red and yellow were produced in cells. The TLR can be readily used in maize mesophyll protoplast for a quick test of promoter activity. The TLR assay indicates the variation in transcription activities of the seven Pol III promoters, from 3.4%(U6-1) to over 21.0%(U6-6). The U6-2 promoter, which was constructed to drive sg RNA expression targeting the Zm Wx1 gene, yielded mutation efficiencies ranging from 48.5% to 97.1%. Based on the reported and unpublished data, the in vitro TLR assay results were confirmed to be a readily system and may be extended to other plant species amenable to efficient genome editing via CRISPR/Cas. Our efforts provide an efficient method of identifying native Pol III-recognized promoters for RNA guide-based genome-editing systems in maize.
基金supported by the Cooperative Research Program for Agriculture Science and Technology Development(Project title:Global identification and functional study of rice genes for enhancement of root development and nutrient use efficiency using genome information,project no.PJ01100401)
文摘Rice （Oryza sativa） is one of the most important staple food crops for more than half of the world＇s population. The demand is increasing for food security because of population growth and environmental challenges triggered by climate changes. This scenario has led to more interest in developing crops with greater productivity and sustainability. The process of genetic transformation, a major tool for crop improvement, utilizes promoters as one of its key elements. Those promoters are generally divided into three types： constitutive, spatiotemporal, and condition-dependent. Tran- scriptional control of a constitutive promoter often leads to reduced plant growth, due to a negative effect of accumu- lated molecules during cellular functions or energy consump- tion. To maximize the effect of a transgene on transgenic plants, it is better to use condition-dependent or tissue- specific promoters. However, until now, those types have not been as widely applied in crop biotechnology. In this review, we introduce and discuss four groups of tissue-specific promoters （5o promoters in total） and six groups of condition-dependent promoters （27 promoters）. These pro- moters can be utilized to fine-tune desirable agronomic traits and develop crops with tolerance to various stresses, enhanced nutritional value, and advanced productivity.
