According to the relationship between the information systems of provincial management industrial units of state grid, through the construction of data management visualization platform, unified data standards and dat...According to the relationship between the information systems of provincial management industrial units of state grid, through the construction of data management visualization platform, unified data standards and data structure, standardized data generation and use process, improved enterprise system data quality level and data security level, ensured that all kinds of data were accurate, true, timely and standardized, better met the requirements of enterprise statistical analysis, capacity building, industry classification and business upgrading, and improved data utilization ability and data value.展开更多
CRISPR/Cas is a simple,robust,versatile tool for plant biology studies and precision plant breeding.However,establishing a high-efficiency gene editing system for multiplex editing of the autotetraploid crop alfalfa(M...CRISPR/Cas is a simple,robust,versatile tool for plant biology studies and precision plant breeding.However,establishing a high-efficiency gene editing system for multiplex editing of the autotetraploid crop alfalfa(Medicago sativa L.),the most important forage legume worldwide,remains a formidable challenge.Here,we systematically identified endogenous U6 promoters in alfalfa through transient expression via Agrobacterium-mediated infiltration of alfalfa leaves.We further demonstrated the efficacy of the three most active promoters for genome editing using an optimized alfalfa hairy root system.Subsequently,we established an improved CRISPR/Cas9 multiplex system containing three or four tandemly arrayed MsU6-promoter-driven polycistronic tRNA-sgRNA(PTG)expression cassettes,each consisting of three tRNA-sgRNA units,to simultaneously edit three or four alfalfa genes,coupled with the visual reporter RH1 or RUBY.This toolkit showed efficient multiplex editing in the hairy root system with visual selection.We successfully obtained regenerated,red-colored shoots resulting from the stable transformation of alfalfa.These results highlight the potential application of the visual reporter system for the stable transformation of alfalfa.Our improved CRISPR/Cas9 multiplex system enables convenient,high-efficiency multiplex genome editing in alfalfa,providing a versatile toolset to facilitate functional studies of multiple genes and gene families for basic research and the genetic improvement of alfalfa.展开更多
Current systems to screen for transgenic soybeans(Glycine max)involve laborious molecular assays or the expression of fluorescent markers that are difficult to see in soybean plants.Therefore,a visual system for early...Current systems to screen for transgenic soybeans(Glycine max)involve laborious molecular assays or the expression of fluorescent markers that are difficult to see in soybean plants.Therefore,a visual system for early screening of transgenic plants would increase the efficiency of crop improvement by genome editing.The RUBY reporter system,which consists of three genes encoding betalain biosynthetic enzymes,leading to the accumulation of purple pigment in transgenic tissue,has been employed in some plants and dikaryon fungi.Here,we assessed the RUBY reporter for visual verification during soybean transformation.We show that RUBY can be expressed in soybean,allowing for visual confirmation of transgenic events without the need for specialized equipment.Plants with visible accumulation of purple pigment in any tissue were successfully transformed,confirming the accuracy of the RUBY system as a visual indicator.We also assessed the genetic stability of the transgene across generations,which can be performed very early,using the cotyledons of the progeny.Transgene-free seedlings have a distinct green color,facilitating the selection of genome-edited but transgene-free soybean seedlings for harvest.Using the RUBY system,we quickly identified a transgene-free Gmwaxy mutant in the T1 generation.This system thus provides an efficient and convenient tool for soybean genome editing.展开更多
文摘According to the relationship between the information systems of provincial management industrial units of state grid, through the construction of data management visualization platform, unified data standards and data structure, standardized data generation and use process, improved enterprise system data quality level and data security level, ensured that all kinds of data were accurate, true, timely and standardized, better met the requirements of enterprise statistical analysis, capacity building, industry classification and business upgrading, and improved data utilization ability and data value.
基金supported by grants from the Biological Breeding-National Science and Technology Major Project(2022ZD04011,2023ZD04073)National Natural Science Foundation of China(32325035,32071864)+4 种基金Fundamental Research Funds for Central Non-profit Scientific Institution(1610392024008)Central Laboratory of Biotechnology Research Institute,Chinese Academy of Agricultural Sciences,Hainan Seed Industry Laboratory(B23CJ0208)Yunnan Academy of Tobacco Agricultural Sciences(CNTC-YC2022530000241013,CNTC110201501015(JY-02))Hohhot Key R&D Project(2023-JBGS-S-1)Inner Mongolia Center of Pratacultural Technology Innovation(CCPTZX2023B01).
文摘CRISPR/Cas is a simple,robust,versatile tool for plant biology studies and precision plant breeding.However,establishing a high-efficiency gene editing system for multiplex editing of the autotetraploid crop alfalfa(Medicago sativa L.),the most important forage legume worldwide,remains a formidable challenge.Here,we systematically identified endogenous U6 promoters in alfalfa through transient expression via Agrobacterium-mediated infiltration of alfalfa leaves.We further demonstrated the efficacy of the three most active promoters for genome editing using an optimized alfalfa hairy root system.Subsequently,we established an improved CRISPR/Cas9 multiplex system containing three or four tandemly arrayed MsU6-promoter-driven polycistronic tRNA-sgRNA(PTG)expression cassettes,each consisting of three tRNA-sgRNA units,to simultaneously edit three or four alfalfa genes,coupled with the visual reporter RH1 or RUBY.This toolkit showed efficient multiplex editing in the hairy root system with visual selection.We successfully obtained regenerated,red-colored shoots resulting from the stable transformation of alfalfa.These results highlight the potential application of the visual reporter system for the stable transformation of alfalfa.Our improved CRISPR/Cas9 multiplex system enables convenient,high-efficiency multiplex genome editing in alfalfa,providing a versatile toolset to facilitate functional studies of multiple genes and gene families for basic research and the genetic improvement of alfalfa.
基金supported by grants from the National Key Research and Development Program of China(2023YFD1202900)Hainan Yazhou Bay Seed Laboratory(B23YQ1504,B23CQ154P).
文摘Current systems to screen for transgenic soybeans(Glycine max)involve laborious molecular assays or the expression of fluorescent markers that are difficult to see in soybean plants.Therefore,a visual system for early screening of transgenic plants would increase the efficiency of crop improvement by genome editing.The RUBY reporter system,which consists of three genes encoding betalain biosynthetic enzymes,leading to the accumulation of purple pigment in transgenic tissue,has been employed in some plants and dikaryon fungi.Here,we assessed the RUBY reporter for visual verification during soybean transformation.We show that RUBY can be expressed in soybean,allowing for visual confirmation of transgenic events without the need for specialized equipment.Plants with visible accumulation of purple pigment in any tissue were successfully transformed,confirming the accuracy of the RUBY system as a visual indicator.We also assessed the genetic stability of the transgene across generations,which can be performed very early,using the cotyledons of the progeny.Transgene-free seedlings have a distinct green color,facilitating the selection of genome-edited but transgene-free soybean seedlings for harvest.Using the RUBY system,we quickly identified a transgene-free Gmwaxy mutant in the T1 generation.This system thus provides an efficient and convenient tool for soybean genome editing.
