In plants and mammals,non-homologous end-joining is the dominant pathway to repair DNA doublestrand breaks,making it challenging to generate knock-in events.In this study,we identified two groups of exonucleases from ...In plants and mammals,non-homologous end-joining is the dominant pathway to repair DNA doublestrand breaks,making it challenging to generate knock-in events.In this study,we identified two groups of exonucleases from the herpes virus and the bacteriophage T7 families that conferred an up to 38-fold increase in homology-directed repair frequencies when fused to Cas9/Cas12a in a tobacco mosaic virus-based transient assay in Nicotiana benthamiana.We achieved precise and scar-free insertion of several kilobases of DNA both in transient and stable transformation systems.In Arabidopsis thaliana,fusion of Cas9 to a herpes virus family exonuclease led to 10-fold higher frequencies of knock-ins in the first generation of transformants.In addition,we demonstrated stable and heritable knock-ins in wheat in 1%of the primary transformants.Taken together,our results open perspectives for the routine production of heritable knock-in and gene replacement events in plants.展开更多
Dear Editor,Rauvolfia tetraphylla(aka the Devil pepper)(Supplemental Figure 1)is a well-known medicinal plant that produces monoterpenoid indole alkaloids(MIAs).This MIA biosynthesis occurs in several organs,including...Dear Editor,Rauvolfia tetraphylla(aka the Devil pepper)(Supplemental Figure 1)is a well-known medicinal plant that produces monoterpenoid indole alkaloids(MIAs).This MIA biosynthesis occurs in several organs,including leaves,stems,fruit,and roots,which accumulate the famous antiarrhythmic ajmaline(Kumar et al.2016a,2016b;Kumara et al.,2019).MIAs are natural products notably involved in plant adaptation to the environment and defense against aggressors.This mainly results from their high biological activities,which also explain their pharmacological properties.展开更多
基金funded by grant no.031B0548 in the frame of the program"Crop plants of the future"from the Bundesministerium fur Bildung und Forschung to A.T.funded by the Investissement d’Avenir program of the French National Agency of Research for the project GENIUS(ANR-11-BTBR-0001_GENIUS).
文摘In plants and mammals,non-homologous end-joining is the dominant pathway to repair DNA doublestrand breaks,making it challenging to generate knock-in events.In this study,we identified two groups of exonucleases from the herpes virus and the bacteriophage T7 families that conferred an up to 38-fold increase in homology-directed repair frequencies when fused to Cas9/Cas12a in a tobacco mosaic virus-based transient assay in Nicotiana benthamiana.We achieved precise and scar-free insertion of several kilobases of DNA both in transient and stable transformation systems.In Arabidopsis thaliana,fusion of Cas9 to a herpes virus family exonuclease led to 10-fold higher frequencies of knock-ins in the first generation of transformants.In addition,we demonstrated stable and heritable knock-ins in wheat in 1%of the primary transformants.Taken together,our results open perspectives for the routine production of heritable knock-in and gene replacement events in plants.
基金supported by the EU Horizon 2020 research and innovation program (MIAMi project-grant agreement N°814645)the ARD CVL Biopharmaceutical program of the Region Centre-Val de Loire (ETOPOCentre project)the ANR (project MIACYC–ANR-20-CE43-0010).
文摘Dear Editor,Rauvolfia tetraphylla(aka the Devil pepper)(Supplemental Figure 1)is a well-known medicinal plant that produces monoterpenoid indole alkaloids(MIAs).This MIA biosynthesis occurs in several organs,including leaves,stems,fruit,and roots,which accumulate the famous antiarrhythmic ajmaline(Kumar et al.2016a,2016b;Kumara et al.,2019).MIAs are natural products notably involved in plant adaptation to the environment and defense against aggressors.This mainly results from their high biological activities,which also explain their pharmacological properties.
