Agrobacterium rhizogenes-mediated marker-free transformation and gene editing system revealed that AeCBL3 mediates the formation of calcium oxalate crystal in kiwifruit  

作　　者：Pengwei Li Yiling Zhang Jing Liang Xufan Hu Yan He Tonghao Miao Zhiyin Ouyang Zuchi Yang Abdul Karim Amin Chengcheng Ling Yize Liu Xiuhong Zhou Xiaoran Lv Runze Wang Yajing Liu Heqiang Huo Yongsheng Liu Wei Tang Songhu Wang 

机构地区：[1]Anhui Provincial Key Laboratory of Horticultural Crop Quality Biology,School of Horticulture,Anhui Agricultural University,Hefei 230036,China [2]Mid-Florida Research and Education Center,University of Florida,Institute of Food and Agricultural Sciences,Apopka,FL 32703,USA

出　　处：《Molecular Horticulture》2024年第1期574-588,共15页分子园艺(英文)

基　　金：supported by the National Natural Science Foundation of China(U23A20204),Anhui Provincial Key Laboratory of Horticultural Crop Quality Biology,Startup Funding from Anhui Agricultural University,and the“Wanjiang Scholar”Program(Anhui Province).

摘　　要：The transformation and gene editing of the woody species kiwifruit are difficult and time-consuming.The fast and marker-free genetic modification system for kiwifruit has not been developed yet.Here,we establish a rapid and efficient marker-free transformation and gene editing system mediated by Agrobacterium rhizogenes for kiwifruit.Moreover,a removing-root-tip method was developed to significantly increase the regeneration efficiency of transgenic hairy roots.Through A.rhizogenes-mediated CRISPR/Cas9 gene editing,the editing efficiencies of CEN4 and AeCBL3 achieved 55 and 50%,respectively.And several homozygous knockout lines for both genes were obtained.Our method has been successfully applied in the transformation of two different species of kiwifruit(Acti-nidia chinensis‘Hongyang’and A.eriantha‘White’).Next,we used the method to study the formation of calcium oxalate(CaOx)crystals in kiwifruit.To date,little is known about how CaOx crystal is formed in plants.Our results indicated that AeCBL3 overexpression enhanced CaOx crystal formation,but its knockout via CRISPR/Cas9 significantly impaired crystal formation in kiwifruit.Together,we developed a fast maker-free transformation and highly efficient CRISPRCas9 gene editing system for kiwifruit.Moreover,our work revealed a novel gene mediating CaOx crystal formation and provided a clue to elaborate the underlying mechanisms.

关 键 词：Marker-free transformation Agrobacterium rhizogenes CRISPR/Cas9 KIWIFRUIT AeCBL3 

分 类 号：S66[农业科学—果树学]