Construction and optimization of a genetic transformation system for efficient expression of human insulin-GFP fusion gene in flax  

作　　者：Wei Zhao Rui Zhang Luyang Zhou Zhongxia Zhang Fei Du Ruoyu Wu Jing Kong Shengjun An 

机构地区：[1]School of Medicine,Hebei University of Engineering,Handan Economic and Technological Development Zone,No.19 Taiji Road,Handan,Hebei Province 056038,China [2]Hebei Provincial Engineering Laboratory of Plant Bioreactor Preparation Technology,Hebei University of Chinese Medicine,No.326 Xinshi South Road,Qiaoxi District,Shijiazhuang,Hebei 050090,China [3]The Second Hospital of Hebei Medical University,No.215 Heping West Road,Changan District,Shijiazhuang,Hebei 050000,China [4]Shijiazhuang Medical College,No.1 Tongxin Road,Lingshou County,Shijiazhuang,Hebei 050500,China [5]Department of Ultrasound Medicine,Hengshui People’s Hospital,Hengshui,Hebei 053000,China

出　　处：《Bioresources and Bioprocessing》2024年第1期1102-1114,共13页生物资源与生物加工(英文)

基　　金：funded by Hebei Development and Reform Commission of China,Hebei Engineering Laboratory,grant number(2021)1157 and the Innovation Fund Project of Hebei University of Engineering(114/SJ2401002149).

摘　　要：The human insulin gene modified with a C-peptide was synthesized according to the plant-preferred codon,and a fusion gene expression vector of insulin combined with green fluorescent protein(GFP)was constructed.The optimization of the flax callus culturing was undertaken,and a more efficient Agrobacterium-mediated genetic transformation of the flax hypocotyls was achieved.The critical concentration values of hygromycin on the flax hypocotyl development,as well as on its differentiated callus,were explored by the method of antibiotic gradient addition,and the application of antibiotic screening for the verification of positive calluses was assessed.The fusion gene of insulin and GFP was successfully inserted into the flax genome and expressed,as confirmed through polymerase chain reaction and Western blotting.In conclusion,we have established a flax callus culture system suitable for insulin expression.By optimizing the conditions of the flax callus induction,transformation,screening,and verification of a transgenic callus,we have provided an effective way to obtain insulin.Moreover,the herein-employed flax callus culture system could provide a feasible,cheap,and environmentally friendly platform for producing bioactive proteins.

关 键 词：Flax callus Insulin-GFP Genetic transformation Agrobacterium-induced infection PROTOPLASTS 

分 类 号：R73[医药卫生—肿瘤]