An Efficient Genetic Transformation Method for Broomcorn Millet Mediated by Agrobacterium tumefaciens
Broomcorn millet(Panicum miliaceum L.)is a traditional grain crop in China.Currently,there is still limited research on the genetic transformation methods for P.miliaceum.This study aimed to establish an efficient genetic transformation method for P.miliaceum mediated by Agrobacterium.Mature seeds of the millet variety Jishu 5 were used as the explants to induce embryogenic callus of millet on CIM and MSD induction media,respectively.They were used as the transformation receptor materials and infected with Agrobacterium containing plant expression vectors for 60 minutes and co-cultured for 6 days.The calli were then transferred to a screening medium containing 0.025 g/L of hygromycin to screen for resistant calli.Then,they differentiated on a differentiation medium containing 0.015 g/L of hygromycin.Finally,they rooted into seedlings on a rooting medium containing 0.015 g/L of hygromycin.PCR detec-tion was performed on the regenerated plants using vector specific primers to identify whether they were positive transgenic plants.Based on the multiple batches of transformation experiments,the screening efficiency and transfor-mation efficiency of resistant regenerated plants of broomcorn millet were statistically analyzed.The results showed that both CIM and MSD media could induce embryogenic callus of broomcorn millet,but CIM medium had a better effect on inducing embryogenic callus of broomcorn millet.After being infected by Agrobacterium and co-cultured with Agrobacterium,the multiple resistant regenerated plants were obtained through screening,differentiation,and rooting on a medium containing hygromycin.The PCR identification positive rate of the resistant regenerated plants obtained from three experiments was 100%,and the average transformation efficiency was over 30%.This study successfully estab-lished an Agrobacterium mediated genetic transformation method for broomcorn millet,which is simple to operate,ef-ficient,cost-effective,and not limited by seasons.It could be carried out on a large scale,providing an effective techni-cal means for genetic improvement of broomcorn millet.
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