Optimization of Agrobacterium rhizogenes-Mediated Genetic Transformation System of Morus alba L.

Wu Min; Liao Yangzhen; Pi Jilan; Liu Shangyu; Wei Yuan; Ouyang Zhen

doi:10.14083/j.issn.1001-4942.2025.03.004

Abstract：On the basis of the existing genetic transformation system,the genetic transformation system was optimized in this study to improve the transformation efficiency and lay a foundation for the subsequent re-search on gene function and molecular breeding of mulberry(Morus alba L.).Firstly,the Agrobacterium-me-diated mulberry genetic transformation system was optimized from five aspects:Agrobacterium rhizogenes spe-cies,explants types,pre-culture time,infection time and co-culture time;then the MaCYP90A-pBI121 plas-mid was constructed and transported into Agrobacterium rhizogenes to induce hairy roots.The results showed that the optimized genetic transformation conditions were as follows:using Agrobacterium LBA9402 with OD600=0.6 to infect the true leaves of sterile mulberry seedlings pre-cultured for 2 d for 10 min,and coculturing for 2 d.In the optimized conditions,the induction rate of hairy roots reached 84.51％.The recombinant plasmid MaCYP90A-pBI121 was transformed into mulberry leaves,and the positive transgenic efficiency was 81.84％after hydomycin resistance screening and PCR identification.qRT-PCR results showed that the expression level of MaCYP90A gene in the positive transgenic hairy roots was 39.97 times higher than that of the blank control.Based on the established mulberry genetic transformation system,this study successfully optimized the system,and the conversion efficiency was 1.8 times higher than that(30.21％)of the system before optimization,which could provide technical supports for the follow-up study of mulberry gene function,analysis of secondary metabolite synthesis pathway and molecular breeding.