根癌农杆菌介导的蒙古冰草稳定遗传转化体系建立

Establishment of Agrobacterium-mediated Transformation System for Agropyron mongolicum

蒙古冰草(Agropyron mongolicum)亦称沙芦草,为禾本科(Poaceae)小麦族(Triticeae)冰草属(Agropyron)多年生疏丛型牧草,具有饲用价值高、抗寒耐旱以及耐盐、耐瘠薄、耐风沙等特性,是改良天然草场的适宜草种与挖掘优良耐逆基因资源的重要材料。然而,目前尚未建立蒙古冰草高效遗传转化体系,制约了该物种的基因资源鉴定与遗传改良应用。以蒙农1号蒙古冰草种子为来源的高再生效率的胚性愈伤系#89为外植体,建立了根癌农杆菌(Agrobacterium tumefaciens)EHA105介导的蒙古冰草稳定遗传转化体系,转化效率达30%。此外,针对多次继代后蒙古冰草愈伤系再生能力退化的难题,通过在再生培养基中添加1 mg·L^(-1) ABA或提高蔗糖浓度至45 g·L^(-1),成功将再生能力衰退的蒙古冰草愈伤系再生效率由5%分别提高至35%与42%。研究结果为后续蒙古冰草基因编辑体系建立、基因功能鉴定和新品种培育奠定了重要技术基础。

Agropyron mongolicum is a perennial shrub grass in the Triticeae tribe of Poaceae family.It has high forage quality and tolerance to cold,drought,salt and sandstorm.Therefore,it is an excellent grass species for mining stress tolerant genes and restoring deteriorated grasslands.However,a highly efficient transformation system has yet to be established in A.mongolicum.It restricts the identification of genetic resources and the application of genetic improve-ment of this species.Here,we report an embryogenic callus line(ECL)#89 that was induced from the seeds of A.mon-golicum cultivar Mengnong No.1.Our studies showed the ECL#89 had high regeneration and Agrobacterium-infection efficiencies.An Agrobacterium-mediated transformation system with a transformation efficiency up to 30%was estab-lished by infecting the ECL#89 with EHA105.Additionally,we found that subcultures could decrease severely the rege-neration capacity of ECL#89,which could be partially restored by adding(1 mg·L^(-1))ABA or high concentration(45 g·L^(-1))sucrose in the regeneration medium,from 5%to 35%and 42%,respectively.Our work will facilitate the developing of genome editing system,gene function characterization,and molecular breeding in A.mongolicum.