金铁锁毛状根诱导及培养体系的建立

Establishment of induction and culture system for hairy roots of Psammosilene tunicoides

目的:研究金铁锁毛状根制备方法及液体培养条件,为规模化生产金铁锁提供技术支持。方法:利用发根农杆菌ACCC10060菌株侵染金铁锁幼叶及茎段。结果:将外植体与菌液(A6000.8)在含有20 mg.L-1乙酰丁香酮的B5培养基上共培养48 h,可以有效地诱导金铁锁毛状根;叶片比茎段的诱导率高。转化的毛状根具有激素自养和抗卡那霉素的特性,并且组织中含有冠瘿碱,证明该菌对金铁锁组织成功地进行了转化。在无激素的1/2 MS液体培养基中培养35 d后,金铁锁毛状根生物量增加了14.11倍(鲜重)和8.39倍(干重),其总皂苷为0.857%(干重),愈伤组织和原植物中分别为0.388%,0.217%。结论:金铁锁毛状根离体培养系统的成功建立,对进一步规模化培养金铁锁毛状根具有重要意义。

Objective：To establish a culture system for Psammosilene tunicoides hairy roots,and provide technological aid for the large-scale production of P.tunicoides material.Method：The young leaves and stem segments of sterile plantlets were infected with ACCC10060 strain,and subsequently a culture system suitable for hairy roots growth was further established.Result：When explants were co-cultured with ACCC10060（A600 0.8） on B5 media containing 20 mg·L-1 Acetosyringo（AS） for 48 h,the hairy roots could be successfully induced,and it could achieve a higher induction rate using young leaves as explants than that of stem segments.The transfected hairy roots possessed the ability of kanamycin resistance and growth on hormone-free media,and synthesis of opines.All above results demonstrated that the present hairy roots originated in the infection of P.tunicoides tissues by ACCC10060 strains.After 35 d culture in liquid hormone-free MS（1/2 strength）,the biomass of hairy roots increased 14.11 times（fresh weight） and 8.39 times（dry weight）,respectively,and the content of total saponins in hairy roots reached to 0.857%（DW）,by contrast,it＇s only 0.388% and 0.217% in callus and seedlings respectively.Conclusion：Establishment of hairy roots culture of P.tunicoides provided a foundation for industrial production of active components from P.tunicoides culture.