颠茄PMT·TR-I和H6H基因植物高效表达载体的构建

Construction of Plant Expression Vectors of PMT,TR-I and H6H

[目的]构建1,4-丁二胺-氮-甲基转移酶(Putrescine N-methyltransferase,PMT)基因、托品酮还原酶-I(Tropinone reductase-I,TR-I)基因和莨菪碱6-β-羟化酶(Hyoscyamine 6β-Hydroxylase,H6H)基因的植物高效表达载体。[方法]采用RT-PCR方法从颠茄中克隆到PMT、TR-I和H6H基因编码区序列,并连接到植物表达载体pCAMBIA1304+(p1304+)中。[结果]成功构建了植物高效表达载体p1304+-PMT、p1304+-TR-I、p1304+-H6H、p1304+-PMT-H6H、p1304+-TR-I-PMT和p1304+-TR-I-H6H,并转化农杆菌,获得可直接用于遗传改良颠茄的工程菌。[结论]该研究为利用植物基因工程技术提高东莨菪碱的产量奠定了基础。

[Objective]To constrct the plant expression vectors of putrescine N-methyltransferase（PMT）,tropinone reductase-I（TR-I） and hyoscyamine 6β-hydroxylase （H6H）. [Method]The coding sequences of PMT,TR-I and H6H were obtained by the method of RT-PCR and ligated to the plant expression vector pCAMBIA1304^＋ （p1304^＋ ）. [Result]The expression vectors p1304^＋ -PMT,p1304^＋ -TR-I,p1304^＋ -H6H,p1304^＋ -PMT-H6H,p1304^＋ -TR-I-PMT and p1304^＋ -TR-I-H6H were constructed and transferred to Agrobacterium successfully. Engineered bacteria were obtained that can meliorate Atropa belladonna L.. [Conclusion]It establishes the base of improving production of scopolamine by genetic engi-neering technology.