延缓叶片衰老ZmIPT2基因的玉米遗传转化及功能验证

Transformation of Zm IPT2 Gene Related to Leaf Senescence into Maize and Function Identification

从玉米自交系郑58中克隆Zm IPT2基因并构建单子叶植物表达载体,通过农杆菌侵染萌动胚方法将其转入玉米自交系K10中,对转基因后代进行分子检测和功能验证。结果表明,Zm IPT2基因c DNA全长969 bp,成功构建其单子叶植物表达载体p CAMBIA5300-ubi-Zm IPT2。农杆菌侵染萌动胚法共转化K10种子5 163粒,获得T0代PCR阳性幼苗48株,其中13株结实收获种子;获得的3个T2代株系PCR阳性率符合3∶1的分离比,且RT-PCR检测呈阳性;2个T2代转基因株系的成熟期叶绿素含量和细胞分裂素含量极显著高于对照,相对绿叶面积和叶面积持绿期极显著或显著高于对照,百粒重和小区产量显著高于对照。结果初步证明,Zm IPT2基因在玉米中的过表达可延缓叶片衰老,提高玉米产量。

ZmlPT2 gene was cloned from maize inbred line Zheng58 and then was constructed into the expression vector of monocotyledon. The gene was introduced into maize inbred lines K 10, and the molecular test and functional verification were employed to analyze transgenie progeny. The results showed that ZmlPT2 gene with 969bp length was cloned and introduced into monocot expression vector pCAMBIA5300-ubi-ZmlPT2. By Agrobacteriummediated transformation method, the gene was transformed into 5 163 germinating embryo of maize inbred lines K10, got 48 PCR positive seedlings in To generation, and 13 plants of them were harvested. The segregation ratio of PCR was 3：1 in 3 T2lines and they were positive in RT-PCR test totally. CTK and chlorophyll content of two transgenie lines were significantly higher than that of control line at mature stage. The relative green leaf area（RGLA）, leaf area to green time（LAD）, 100-seed weight and plot yield of transgenic lines were significantly higher or very significantly higher than that of control. The results proved that the over expression of ZmlPT2 gene in maize could delay leaf-senescence and increased yield.