Huang L, Wei Z M
Shanghai Institute of Plant Physiology, The Chinese Academy of Sciences, Shanghai 200032.
Shi Yan Sheng Wu Xue Bao. 1999 Dec;32(4):381-9.
Several maize inbreds were transformed with Agrobacterium tumefaciens EHA101 (pGIH). Transgenic maize plants were obtained. Frequency of transformation of maize inbred Suyu No. 1 can reach 8.1%. Results of PCR and Southern blot analysis proved that T-DNA was stably integrated into the genome of maize. Staining with X-gluc confirmed the expression of GUS gene in maize cells. The band amplified by inverse PCR showed that the copy number of transgene in three transformants was single. After long term of subculture, some hygromycin resistant calli lost their regeneration ability. Although Southern blot probed the integration of gusA gene in their genome, GUS activity cannot be detected in those calli. Southern blot analysis of HpaII digest DNA showed that transgenic gusA gene was highly methylated.
用根癌农杆菌EHA101（pGIH）转化了几个玉米自交系。获得了转基因玉米植株。玉米自交系苏玉1号的转化频率可达8.1%。PCR和Southern杂交分析结果证明T-DNA已稳定整合到玉米基因组中。用X-葡糖苷酸染色证实了GUS基因在玉米细胞中的表达。反向PCR扩增的条带表明三个转化体中转基因的拷贝数为单拷贝。经过长期继代培养后，一些潮霉素抗性愈伤组织失去了再生能力。虽然Southern杂交检测到gusA基因已整合到它们的基因组中，但在这些愈伤组织中检测不到GUS活性。对经HpaII消化的DNA进行Southern杂交分析表明，转基因gusA基因高度甲基化。
