PuCAT/Bar抗逆抗除草剂基因共转化紫花苜蓿的研究

Co-transformation of Puccinellia chinampoensis PuCAT Gene and Bar Gene into Alfalfa with Agrobacterium-mediated Method

为了提高紫花苜蓿的抗盐碱能力,利用盐生植物碱茅的PuCAT基因成功构建了p3300-35S-PuCAT-NOS植物表达载体,以Bar基因为筛选标记基因,通过农杆菌介导转化公农5号紫花苜蓿子叶,并对其遗传转化体系进行了优化(预培养、菌液浓度、侵染时间、共培养、抗生素浓度),经草铵膦筛选获得72株抗性植株,用PCR检测和RT-PCR鉴定,最终获得30株RT-PCR阳性植株。结果表明,PuCAT基因已经整合在苜蓿基因组中,并且能够在RNA水平上表达。

In order to improve the salt and alkali tolerance of alfalfa,the plant expression vector p3300-35S-PuCAT-NOS was constructed and transformed into alfalfa via Agrobacterium mediated.The selectable marker gene was Bar gene.And the genetic transformation system was optimized,including:pre-incubation time,bacteria concentration,inoculation time,co-cultivation time and antibiotic concentration.The transformed plants were selected with glufosinate ammonium.30 transformed plants were identified as positive in the gene expression by PCR and RT-PCR which suggested that the PuCAT gene had been integrated into the genome of the regenerated plants.