基因枪介导转hpa99基因小麦的获得和检测

为了探究来源于水稻细条斑病菌(Xanthomonas oryzae pv.oryzicola)的hpa99基因改良小麦抗赤霉病的可行性,以小麦幼胚诱导的愈伤组织为转化受体,以磷酸甘露糖异构酶(phosphomannose isomerase,PMI)作为选择标记,通过基因枪法将带有hpa99基因的表达质粒AF234296导入普通小麦品种山农15。2～3次PMI筛选后,获得38株再生植株。通过氯酚红(chlorophenol red,CPR)染色以及PMI基因和目的基因hpa99的PCR检测,获得7株阳性植株,转化率为0.108%。结果表明,hpa99已整合到小麦基因组中,获得了无抗生素标记基因的转基因小麦。

Acquisition and Determination of Wheat with Transferred hpa99 Gene Mediated by Biolistic Particle

To assess the role of hpa99 gene isolated from Xanthomonas oryzae pv.oryzicola against wheat scab,an efficient acceptor system for gene transformation was established with callus of wheat immature embryos.The AF234296 vector containing hpa99 gene and phosphomannose isomerase(PMI) gene was transferred into Triticum aestivum cv.Shannong 15,a common wheat variety,by particle bombardment-mediated transgenic method.Thirty-eight regenerated plants were obtained on the selection medium complemented with L-mannose.Seven transgenic plants were identified by chlorophenol red staining and PCR assay in the T0 generation with the transformation frequency as 0.108%.These results suggested that hpa99 gene had been successfully transferred into wheat genome,and the transgenic wheat without antibiotic marker genes was obtained.