冰草种间杂种蒙农杂种组织培养再生和遗传转化体系的建立

Plant Regeneration and Genetic Transformation in Wheatgrass(Agropyron cristatum×A.desertorum cv.‘Mengnong')

以冰草属(AgropyronGaertn.)中的1个优质种间杂种——“蒙农杂种”冰草(A.cristatumA.desertorumcv.‘Mengnong’)为材料,以幼穗为外植体,建立了组织培养再生与遗传转化体系。试验中所用诱导愈伤组织的培养基为改良MS+2,4-D2.0～3.0mg·L-1,愈伤组织诱导率平均为83.4%;分化培养基为MS(无附加成分),分化率达59.6%;在1/2MS培养基上生根后得到完整小植株。在此基础上,以抗除草剂基因bar为目标基因,用基因枪法转化幼穗诱导的愈伤组织,并对再生植株进行PCR和Southern鉴定,以及在含除草剂(0.5mg·L-1的Glufosinate)培养基中可正常生长的结果表明,外源基因已整合到受体基因组DNA中,转化率为1.1%。

Plant regeneration from immature inflorescence through direct callus induction was investigated in the caseof a hybrid wheatgrass (A. cristatum × A. desertorum cv.‘Mengnong’). Explants were directly cultured on advancedMS solid medium containning 2.0-3.0 mgL-1 of 2,4-D to induce calli and then transferred to hormone-free MS medium forcallus differentiation and 1/2 MS medium for rooting. It was found that the callus induction frequency was 83.4% and plantregeneration frequency was 59.6%. Phosphinothricin acetyltransferase (bar) gene was transformed into hybrid wheatgrassby particle bombardment. Upon selection with 0.5mgL-1 glufosinate, resistant callus were obtained and some transformedplants recovered in vitro. The transgenic wheatgrasses were identified by PCR and Southern analysis of digested genomicDNA, and they could develop normally in the medium containing glufosinate while the untransgenic plants could not. Thisis the first report on genetic transformation in wheatgrass.