应用发根农杆菌RiT-DNA建立菘蓝的毛状根优质株系和植株再生系统

Establishing better lines and regenerating system of hairy roots of Isatis indigotica with Ri T-DNA

目的 :建立菘蓝 ( Isatis indigotica Fort)的毛状根离体培养系统 ,并诱导毛状根再生植株。 方法 :分别用发根农杆菌A4,R16 0 1和 ATCC15 834三种菌株感染菘蓝的子叶外植体 ,获得毛状根 ,并筛选了优质株系 ;测定了毛状根的生长曲线 ;在含不同激素的培养基上诱导毛状根再生植株 ;利用高压纸电泳法对毛状根和再生植株进行 T- DNA转化的检测。 结果 :首次利用发根农杆菌 A4,R16 0 1和 ATCC15 834三种菌株成功地从菘蓝中诱导出毛状根 ;菘蓝毛状根在含 BA的 MS培养基中成功地诱导出再生植株 ;经高压纸电泳检测 ,菘蓝毛状根及其再生植株中均含有甘露碱 ,表明 Ri质粒的 T- DNA已整合进毛状根和再生植株中。 结论 :菘蓝毛状根离体培养和植株再生系统的建立 ,为进一步进行药用活性成分的工业化生产和引入外源基因改良性状奠定了基础。

Objective: To establish the culture system of hairy root of Isatis indigotica and to induce the regeneration plant of hairy root. Methods: Hairy root of Isatis indigotica was obtained from infected cotyledon explants after infection with Agrobacterium rhizogenes strains A4, R1601 and ATCC15834. The better lines were selected. The growth curve was surveyed and extrinsic factors affecting the growth of hairy roots were investigated. The regeneration plant was induced on MS media with different hormones. The transformation of Ri T DNA was examined through high voltage paper electrophoresis. Results: The hairy root was originally obtained from Isatis indigotica . The regeneration plant was induced on MS media with BA. The result of high voltage paper electrophoresis confirmed the transformation of T DNA from Ri plasmid to the hairy root and regeneration plant. Conclusion: The acquisition of hairy root of Isatis indigotica and regeneration plant of it lay a foundation for the production of active component and introduction of foreign gene. [