抗草甘膦基因aroA转化陆地棉胚性愈伤组织的研究

为了建立以草甘膦为选择标记的农杆菌介导的棉花胚性愈伤组织转化体系,本研究将草甘膦抗性基因(aroA)转化陆地棉WC和YZ-1的胚性愈伤组织,分别以5 mg·L-1和10 mg·L-1的草甘膦浓度作为选择压,经过2~3轮筛选,获得了抗性胚性愈伤组织,诱导出胚状体并发育成苗。对T0再生植株进行PCR及草甘膦抗性检测,以WC和YZ-1为受体的转抗草甘膦基因(aroA)分别获得了33株和10株阳性植株,转化效率分别为82.5%和62.5%。该体系为棉花遗传转化提供了实验方法,获得的抗草甘膦棉花为棉花育种提供种质资源;同时抗除草剂基因还可以作为筛选标记与其它基因串联在一起转化棉花,进行基因功能验证。

Study on Transformation of Glyphosate-resistant Gene aroA into Embryogenic Calli of Upland Cotton via Agrobacterium tumefaciens

For producing the glyphosate-resistant cotton plants, we developed an Agrobacterium-mediated transformation system from embryogenic calli(EC) in which glyphosate was used as a selective agent. In this study, glyphosate-resistant gene aroA was transformed to embryogenic calli(EC) of Gossypium hirsutum L. cv. WC and YZ-1. And the calli were transferred onto culture medium, respectively, with 5 and 10 mg·L^-1 glyphosate for selection. After two or three cycles of glyphosate-resistant screening, resistant EC was obtained and transgenic plantlets were regenerated from embryoid and grafted in greenhouse. Glyphosate-resistant positive plants were identified by PCR analysis and glyphosate-resistant screening. In this study, 33 and 10 glyphosate-resistant positive plants were obtained from WC and YZ^-1, respectively, and the corresponding transformation efficiency was 82.5% and 62.5%. This transformation system provides not only an efficient method for cotton genetic transformation but also important germplasm resources for cotton breeding. Meanwhile, the selection marker gene aroA can be transformed into cotton genome together with other target gene for functional analysis.