Genetic transformation of Agave salmiana by Agrobacterium tumefaciens and particle bombardment
Genetic transformation of Agave salmiana by Agrobacterium tumefaciens and particle bombardment
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DOI：
10.1007/s11240-007-9287-3
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发表时间：
2007-12-01
影响因子：
3
通讯作者：
Alpuche-Solis, Angel Gabriel
中科院分区：
文献类型：
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作者：
Flores-Benitez, Silvia;Jimenez-Bremont, Juan F.;Alpuche-Solis, Angel Gabriel
采用农杆菌共培养和基因枪轰击两种方法对龙舌兰进行转化。uidA（β-葡糖醛酸酶）基因用作两种方法的报告基因，而npt II和bar基因用作A. tumefaciens和基因枪转化。以往的报道，在体外再生的A。salmiana的研究尚未公开;因此，使用龙舌兰的叶和胚性愈伤组织优化了芽再生和生根的条件。用聚合酶链反应（PCR）检测了11个月龄的转基因植株。转基因植物的性质也证实了使用GUS组织化学测定。通过与携带pBII 21二元载体的农杆菌共培养外植体的转化是最有效的转化方法，产生32个转基因植株，转化效率为2.7%。另一方面，基因枪方法产生的转基因愈伤组织在选择性培养基上培养14个月后，GUS检测呈阳性，同时仍在再生。
Agave salmiana was transformed using two different protocols: co-cultivation with Agrobacterium tumefaciens and particle bombardment. The uidA (beta-glucuronidase) gene was used as a reporter gene for both methods whereas the nptII and bar genes were used as selectable markers for A. tumefaciens and biolistic transformation respectively. Previous reports for in vitro regeneration of A. salmiana have not been published; therefore the conditions for both shoot regeneration and rooting were optimized using leaves and embryogenic calli of Agave salmiana. The transgenes were detected by Polymerase Chain Reaction (PCR) in 11 month old plants. The transgenic nature of the plants was also confirmed using GUS histochemical assays. Transformation via co-cultivation of explants with Agrobacterium harbouring the pBII21 binary vector was the most effective method of transformation, producing 32 transgenic plants and giving a transformation efficiency of 2.7%. On the other hand, the biolistic method produced transgenic calli that tested positive with the GUS assay after 14 months on selective medium while still undergoing regeneration.