Al-Forkan Mahammad, Power J Brian, Anthony Paul, Lowe Kenneth C, Davey Michael R
Plant Sciences Division, School of Biosciences, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
Cell Mol Biol Lett. 2004;9(2):287-300.
Morphologically normal, fertile transgenic plants were obtained by co-culturing embryogenic calli of the Bangladeshi Indica rice cultivars BR26 and Binni with Agrobacterium tumefaciens strain LBA4404 carrying the super binary vector pTOK233. Acetosyringone (100 microM) in the medium during co-culture (25-28 degrees C) and selection on hygromycin B (50 mg l(-1)) were essential for efficient transformation. Stable integration and expression of beta-glucuronidase, neomycin phosphotransferase and hygromycin phosphotransferase genes in regenerated plants were confirmed by histochemical and fluorometric assays, ELISA and Southern analysis. Two to 3 copies of T-DNA were integrated into regenerated plants; transgene expression did not correlate with gene copy number. Mendelian segregation of transgenes occurred in T1 seed progeny.
通过将孟加拉国籼稻品种BR26和Binni的胚性愈伤组织与携带超级双元载体pTOK233的根癌农杆菌菌株LBA4404共培养，获得了形态正常、可育的转基因植株。共培养期间（25 - 28摄氏度）培养基中加入乙酰丁香酮（100微摩尔）以及在潮霉素B（50毫克/升）上进行筛选对于高效转化至关重要。通过组织化学和荧光测定、酶联免疫吸附测定以及Southern分析，证实了再生植株中β-葡萄糖醛酸酶、新霉素磷酸转移酶和潮霉素磷酸转移酶基因的稳定整合与表达。两到三个T-DNA拷贝整合到了再生植株中；转基因表达与基因拷贝数无关。转基因在T1种子后代中发生孟德尔分离。
