【摘要】 本研究选用小黑麦小孢子作为供试材料,进行细胞穿透肽介导的植物单倍体细胞遗传转化可行性研究。提取单核靠边期的小黑麦小孢子进行悬浮培养,将细胞穿透肽与报告基因GFP的表达框复合物对小孢子进行转化。转化植株进行了PCR分子检测,并在荧光体视显微镜下观察外源GFP基因的表达情况。共获得21株候选转基因植株,其中染色体自然加倍植株为10株。T1代加倍植株的PCR检测结果显示,加倍的转基因株系中有3个株系的PCR检测结果呈阳性。通过荧光体视显微镜可在转基因植株的幼胚部位观测到GFP蛋白的荧光。以上结果表明外源GFP基因已经整合到小黑麦基因组上并且得到表达并稳定遗传。因此,由细胞穿透肽介导的植物单细胞转化是一种有效、可行的植物单细胞转基因方法。更多还原

【Abstract】 In this study, the microspores from Triticale c.v. AC Ultima were used as explant to study thefeasibility of cell penetrating peptides(CPPs) mediated transformation to a plant single cell. The microspores atmid-to-late-uninucleate stage were isolated and cultured in liquid suspension midium. The mexture of cellpenetrating peptides(CPPs) and the plasmid DNA only with GFP gene’s expression frame were introducedinto microspores. The transformants were screened by PCR and the expression of foreign GFP gene wasidentfied with fluorescence stereoscope. 21 transformed plants were regenerated, and 10 transformants werehomozygous doubled haploid(DH) plants among them. The analysis of PCR confirmed 3 DH lines with positiveresults in T1 progeny, and GFP fluorescence could also be observed in the immature embryo of these lines. Allthe results indicated that exogenous GFP gene was well integrated into the genome of Triticale and inherited tooffspring. This paper provides an effctive and feasible approch of CPPs mediated transformation, which will beapplied in genetic transformation of plant single cell.更多还原