细胞穿透肽介导的小黑麦小孢子遗传转化研究  

Cell Penetrating Peptides Mediated Transformation of Triticale Microspore

作　　者：刘大普[1,2] 杨凤萍[1] 张晓东[1] Liu Dapu Yang Fengping Zhang Xiaodong(Beijing Agricultural Biotechnology Research Center, Beijing Academy of Agriculture and Forestry Science, Beijing 100097 Plant Science and Technology College, Beijing University of Agriculture, Beijing 102206)

机构地区：[1]北京市农林科学院北京农业生物技术研究中心,北京100097 [2]北京农学院植物科学技术学院,北京102206

出　　处：《中国农学通报》2017年第20期29-34,共6页Chinese Agricultural Science Bulletin

基　　金：北京市优秀人才培养资助项目"麦类作物抗逆转录因子基因的挖掘和利用"(PYZZ130826001696);北京市农林科学院科技创新能力建设专项"小麦小孢子转化系统的建立"(KJCX201102003);北京市留学人员科技活动择优资助项目资助"利用大麦条纹花叶病毒研究小黑麦茎秆NAC转录因子基因"

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。In this study, the microspores from Triticale c.v. AC Uhima were used as explant to study the feasibility of cell penetrating peptides （CPPs） mediated transformation to a plant single cell. The microspores at mid-to-late-uninucleate stage were isolated and cultured in liquid suspension midium. The mexture of cell penetrating peptides （CPPs） and the plasmid DNA only with GFP gene＇ s expression frame were introduced into microspores. The transformants were screened by PCR and the expression of foreign GFP gene was identfied with fluorescence stereoscope. 21 transformed plants were regenerated, and 10 transformants were homozygous doubled haploid （DH） plants among them. The analysis of PCR confirmed 3 DH lines with positive results in T~ progeny, and GFP fluorescence could also be observed in the immature embryo of these lines. All the results indicated that exogenous GFP gene was well integrated into the genome of Triticale and inherited to offspring. This paper provides an effctive and feasible approch of CPPs mediated transformation, which will be applied in genetic transformation of plant single cell.

关 键 词：小黑麦 小孢子 细胞穿透肽 转化 

分 类 号：Q789[生物学—分子生物学]