Gürel F, Gözükirmizi N
University of Istanbul, Department of Biology, Molecular Biology Section, Vezneciler-34459 Istanbul, Turkey e-mail:
TUBITAK MRC, Research Institute of Genetic Engineering and Biotechnology, 41470-Gebze Kocaeli, Turkey, , , , , , TR.
Plant Cell Rep. 2000 Jul;19(8):787-791. doi: 10.1007/s002999900182.
This study was conducted to detect the optimum conditions for DNA transfer into mature embryos of barley via electroporation. Cultured mature embryos of barley were directly electroporated in the presence of the pBI 121 vector carrying both the β-glucuronidase (GUS) and neomycin phosphotransferase II (npt II) genes. It was found that 500 v/cm and 500 μFd capacitance was the optimum combination for healthy germination of the transformed plants from mature electroporated embryos. Effects of culture duration before electroporation and selection antibiotic concentrations on germination were also examined. Gene transfer performed on 3-day-old cultures resulted in the highest germination frequencies. GUS expression was observed on transversal sections of embryos and mature leaves from 3 month-old regenerants. PCR and Southern blot analyses show the presence of the npt II transgene in the genome of a plant.
本研究旨在检测通过电穿孔法将DNA导入大麦成熟胚的最佳条件。将培养的大麦成熟胚在携带β-葡萄糖醛酸酶（GUS）和新霉素磷酸转移酶II（npt II）基因的pBI 121载体存在下直接进行电穿孔。结果发现，500 V/cm的电场强度和500 μFd的电容是使经电穿孔处理的成熟胚发育出的转化植株健康萌发的最佳组合。同时还研究了电穿孔前培养时间和选择抗生素浓度对萌发的影响。对3日龄培养物进行基因转移，得到的萌发频率最高。在3个月大的再生植株的胚横切片和成熟叶片上观察到了GUS表达。PCR和Southern杂交分析表明，植株基因组中存在npt II转基因。
