Taniguchi T, Kurita M, Ohmiya Y, Kondo T
Forest Tree Breeding Center, 3809-1 Ishi, Juo, Hitachi, Ibaraki, 319-1301, Japan.
Plant Cell Rep. 2005 Mar;23(12):796-802. doi: 10.1007/s00299-004-0895-7. Epub 2004 Nov 13.
A genetic transformation procedure for Chamaecyparis obtusa was developed after co-cultivation of embryogenic tissues with disarmed Agrobacterium tumefaciens strain C58/pMP90, which harbours the sgfp (synthetic green fluorescent protein) visual reporter and nptII (neomycin phoshotransferase II) selectable marker genes. The highest transformation frequency was 22.5 independent transformed lines per dish (250 mg embryogenic tissue) following selection on kanamycin medium. Transgenic plantlets were regenerated through the maturation and germination of somatic embryos. The intensity of GFP fluorescence, observed under a fluorescence microscope, varied from very faint to relatively strong, depending on the transgenic line or part of the transgenic plant. The integration of the genes into the genome of regenerated plantlets was confirmed by Southern blot analysis.
在用携带sgfp（合成绿色荧光蛋白）视觉报告基因和nptII（新霉素磷酸转移酶II）选择标记基因的无致病力根癌农杆菌菌株C58/pMP90与胚性组织共培养后，开发了一种钝叶扁柏的遗传转化程序。在卡那霉素培养基上筛选后，最高转化频率为每皿（250毫克胚性组织）22.5个独立转化株系。通过体细胞胚的成熟和萌发再生出转基因植株。在荧光显微镜下观察到的GFP荧光强度因转基因株系或转基因植物的部位而异，从非常微弱到相对较强。通过Southern杂交分析证实了基因整合到再生植株的基因组中。
