摘要
为了验证二氢黄酮醇-4-还原酶基因(DFR)在杨树上的功能,明确DFR对抗病物质儿茶素合成的影响,利用抗病基因防治树木溃疡病提供候选基因。以接种欧美杨细菌性溃疡病菌后6 d的一年生中林46杨树苗干的树皮为材料,利用RT-PCR技术克隆DFR基因的ORF序列,并构建DFR的反义表达载体anti-p BI121-DFR。采用农杆菌介导叶盘法转化84K杨,获得了转反义DFR基因的84K杨4株。用高效液相色谱法检测转基因植株叶片中的儿茶素质量分数,结果显示,4株转反义DFR株其内源儿茶素质量分数分别为0.97、2.4、1.6和0.87 ng/g,与84K野生型植株中儿茶素质量分数(8.9 ng/g)相比显著降低。上述结果表明DFR基因参与了杨树类黄酮生物合成途径,该基因与儿茶素的合成有关。
The experiment was conducted to verify the function of Dihydro flavonol-4-reductase gene（ DFR）,and analyze the influence on the synthesis of disease-resistant substances catechins for the use of resistance genes resistant canker providing candidate genes. Total RNA was isolated from the infected bark of Populus×euramericana‘Zhonglin 46＇after inoculating 6days with Lonsdalea quercina subsp. populi. A specific DFR fragment of the expected size was amplified by RT-PCR and sequenced,and then successfully constructed into an expression vector with antisense-orientation driven by CaM V 35 S promoter（ anti-pB I121-DFR）. Subsequently,anti-pB I121-DFR was transformed into Populus alba × P. glandulosa using Agrobacterium tumefaciens mediated method. Four transgenic lines was finally obtained and confirmed by PCR. High performance liquid chromatography（ HPLC） was further used to detect catechin in transgenic plants and wild type plants. The contents of catechin in transgenic plants were 0.97,2.4,1.6,and 0.87 ng / g,respectively,significantly lower than that of wild type plants（ 8.9 ng / g）. Therefore,DFR of Populus×euramericana ‘Zhonglin 46＇was involved in flavonoids biosynthesis pathway,and associated with the synthesis of catechin.
出处
《东北林业大学学报》
CAS
CSCD
北大核心
2016年第10期49-55,共7页
Journal of Northeast Forestry University
基金
国家自然科学基金项目(31470668、31200511、J1103516)
国家林业公益性行业科研专项(201104054)
