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本研究以生长在3个不同施氮水平(60 kg/hm~2,90 kg/hm~2和120 kg/hm~2)下的“云烟87”为材料,运用超高效液相色谱法、分光光度法和荧光定量PCR法,分别研究了不同施氮水平对烟叶生长发育过程中黄酮类化合物芸香苷、山奈酚-3-O-芸香苷和总黄酮含量、代谢相关酶活性及相关基因表达的影响。结果表明:随着生长发育的进行,黄酮类化合物代谢相关基因PAL、C4H、4CL、CHS、CHI、F3H、F3’H、FLS、A3RT的基因表达总体上呈逐渐降低的趋势,从而调控了山奈酚-3-O-芸香苷和总黄酮含量呈逐渐降低的趋势,芸香苷含量呈先略有升高后降低的趋势。不同施氮水平下比较,处理前期(50 d和83 d),较低施氮水平(60 kg/hm~2)上调了烟草叶片黄酮类化合物代谢相关基因PAL、C4H、4CL、CHS、CHI、F3H、F3’H、FLS、A3RT和ANS的基因表达,提高了黄酮类化合物代谢关键酶PAL、CHI的活性,促进了黄酮类化合物芸香苷、山奈酚-3-O-芸香苷和总黄酮含量的积累(p<0.05);处理后期(106 d),较高施氮水平(120 kg/hm~2)上调了烟草叶片黄酮类化合物代谢相关基因PAL、C4H、4CL、CHS、CHI、F3H、F3’H、A3RT和ANS基因的表达,提高了黄酮类化合物代谢关键酶PAL、CHI的活性,促进了黄酮类化合物芸香苷、山奈酚-3-O-芸香苷和总黄酮含量的积累(p<0.05)。
In this study, “Yunyan 87” grown at three different nitrogen levels (60 kg / hm ~ 2,90 kg / hm ~ 2 and 120 kg / hm ~ 2) was used as material and analyzed by ultra performance liquid chromatography The contents of flavonoids rutin, kaempferol-3-O-rutin and total flavonoids, the related enzyme activities and their correlation in the growth and development of tobacco leaves were studied under different nitrogen application rates, spectrophotometry and fluorescence quantitative PCR. Effects of gene expression. The results showed that the gene expression of flavonoid metabolism related genes PAL, C4H, 4CL, CHS, CHI, F3H, F3’H, FLS and A3RT tended to decrease as the growth and development progressed, Phenol-3-O-rutin and total flavonoids showed a trend of decreasing gradually, while rutin showed a tendency of slightly increasing and then decreasing. At the early stage of treatment (50 d and 83 d), lower nitrogen application level (60 kg / hm ~ 2) up regulated the PAL, C4H, 4CL, CHS and CHI of flavonoid metabolism in tobacco leaves, F3H, F3H, FLS, A3RT and ANS increased the activity of flavonoid key enzymes PAL and CHI, and promoted the flavonoid rutin, kaempferol-3-O-rutin and total flavonoids (P <0.05). At the late stage of treatment (106 d), higher nitrogen application level (120 kg / hm ~ 2) upregulated the metabolism of flavonoid related genes PAL, C4H, 4CL, CHS, F3’H, A3RT and ANS genes increased the activity of flavonoid key enzymes PAL and CHI and promoted the accumulation of flavonoid rutin, kaempferol-3-O-rutin and total flavonoids (p <0.05).