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wovs211木蟲 (正式寫手)
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[求助]
有關(guān)植物逆境脅迫下ASA-GSH關(guān)鍵代謝酶的測定
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各位蟲友，關(guān)于ASA-GSH代謝關(guān)鍵酶MDHAR的測定方法，我用的是下面的方法，飯是我測的黑麥草根系中這個(gè)MDHAR的OD值沒有變化，不知道是怎么回事，十分著急，望蟲友們提供幫助。 我的參考文獻(xiàn)是Exogenous sodium nitroprusside and glutathione alleviate copper toxicity by reducing copper uptake and oxidative damage in rice (Oryza sativa L.) seedlings，Protoplasma (2014) 251:1373–1386。酶的提取液方法是To extract enzymes, fresh leaf samples (0.5 g) were homogenized separately with a reaction mixture containing 50 mM Kphosphate buffer (pH 7.0), 100 mM KCl, 1 mM AsA, 5 mMβ-mercaptoethanol, and 10 % (w/v) glycerol in pre-chilled mortars and pestles. The homogenate was centrifuged at12,500×g for 15 min and the resultant supernatants werecollected for analysis of enzyme activities and protein content.All procedures were performed at 0–4°C. 我提取的酶液定容到3ml.MDHAR (EC 1.6.5.4) activity was determined according to the method of Hossain et al. (1984). The reaction mixture contained 50 mM Tris–HCl buffer (pH 7.5), 0.2 mM NADPH, 2.5 mM AsA, 1.0 U of AO and enzyme solution in a final volume of 0.7 ml. The activity was calculated from the change in ascorbate at 340 nm for 1 min using the extinction coefficient of 6.2 mM−1 cm−1. 各位蟲友，我測出OD值沒有變化啊，怎么辦啊。望蟲友們提供幫組，謝謝。 |
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