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[交流] 抽提核蛋白为什么要用spermine？

抽提核蛋白为什么要用spermine？

以下核抽提的porocol中，为什么在buffer中要用到Spermidine，Spermine，是必须的吗，具体有什么作用？谢谢各位！

1. The monocytes were pelleted and resuspended in 300 ul of buffer A (50 mM NaCl, 10 mM HEPES pH 8, 500 mM sucrose, 1 mM EDTA, 0.5 mM Spermidine, 0.15 mM Spermine, 0.2% Triton X-100) containing -mercaptoethanol and the protease inhibitors PMSF, leupeptin, aprotinin and pepstatin.

2. After 15 min on ice and centrifugation, the supernatant (cytoplasmic fraction) was collected and stored while the pellet was washed with 200 ul Buffer B (50 mM NaCl, 10 mM HEPES pH 8, 25% glycerol, 0.1 mM EDTA, 0.5 mM Spermidine, 0.15 mM Spermine) and then resuspended in 100ul buffer C (350 mM NaCl, 10 mM HEPES, 25% glycerol, 0.1 mM EDTA, 0.5 mM Spermidine, 0.15 mM Spermine).

3. After centrifugation, the supernatant (nuclear fraction) was recovered. Cytoplasmic and nuclear fractions were quantitated for protein content by micro-BCA method (Pierce) and subjected to Western blot analysis.

[ Last edited by 350784478 on 2009-10-13 at 11:57 ]

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