文档介绍：-
. z.
*t大人物的悲哀在于他们需要不停地做出选择；而小人物的悲哀在于他们从来没有选择的时机。男人因沧桑而成熟，女人因成熟而沧桑。男人有了烟，有了酒，也就有tube.
7. If necessary, dialize the supernatant against PBS with
50mM/L Tris-HCl pH .
五、
植物材料：水稻苗，叶鞘，根〔ynibcas〕
1、200 毫克样品置于冰上磨碎
2、加lysis buffer，离心，10000rpm，4度，5min 取上清
3、重复离心5min
lysis buffer：urea np-40 ampholine 2-me pvp-40
六、
蛋白质样品制备〔sigma〕
秧苗蛋白质样品的提取按Davermal 等〔1986〕的方法进展。
100mg材料剪碎后参加10mgPVP-40(聚乙烯吡咯烷***)及少量石英砂，用液氮研磨成粉， ml 10% 三
***乙酸〔***配制，含10mM %β-巯基乙醇〕，混匀，-20℃沉淀1 小时，4℃，15000 r/min离心15
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. z.
min，弃上清，***(含10 mMβ-巯基乙醇)，再于-20℃沉淀1 小时，同上离心弃上清，
〔有必要再用80％***(含10 mMβ-巯基乙醇所得沉淀低温冷冻真空抽干。
按每mg干粉参加20μl〔可调〕 UKS液[ M尿素，5mM 碳酸钾，%SDS，%DTT(二硫苏糖醇)，
2% Ampholine (Amersham Pharmacia Biotech Inc，-10)，6% Triton *-100]，37℃温育30min，期间搅
动几次，28度 〔温度低，高浓度的尿素会让溶液结冰〕16000 r/min离心15 min，离心力越大时间长一点
越好！上清即可上样电泳。或者-70 度保存
七、
植物根中蛋白质的抽取〔phenol〕
(1) sample, 液氮研磨
(2) ml centrifuge 用tube
(3) 加 1M KH2PO4+K2HPO4 700 ul
(4) 12000 rpm, 4度, 10-15minite
(5) 取上层液，蛋白质就在里面
八、
SDS e*traction followed by acetone precipitation – simple e*traction protocol that does not require phenol.
Remended start protocol for whole tissue e*tractions.〔hgp〕
1. Grind 1 g of fresh tissue to a powder with liquid nitrogen in a mortar and pestle.
2. Add 5 mL of e*traction media ( M Tris-HCl, pH , 5% SDS, 15% glycerol, M DTT) directly to
mortar and continue grinding for an additional 30 sec.
3. Filter homogenate through two layers of miracloth into a 50 mL Falcon tube at room temperature.
4. Immediately add 4 volumes of ice cold 100% acetone to filtered homogenate, mi* by vorte*ing and place at -20
C for at least one hour to precipitate proteins.
5. Centrifuge at 5000 g for 15 min to collect precipitated protein, decant supernatant.
6. Gently blot residual acetone from container with Kimwipe and then wash pellet in 15-20 mL of cold 80%
acetone. Be sure to thoroughly break-up pellet by pipetting, vorte