哺乳动物细胞的融解,9生产厂家说明书

中文名称:潮霉素B/湿霉素乙/效高素英文名称:Hygromycin B;O-6-Amino-6-deoxy-L-glycero-D-galacto-heptopyranosylidene--o-beta-D-talopyranosyl--2-deoxy-N3-methyl-D-streptamineCAS号:31282-04-9C20H37N3O13=527.52级别:Ultra pure grade含量:≥80%浓度:50mg/mlWorking concentration: 50~1000µg/ml in cell culture. A commonly used concentration for the selection of mammalian cells is 200 µg/ml. However, the optimal concentration must be experimentally determined, since it may vary depending on the type of cell usedLD50:6 mg/kg in mouse ;13 mg/kg in guinea pig ;63 mg/kg in rat 性状:液体,50 mg/ml, in PBS (phosphate-buffered saline), sterile-filtered用途:生化研究。是一种氨基糖苷类抗生素,通过阻断多肽的合成,抑制多肽延长,用于原核和真核细胞的筛选。Hygromycin B is an aminoglycosidic antibiotic that inhibits protein synthesis in prokaryotes and eukaryotes. It is used to selectand maintain eukaryotic cells that are stably transfected with the E. coli hygromycin-resistance gene .保存:2~8℃

凤凰时时彩平台,核心提示:This extraction procedure was developed by the Simm Laboratory and described in Rudolph et al., 1999. It’s a great way to make very concentrated extracts for Western blotting, andis particularly useful when studying low-abundance proteins . The protocol is described forharvesting mammalian cells from 6 or 10 cm tissue-culture dishes.

哺乳动物细胞的融解,9生产厂家说明书。A. CELL HARVEST.1. Remove media from cells. Wash in ice-cold PBS.2. Scrape cells in 1 ml of ice-cold PBS. Transfer to 1.5 ml microfuge tube.3. Pellet cells at top speed in microfuge .4. Resuspend cell pellet in FT LYSIS Buffer . Resuspend by pipeting up anddown.5. Drop tube in liquid nitrogen to freeze. Thaw sample on ice . Vortex briefly.6. Repeat freeze-thaw cycle two more times.7. Thaw cell lysate. Add 250 U of Benzonase to digest DNA.Mix. Incubate at room temperature for 10 minutes.8. Measure protein concentration.9. Add an equal volume of 2xLaemmli Buffer prior to analysis by SDSPAGE.Heat at 95oC for 10 minutes. Load immediately on gel. .

B. SOLUTIONS.FT LYSIS BUFFER For 100 ml600 mM KCl 20 ml 3 M KCl20 mM Tris-Cl 2 ml 1 M Tris-Cl 20% Glycerol 20 ml glycerolAdd fresh each time:0.4 mg/ml Pefabloc 1:100 of 40 mg/ml stock10 μg/ml Leupeptin 1:1000 of 10 mg/ml stock10 μg/ml Pepstatin 1:500 of 2 mg/ml stock5 μg/ml Aprotinin 1:2000 of 10 mg/ml stock

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