摘要:Horvath and Riezman,Yeast,1994; Gottschling Lab[阅读全文:]
摘要:Screen of GST-Fusion Protein Expression(pGEX system by Amersham: for check clones for expression of the desired fusion protein prior to large-scale purification)Pick several colonies of E.coli transfo[阅读全文:]
摘要:Small scale fusion protein preparationGrow 5ml culture o/n in TB with amp.Add o/n culture to 50ml of TB amp and grow for 3 hours in 37℃ shaker.Induce culture by adding 20μl of 1M IPTG (final [0.4mM[阅读全文:]
摘要:Making GST fusion proteins:(07/19/03)ver.1Grow up 5ml LB with Amp o/n.Add to 45ml LB with Amp37o shake 2.5 - 3 hrs,till OD600 0.4-0.8Put bottles in room temperature water for 10 min to cool down.Add 1[阅读全文:]
摘要:GST Protein Prep.Ver.21)Grow 50ml of culture in LB or TB antibiotic o/n at 37℃ shaker.2)Dilute culture in LB or TB antibiotic 1:103)Grow 3hrs at 37℃.4)Induce culture by adding 0.4 mM IPTG final concen[阅读全文:]
摘要:1.Start a 5 ml overnight culture from a frozen stock (in the -70℃)in the following media:For 100 ml of M9CA media tyrp 10 ml of 10X M9 salts5 ml of 10% casamino acids0.2 ml 1M MgSO40.01 ml 1M CaCl20.5[阅读全文:]
摘要:Low (“Low”)Imidazole Buffer 0.5L100mM Imidizole 3.4g5% glycerol 25ml 100% glycerol50mM Tris-HCl (pH 7.9)50ml 0.5M Tris-HCl (pH 7.9)0.1% Tween-20 0.5M 100% Tween-20500mM NaCl50ml 5M NaCldH2[阅读全文:]
摘要:Inoculate 2ml of 5ml o/n culture of either p3133 (empty vector pET11a)or p3134 (dnEBNA-1/Soft)in E.coli BL21 LysS per 0.5L LB ampicillin (grow two 0.5L cultures of each)Incubate ~2hrs 37℃250rpm until [阅读全文:]