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Available from Riken BioResource Center.

Plasmid & Protein Purification Database -Purification-

Species  
Locus tag ST1969
Products  
Plasmid ID 248
Vector  
Purification Lot ST1969_1a
Purified Date 2004.12.14-12.28
Protein conc. (mg/ml) 9.4
Total volume (ml) 0.7
Total protein (mg) 6.58

Culture Lot:

date host cell scale label weight of bacteira (g)
        30.9

Cell Lysis and Heat Treatment:

30.9 g of E. coli.
↓
125 ml of 20mM Tris-HCl (pH8.0), 5mM B-ME, 50mM NaCl.
↓
Sonication, output 8, duty 50/2 min, 5 times.
↓
Heat 10 min, 70 deg C.
↓
Centrifuge at 100,000 rcf, 60 min.
↓
125 ml of supernatant.
↓
 
↓
 
 

Purification Procedure:

Step Column Buffer Gradient 1 Gradient 2 FlowRate (ml/min) Eluted
1 Resource Phe 6ml 20mM Tris-HCl(pH8.0),5mM B-ME,1.2M AS 1.2M AS 0M AS 6 1.1M AS
2 Resource S 6 ml 50mM MES(pH5.7),5mM B-ME 0M NaCl 2M NaCl 6 72mM NaCl
3 HiLoad 16/60 Superdex 75 20mM Tris-HCl(pH8.0),5mM B-ME,150mM NaCl     1 66min
4            
5            
6            
7            
8            
9            

Comment:

Products were confirmed by MS.

2011.10.20

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