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Available from Riken BioResource Center.

Plasmid & Protein Purification Database -Purification-

Species  
Locus tag ST1096
Products  
Plasmid ID 0198
Vector  
Purification Lot ST1096_2
Purified Date 2005.6.8
Protein conc. (mg/ml) 17.15
Total volume (ml) 0.51
Total protein (mg) 8.7

Culture Lot:

date host cell scale label weight of bacteira (g)
        30

Cell Lysis and Heat Treatment:

30.0 g of E. coli.
↓
140 ml of 20mM Tris-HCl (pH8.0), 5mM B-ME, 50mM NaCl.
↓
Sonication, output 8/duty 50, 2 min, 5 times.
↓
Heat 15min, 75 deg C.
↓
Centrifuge at 100,000 rcf for 60min.
↓
120 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,2.0M AS 2.0M AS 0M AS 3 1.74M AS
2 Resource Q 6mL 20mM Tris-HCl(pH8.0),5mM B-ME 0M NaCl 1.0M NaCl 4 330mM NaCl
3 HiPrep 16/60 Heparin FF 20mM Tris-HCl(pH8.0),5mM B-ME 0M NaCl 1.5M NaCl 5 800mM NaCl
4 HiLoad 16/60 Superdex 75 20mM Tris-HCl(pH8.0),5mM B-ME,150mM NaCl     1 72ml
5 HiPrep 26/10 Desalting 53ml 20mM Tris-HCl(pH8.0),5mM B-ME        
6            
7            
8            
9            

Comment:

Products were confirmed by MS in the previous purification lot.

2011.10.20

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