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Available from Riken BioResource Center.

Plasmid & Protein Purification Database -Purification-

Species  
Locus tag APE0405
Products  
Plasmid ID 4016
Vector  
Purification Lot APE0405_a
Purified Date 2005.10.3-10.11
Protein conc. (mg/ml) 13.44
Total volume (ml) 1.2
Total protein (mg) 16.13

Culture Lot:

date host cell scale label weight of bacteira (g)
        20.6

Cell Lysis and Heat Treatment:

20.6 g of E. coli.
103.1 ml of 20mM Tris-HCl (pH8.0), 5mM B-ME, 150mM NaCl.
Sonication, output 8, duty 50/2min, 8 times.
Heat 15 min, 85 deg C.
Centrifuge at 100,000 rcf for 50 min.
94 ml of supernatant.
 
 
 

Purification Procedure:

Step Column Buffer Gradient 1 Gradient 2 FlowRate (ml/min) Eluted
1 Resource Iso 6ml 20mM Tris-HCl(pH8.0),5mM B-ME,1.5M AS 1.5M AS 0M AS 4 0.8M AS
2 Resource Q 6 ml 20mM Tris-HCl(pH8.0),5mM B-ME 0M NaCl 1M NaCl 4 0.27M NaCl
3 HiLoad 16/60 Superdex 75 20mM Tris-HCl(pH8.0),5mM B-ME,150mM NaCl     1 65min
4            
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9            

Comment:

Products containing less contaminant from Reso Q column were concentrated with a gel purification.

2011.10.20

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