Whole Cell Project Database -Purification-

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Thermus thermophilus HB8  TTHB223
CRISPR-associated protein, Cas2

Lot Information:

Purification Lot: PC014223-42-02-02
sPlasmid ID: PC014223-42
Plasmid Vector: pET-HisTEV
Label: SeMet
Person: Takemoto N.
Purified Date: 02/26/2007
Total Protein Yield: .8 mg
Final Volume: 0.1 mL
Conc.: 7.7 mg/mL

Final SDS-PAGE:

Culture Lot:

date

host cell

scale

SDS-PAGE

02/16/2007Rosetta2 834(DE3)pLysS9 L

Cell Lysis and Heat Treatment:

22.3g of E. coli cells
Ultra Sonication with 20mM Tris-HCl + 500mM NaCl (pH8.0) + 5mM 2-ME + 1mM PMSF
Heat Treatment at 70°C for 13 min
Ultra Cengrifuge at 200,000G for 1 hr

Purification Procedure:

ColumnBufferFlow Rate (mL/min)Eluted
HisTrap HP 5ml20 mM Tris-HCl (pH 8.0), 0.5 M NaCl, 20 mM Imidazole50.5M Imidazole
HiLoad 16/60 Superdex 200 pg20 mM Tris-HCl (pH 8.0), 200 mM NaCl, 20 mM Imidazole1.591ml
HisTrap HP 5ml20 mM Tris-HCl (pH 8.0), 0.5 M NaCl, 20 mM Imidazole50.5M Imidazole
Resource S 6ml20 mM Tris-HCl (pH 8.0), 10 mM NaCl, 20 mM Imidazole4Flowthrough
HisTrap HP 5ml20 mM Tris-HCl (pH 8.0), 0.5 M NaCl, 20 mM Imidazole50.5M Imidazole
Resource Q 6ml20 mM Tris-HCl (pH 8.0), 10 mM NaCl, 20 mM Imidazole40.27M NaCl