Method for purifying pyruvate kinase
A technology of pyruvate kinase and crude enzyme solution, which is applied in the direction of transferase, etc., and can solve the problems of easy clogging of affinity adsorption columns or desalting columns
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0061] (1) Pyruvate kinase expression strain: according to the method of patent application CN101698837, a recombinant host cell—Escherichia coli BL21(DE3)—PET286PK was obtained.
[0062] (2) Preparation of pyruvate kinase crude enzyme solution
[0063] Prepare 6 liters of LB medium and add it to a 10-liter fermenter, sterilize at 121°C for 15 minutes, and add kanamycin after complete cooling to make the final concentration 50 μg / ml. At the same time, 1000 ml of LB culture medium was prepared and placed in three 1000 ml Erlenmeyer flasks, the mouths of which were sealed with air-permeable membranes, and sterilized at 121° C. for 15 minutes.
[0064] The above-mentioned Escherichia coli "BL21(DE3)-pET286PK" was inoculated in 300 ml of culture medium (add kanamycin to 50 μg / ml before inoculation), and a total of 3 bottles were connected (one bottle for spare, one bottle for use) To track and detect the bacterium concentration), at 37 ℃, 200 rev / min culture on the shaker, measur...
Embodiment 2
[0071] (1) Pyruvate kinase expression strain: According to the method of patent application CN101698837, the recombinant host cell—Escherichia coli BL21(DE3)—PET286PK was obtained
[0072] (2) Preparation of pyruvate kinase crude enzyme solution
[0073] Prepare 6 liters of LB medium and add it to a 10-liter fermenter, sterilize at 121°C for 15 minutes, and add kanamycin after complete cooling to make the final concentration 50 μg / ml. At the same time, 1000 ml of LB culture medium was prepared and placed in three 1000 ml Erlenmeyer flasks, the mouths of which were sealed with air-permeable membranes, and sterilized at 121° C. for 15 minutes.
[0074] The above-mentioned Escherichia coli "BL21(DE3)-pET286PK" was inoculated in 300 ml of culture medium (add kanamycin to 50 μg / ml before inoculation), and a total of 3 bottles were connected (one bottle for spare, one bottle for use) To track and detect the bacterium concentration), at 37 ℃, 200 rev / min culture on the shaker, measu...
Embodiment 3
[0081] (1) Pyruvate kinase expression strain: According to the method of patent application CN101698837, the recombinant host cell—Escherichia coli BL21(DE3)—PET286PK was obtained
[0082] (2) Preparation of pyruvate kinase crude enzyme solution
[0083] Prepare 6 liters of LB medium and add it to a 10-liter fermenter, sterilize at 121°C for 15 minutes, and add kanamycin after complete cooling to make the final concentration 50 μg / ml. At the same time, 1000 ml of LB culture medium was prepared and placed in three 1000 ml Erlenmeyer flasks, the mouths of which were sealed with air-permeable membranes, and sterilized at 121° C. for 15 minutes.
[0084] The above-mentioned Escherichia coli "BL21(DE3)-pET286PK" was inoculated in 300 ml of culture medium (add kanamycin to 50 μg / ml before inoculation), and a total of 3 bottles were connected (one bottle for spare, one bottle for use) To track and detect the bacterium concentration), at 37 ℃, 200 rev / min culture on the shaker, measu...
PUM
| Property | Measurement | Unit |
|---|---|---|
| molecular weight | aaaaa | aaaaa |
Abstract
Description
Claims
Application Information
Login to View More