Protocol for Culturing Streptavidin:
Preparation of Materials:
Basic Salts and Trace Elements (TE+BS):
| Salt | Amount |
|---|---|
| H3BO3 | 500 ug |
| CuSO4 5H2O | 40 ug |
| KI | 100 ug |
| FeCl3 6H2O | 200 ug |
| MnSO4 H2O | 400 ug |
| NaMoO4 | 200 ug |
| ZnSO4 7H2O | 400 ug |
| KH2PO4 | 1.0g |
| MgSO4 7H2O | 0.5 g |
| NaCl | 0.1g |
| CaCl2 2H2O | 0.1g |
SLMA:
| Element | Amount |
|---|---|
| L-Asn | 7.0 g |
| Dextrose | 10.0g |
| TE+BS | 1.7g |
| K2HPO4 | 1.0g |
| water | 1L |
SLMB:
| Element | Amount |
|---|---|
| L-Asn | 7.0 g |
| Dextrose | 10.0g |
| TE+BS | 1.7g |
| KH2PO4 | 1.0g |
| K2HPO4 | 1.0g |
| MgSO4 7H2O | 0.5g |
| NaCl | 0.1g |
| CaCl2 2H2O | 0.1g |
| water | 1L |
Procedure:
1) Introduce inoculums into flask (preferably 4L that has been washed and autoclaved) of SLMA on shaker for 200 rev/min @ RT
2) Incubate until heavily growth of balls
3) Spin cells down, discard supernatant and wash with PBS
4) Transfer to SLMB after 1 week in SLMA for optimal cell production
5) Spin cells down, take supernatant out and test on SDS page
6) Transfer to SLMA after 1 week in SLMB for SA production
7) Spin cells down, take supernatant out and test on SDS page
Estimated time for culture: 2.5 weeks
Protocol by Tricia Lin January 2009