| NaCl |
82.0 |
478.9 |
82.0 |
478.9 |
82.0 |
478.9 |
| KCl |
4.9 |
36.3 |
4.9 |
36.3 |
4.9 |
36.3 |
| KH2PO4 |
1.2 |
15.9 |
1.2 |
15.9 |
1.2 |
15.9 |
| MgSO4 · 7H2O
|
1.2 |
29.1 |
1.2 |
29.1 |
1.2 |
29.1 |
| NaHCO3 |
25.0 |
210.0 |
15.0 |
126.0 |
25.0 |
210.0 |
| Glucose |
5.6 |
100.0 |
5.6 |
100.0 |
5.6 |
100.0 |
| Sodium pyruvate |
0.3 |
2.9 |
0.3 |
2.9 |
0.3 |
2.9 |
| CaCl2 · 2H2Oc |
1.7 |
25.1 |
1.7 |
25.1 |
|
|
| SrCl2 · 6H2Oc |
|
|
|
|
2.5–10.0 |
66.7–266.6 |
| HEPES |
|
|
10 |
238.0 |
|
|
| Glutaminec |
1.0 |
14.6 |
1.0 |
14.6 |
1.0 |
14.6 |
| Sodium lactate (60% syrup) |
20 |
370 µL |
20 |
370 µL |
20 |
370 µL |
| EDTA |
0.10 |
3.8 |
0.10 |
3.8 |
0.10 |
3.8 |
| Polyvinyl alcohold |
|
10.0 |
|
10.0 |
|
10.0 |
| BSA |
|
5 mg/mL |
|
|
|
5 mg/mL |
| CZB-based media are used with slight modifications for culturing cloned embryos and micromanipulating oocytes in vitro. These
three media should be made every 2 wk. Sterilize with Millipore filters (0.45 µm), dispense into 5-mL plastic tubes (Falcon
2057), and store at 4°C until they are used. Add glutamine and CaCl2 (or SrCl2) on the day of the experiment.
|
| aKSOM medium can also be used successfully to culture cloned mouse embryos.
|
| bThe CZB-HEPES micromanipulation medium does not contain bovine serum albumin (BSA), which can decrease the survival rate of
oocytes after injection. Bovine calf serum (10%–20%) may be used instead of BSA.
|
| cCaCl2, SrCl2, and glutamine are added from 100×, 10×, and 100× stock solutions, respectively.
|
| dPolyvinyl alcohol (cold-water soluble, e.g., Sigma-Aldrich P8136) is first dissolved in ultrapure water for 1 h at 80°C.
|
