21 Transgenie Animals
323
Table 1. Schedule for the production of transgenic rats
Time
Day
9A.M.
-3
11 A.M.
- 1
4P.M.
- 1
2A.M.
0
7-9 A.M. 0
9- 11 A.M. 0
1-4 P.M. 0
6P.M.
0
7-9 A.M.
Procedure
Im plant minipumps into immature rats. Pump rate is 1 U FSH
and 0.2 lU human choriogonadotropin (HCG)
Administer 30 lU HCG by intraperitoneal injection
Place hormone-treated female with fertile male formating;
place mature females with vasectomized males for mating
Ovulation and mating completed
Examine immature and mature females for the presence of a
copulatory (vaginal) plug
Sacrifize immature females and harvest oocytes
Microinjection
Reimplant surviving oocytes into pseudopregnant females or
Reimplant surviving two-cell embryos into day 1 or day 0
pseudopregnant females
lU HCG are injected intraperitoneally for induction and timing of ovulation. The subcutaneous implantation of the minipump is performed under
light ether anaesthesia by cutting the skin in the neck-region and preparing
a subcutaneous tunnel to the area somewhat lateral to the midline between
the posterior extremities.
Mating
The females carrying the minipumps are placed with fertile males for mating the day before microinjection. At the same time, mature females are
placed with vasectomized males for induction of pseudopregnancy. Vasectomy is carried out under anesthesia using a mixture ofRompun (Xylasin,
0.1 mg/100 g body-weight) and Ketavet (Ketamin, 11.5 mg/100 g bodyweight) in rats and Avertin in mice. Vasectomy is performed according
to the method previously decribed by Hogan et al. (13).
Note: Up to 15 mature females are required to obtain 2 -3 pseudopregnant
foster mothers. U sing females in estrus before mating will increase the yield
of recipients.
On the next morning between 7 and 8 A.M., egg donors and mature females are checked for the presence of a copulatory plug. In rats, careful
323
Table 1. Schedule for the production of transgenic rats
Time
Day
9A.M.
-3
11 A.M.
- 1
4P.M.
- 1
2A.M.
0
7-9 A.M. 0
9- 11 A.M. 0
1-4 P.M. 0
6P.M.
0
7-9 A.M.
Procedure
Im plant minipumps into immature rats. Pump rate is 1 U FSH
and 0.2 lU human choriogonadotropin (HCG)
Administer 30 lU HCG by intraperitoneal injection
Place hormone-treated female with fertile male formating;
place mature females with vasectomized males for mating
Ovulation and mating completed
Examine immature and mature females for the presence of a
copulatory (vaginal) plug
Sacrifize immature females and harvest oocytes
Microinjection
Reimplant surviving oocytes into pseudopregnant females or
Reimplant surviving two-cell embryos into day 1 or day 0
pseudopregnant females
lU HCG are injected intraperitoneally for induction and timing of ovulation. The subcutaneous implantation of the minipump is performed under
light ether anaesthesia by cutting the skin in the neck-region and preparing
a subcutaneous tunnel to the area somewhat lateral to the midline between
the posterior extremities.
Mating
The females carrying the minipumps are placed with fertile males for mating the day before microinjection. At the same time, mature females are
placed with vasectomized males for induction of pseudopregnancy. Vasectomy is carried out under anesthesia using a mixture ofRompun (Xylasin,
0.1 mg/100 g body-weight) and Ketavet (Ketamin, 11.5 mg/100 g bodyweight) in rats and Avertin in mice. Vasectomy is performed according
to the method previously decribed by Hogan et al. (13).
Note: Up to 15 mature females are required to obtain 2 -3 pseudopregnant
foster mothers. U sing females in estrus before mating will increase the yield
of recipients.
On the next morning between 7 and 8 A.M., egg donors and mature females are checked for the presence of a copulatory plug. In rats, careful