A new method to transfect the hypoblast of the chick embryo reveals conservation of the regulation of an Otx2 enhancer between mouse and chick extraembryonic endoderm

Table 1 Comparison of transfection efficiencies

Set	% Lipo-complex	Transfection time	n	(n) % efficiency	Level of expression	(n) % embryo success
A	33%	30 min	8	(4/8) 50%	low	(4/4) 100%
B		60 min	6	(5/6) 83%	low	(4/5) 80%
C		90 min	9	(9/9) 100%	medium	(9/9) 100%
D	66%	30 min	8	(4/8) 50%	medium	(3/4) 75%
E		60 min	7	(7/7) 100%	high	(5/7) 71%
F		90 min	8	(8/8) 100%	high	(6/8) 75%
G	100%	30 min	6	(5/6) 83%	medium	(2/5) 40%
H		60 min	7	(7/7) 100%	high	(3/7) 43%
I		90 min	8	(8/8) 100%	high	(1/8) 13%

Different combinations of transfection period and Lipofectamine-complex were tested. The efficiency of transfection is based on the number of embryos in which fluorescence could be detected following overnight culture. The 'level of expression' is an estimate of the proportion of cells expressing the reporter. Finally, the normality of the embryos ('embryo success') with fluorescent hypoblasts was judged as embryos that developed with a normal axis extension and orientation and in which the hypoblast developed in a characteristic fashion.

ISSN: 1471-213X