Effect of DNA concentration on transgenesis rates in mice and pigs.
A retrospective analysis of transgenesis rates obtained in seven pronuclear microinjection programs was undertaken to determine if a relationship existed between the amount of DNA injected and transgenesis rates in the pig. Logistic regression analysis showed that as the concentration of DNA injected increased from 1 to 10 ng/microl, the number of transgenics when expressed as a proportion of the number liveborn (integration rate) increased from 4% to an average of 26%. A similar relationship was found when the number of molecules of DNA injected per picolitre was analysed. No evidence was obtained to suggest either parameter influenced integration rate in mice when the same constructs were injected. The number of transgenics liveborn when expressed as a proportion of ova injected (efficiency rate), increased as DNA concentration increased up to 7.5 ng/microl and then decreased at 10 ng/microl for both species suggesting that at this concentration DNA (or possible contaminants) may have influenced embryo survival. The relationship between efficiency and the number of molecules injected per picolitre was complex suggesting that the concentration at which DNA was injected was a better determinant of integration and efficiency rates. In conclusion, the present study suggests that transgenes need to be injected at concentrations of between 5 and 10 ng/microl to maximise integration and efficiency rates in pigs.
Nottle
M B
MB
BresaGen Limited, Adelaide, South Australia. mark.nottle@adelaide.edu.au
Haskard
K A
KA
Verma
P J
PJ
Du
Z T
ZT
Grupen
C G
CG
McIlfatrick
S M
SM
Ashman
R J
RJ
Harrison
S J
SJ
Barlow
H
H
Wigley
P L
PL
Lyons
I G
IG
Cowan
P J
PJ
Crawford
R J
RJ
Tolstoshev
P L
PL
Pearse
M J
MJ
Robins
A J
AJ
d'Apice
A J
AJ
D000818:Animals; D030801:Animals, Genetically Modified; D015139:Blotting, Southern; D004247:DNA / Q000032:analysis; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D018014:Gene Transfer Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008822:Mice, Transgenic; D012189:Retrospective Studies; D013552:Swine; D019076:Transgenes
D018076:DNA, Complementary; D004247:DNA