A biolistic process for in vitro gene transfer into chicken embryos.
Chicken embryos kept in culture medium were bombarded using a high helium gas pressure biolistic device. To optimize the factors that affect transformation efficiency, the lacZ gene under control of the human cytomegalovirus immediate early enhancer/promoter was used as a reporter gene. There was an inverse relationship between survival rate and transformation efficiency. The best conditions obtained for high embryo survival and high transformation efficiency were achieved with 800 psi helium gas pressure, 500 mmHg vacuum, gold particles, an 8 cm DNA-coated microparticle flying distance to the embryo and embryo placement 0.5 cm from the center of the particle dispersion cone. Under these conditions, transformation efficiency was 100%, survival rate 25% and the number of expression units in the embryo body cells ranged from 100 to 1,000. Expression of green fluorescent protein was also detected in embryos bombarded under optimal conditions. Based on the results obtained, the biolistic process can be considered an efficient method for the transformation of chicken embryos and therefore can be used as a model system to study transient gene expression and tissue-specific promoters.
Ribeiro
L A
LA
Departamento de Produção Animal, Escola Superior de Agricultura Luiz de Queiroz, Universidade de São Paulo, Piracicaba, SP, Brasil.
Mariani
P D
PD
Azevedo
J L
JL
Rech
E L
EL
Schmidt
G S
GS
Coutinho
L L
LL
D000818:Animals; D019470:Biolistics; D002642:Chick Embryo; D015870:Gene Expression; D017930:Genes, Reporter; D006371:Helium; D066298:In Vitro Techniques; D011312:Pressure
D006371:Helium