Testis-mediated gene transfer in mice: comparison of transfection reagents regarding transgene transmission and testicular damage.
Testis-mediated gene transfer (TMGT) has been used as in vivo gene transfer technology to introduce foreign DNA directly into testes, allowing mass gene transfer to offspring via mating. In this study, we used plasmid DNA (pEGFP-N1) mixed with dimethylsulfoxide (DMSO), N,N-dimethylacetamide (DMA) or liposome (Lipofectin) in an attempt to improve TMGT. Males receiving consecutive DNA complex injections were mated to normal females to obtain F0 progeny. In vivo evaluation of EGFP expression, RT-PCR and PCR were used to detect the expression and the presence of exogenous DNA in the progeny. We also evaluated possible testicular damage by histological procedures. PC R and RT-PCR analyses revealed that liposome and DMSO increased the rate of TMGT. Histological analyses demonstrated that repeated (4 times) injections of DNA complexes can affect spermatogenesis. DMSO was the most deleterious among the reagents tested. In this study, we detected the presence of transgene in the progeny, and its expression in blood cells. Consecutive injections of DNA complexes were associated with impaired spermatogenesis, suggesting requirement of optimal conditions for DNA delivery through TMGT.
Núcleo de Biotecnología, Centro de Desenvolvlmento Tecnológico, Campus Universitario, Unlversldade Federal de Pelotas, Pelotas, RS, Brazil.
Marta G
MG
Amaral
Vinicius F
VF
Campos
Fabiana K
FK
Seixas
Paulo V
PV
Cavalcanti
Lisiane P R
LP
Selau
Joao C
JC
Deschamps
Tiago
T
Collares
D000818:Animals; D030801:Animals, Genetically Modified; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D005260:Female; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000008:administration & dosage / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000008:administration & dosage* / Q000032:analysis / Q000235:genetics; D008081:Liposomes / Q000494:pharmacology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic / Q000235:genetics*; D016133:Polymerase Chain Reaction; D013737:Testis / Q000187:drug effects / Q000473:pathology; D014162:Transfection / Q000379:methods; D019076:Transgenes
D008081:Liposomes; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D004121:Dimethyl Sulfoxide