Efficient gene transfer into zebrafish skeletal muscle by intramuscular injection of plasmid DNA.
The ability of zebrafish skeletal muscles to internalize and express plasmid DNA was demonstrated using pCMVCAT1, a chloramphenicol acetyltransferase (CAT) construct driven by the human cytomegalovirus immediate early (CMV-IE) promoter. We found that CAT activity was correlated to the amount of plasmid DNA injected, with maximal expression at 5 micrograms of pCMVCAT1. CAT activity was also shown to increase steadily over the first seven days after injection, with high levels of CAT expression persisting up to one year. Intramuscular injection of CAT constructs driven by other viral promoters also resulted in high levels of CAT activity. Histochemical localization using a CMV beta-galactosidase construct confirmed that only myofibers at the site of injection expressed beta-galactosidase enzyme. The persistence and strong expression of injected plasmid constructs suggest that zebrafish may be a simple and readily accessible system for direct muscle injection studies.
Tan
J H
JH
School of Biological Sciences, National University of Singapore, Republic of Singapore.
Chan
W K
WK
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003587:Cytomegalovirus / Q000235:genetics; D015870:Gene Expression; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D017874:Immediate-Early Proteins / Q000235:genetics; D007273:Injections, Intramuscular; D008156:Luciferases / Q000235:genetics; D008668:Metallothionein / Q000235:genetics; D018482:Muscle, Skeletal; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012474:Salmon / Q000235:genetics; D015027:Zebrafish / Q000235:genetics*; D001616:beta-Galactosidase / Q000235:genetics
D017874:Immediate-Early Proteins; D008668:Metallothionein; D008156:Luciferases; D015500:Chloramphenicol O-Acetyltransferase; D001616:beta-Galactosidase