Evaluation of nuclear transfer and transcription of plasmid DNA condensed with protamine by microinjection: the use of a nuclear transfer score.
In the present study, the nuclear delivery of a green fluorescence protein (GFP)-encoding pDNA condensed by protamine was investigated in terms of trans-gene expression after cytoplasmic (E(cyt)) and nuclear (E(nuc)) microinjection. To compare the nuclear transfer process, a novel parameter; the nuclear transfer (NT) score was introduced. The E(cyt) value for protamine/pDNA particles increased in a charge ratio-dependent manner. The calculated NT score showed that this increase results from an enhancement in nuclear transfer efficiency, which was also quantitatively confirmed by a recently developed confocal image-assisted three-dimensionally integrated quantification (CIDIQ) method. Moreover, E(nuc) for protamine/pDNA particles was significantly higher than that for poly-L-lysine/pDNA particles, suggesting that pDNA, when condensed with protamine, is more accessible to intra-nuclear transcription. Collectively, protamine is an excellent DNA condenser, with bi-functional advantages: improvement in nuclear delivery and efficient intra-nuclear transcription.
Masuda
Tomoya
T
Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, Hokkaido, Japan.
Akita
Hidetaka
H
Harashima
Hideyoshi
H
D002467:Cell Nucleus; D004247:DNA / Q000008:administration & dosage* / Q000737:chemistry; D008845:Microinjections; D010957:Plasmids; D011479:Protamines / Q000737:chemistry*; D014158:Transcription, Genetic
D011479:Protamines; D004247:DNA