DNA dose and sequence dependence in sperm-mediated gene transfer.
We have tested three parameters in sperm-mediated gene transfer assays with mice and pigs: (i) the epididymal versus ejaculated origin of sperm cells, (ii) the primary structure, and (iii) the amount of the challenging foreign DNA. We have found that the pVLCNhGH construct, of retrotransposon origin, causes a massive embryo lethality and yet increases the yield of genetic transformation among born animals of both species compared to viral constructs. Arrest of embryonic development is a DNA dose-dependent effect, which is observed with high DNA doses, while lower doses are compatible with development. Finally, the overall efficiency of sperm-mediated gene transfer is higher when ejaculated, versus epididymal, spermatozoa are used. We suggest that this difference is related to the highly efficient apoptotic response in epididymal compared to ejaculated spermatozoa, triggered by the interaction of exogenous DNA molecules with the sperm membrane.
Sciamanna
I
I
CNR Institute of Biomedical Technology, Rome, Italy.
Piccoli
S
S
Barberi
L
L
Zaccagnini
G
G
Magnano
A R
AR
Giordano
R
R
Campedelli
P
P
Hodgson
C
C
Lorenzini
R
R
Spadafora
C
C
D000818:Animals; D030801:Animals, Genetically Modified; D000831:Animals, Newborn; D001483:Base Sequence; D015139:Blotting, Southern; D004247:DNA / Q000235:genetics*; D004542:Ejaculation; D005314:Embryonic and Fetal Development; D004822:Epididymis / Q000166:cytology; D005260:Female; D018628:Gene Dosage; D018014:Gene Transfer Techniques; D066298:In Vitro Techniques; D007315:Insemination, Artificial; D008297:Male; D051379:Mice; D013094:Spermatozoa; D013552:Swine
D004247:DNA