Journal of Animal Science, Vol 68, Issue 2 449-453, Copyright © 1990 by American Society of Animal Science

JOURNAL ARTICLE

Incorporation and expression of a neo gene after transfer to caprine hematopoietic cells using a modified retrovirus

D. C. Casey and K. L. White
Dept. of Anim. Sci., Louisiana State University, Baton Rouge.

The ability of the N2 retrovirus to introduce the selectable neo gene into (transform) caprine hematopoietic cells (CHC) was evaluated. Helper-free amphotropic retrovirus producing cells (RPC) were plated at approximately 1.0 x 10(5) cells per 25 cm2 tissue culture flask, cultured to 80% confluence and irradiated (1,500 rads) prior to CHC incubation. The CHC collected from three donor goats were washed and cultured for 24 h in either the RPC or control flasks. Cells were cultured in Dulbecco's Modified Eagles Medium containing 10% fetal calf serum (FCS) (DMEM) and 4 micrograms polybrene/ml. After 48 h of culture in fresh DMEM, cells were recovered and suspended in Iscove's DMEM supplemented with .3% gar, 12% FCS and 400 micrograms geneticin/ml (G418; neomycin) and transferred to 35-mm petri dishes (7.5 x 10(5) cells) for selection of G418 resistant cells. After 17 d of culture, plates were evaluated for total number of colony forming units (CFU, greater than 10 cells). Total number of CFU was greater (P less than .01) in treatment samples (means = 175, SEM = 70) than in control cultures (mean = 0, SEM = 0). The N2 retrovirus appears to be an effective vector for the transformation of CHC and may provide a means to introduce gene(s) into cells of domestic animals.