PMID: 7090350Mar 1, 1982Paper

Transfection conditions for baculovirus DNA in an insect cell line

Voprosy virusologii
E N Oreshkin


The following conditions were found to be necessary to achieve infection of a Spodoptera frugiperda cell line with purified DNA of nuclear polyhedrosis virus of Spodoptera exempta (NPV): (1) cell monolayers shall be prepared one day before the experiment; the number of cells added to 35-mm wells of plastic clusters (Falcon) shall be 1.0 X10(6)-1.1X10(6), and the cells shall be in the stage of logarythmic growth; DNA shall be diluted in HeBS-buffer (pH 6.95-7.05); the volume of the mixtures added per well shall be 0.3 to 0.5 ml; (2) DNA calcium phosphate mixture shall be prepared for 20-30 min at 20 degrees C; (3) the cells shall be inoculated with the DNA-calcium-phosphate mixture for 45-60 min at 28 degrees C; (4) dimethylsulphoxide (DMSO) shall be added 4 hours post-inoculation for 4 min; (5) the upper agarose layer (0.75% agarose) shall be added after 2 hours of incubation in the cell growth medium TC-100 after DMSO had been added. Under these conditions the specific activity of Spodoptera exempta NPHV DNA may achieve the value of 2.26 X 10(3) plaque-forming centres per 1 microgram.

Related Concepts

DNA, Viral
Insect Viruses
Study, Methodological

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