Electroporation: application to human lymphoid cell lines for stable introduction of a transactivator gene of human T-cell leukemia virus type I.
Conditions were developed for stable introduction of foreign DNA into human lymphoid cell lines by electroporation. To introduce stably the p40 gene of human T-cell leukemia virus type I (HTLV-I) into the human lymphoid cell line Jurkat, the p40 expressing plasmid, pMAXRHneo-1, which carries the neo resistant gene, was transfected into Jurkat cells at a voltage of 2500 V and capacitance of 21.7 microF, and stable transformants were screened for neo (G418) resistance. The frequency of transformants was more than one per 2 x 10(5) cells used initially. Clones that were resistant to G418 were shown to have the p40 gene integrated into the host genome and to express mRNA and protein from the introduced plasmid. Expression of p40 in the transformed Jurkat cells was also confirmed by testing the trans-activating effect of HTLV-I enhancer by p40. High frequencies of stable transformations of 10(-4) to 10(-6) were also reproducibly obtained by electroporation of the human T cell lines HSB-2 and TALL-1, a human B cell line Raji, a human monocytic cell line U937, and a human erythroleukemia cell line K562. These results demonstrate that electroporation is a very efficient method for introducing foreign DNA into human lymphoid cell lines.
Ohtani
K
K
Institute for Virus Research, Kyoto University, Japan.
Nakamura
M
M
Saito
S
S
Nagata
K
K
Sugamura
K
K
Hinuma
Y
Y
D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D004558:Electric Stimulation / Q000379:methods; D005786:Gene Expression Regulation; D005814:Genes, Viral; D015368:Human T-lymphotropic virus 1 / Q000235:genetics*; D006801:Humans; D010957:Plasmids; D015183:Restriction Mapping; D013601:T-Lymphocytes; D014162:Transfection