Isolation of the gene encoding the human T-lymphocyte differentiation antigen Leu-2 (T8) by gene transfer and cDNA subtraction.

Abstract

The isolation of genomic and cDNA [complementary DNA] clones encoding the human T-lymphocyte cell-surface differentiation antigen, Leu-2 (T8), by use of a combination of transfection, fluorescence-activated cell-sorting and subtractive cDNA hydridization is reported. A cDNA library was constructed with mRNA from a mouse L-cell transfectant in which the human Leu-2 gene is expressed and amplified. Leu-2 cDNA clones were identified by screening with a selected cDNA probe from a second amplified Leu-2 transfectant. This probe contained cDNA species not removed by hybridization with L-cell mRNA. A Leu-2 cDNA clone was used to isolate a genomic clone. Transfection with DNA from this clone resulted in a high number of Leu-2 transfectants. This approach can be used to clone genes coding for other cell-surface molecules.