Detection of human leukocyte interferon-alpha A and -alpha 2 genes in genomic DNAs by the use of deoxyoctadecyloligonucleotide probes.

Abstract

Two deoxyoctadecyloligonucleotides complementary to the sequence spanning a single base substitution between human leukocyte interferon (HuIFN) alpha A and alpha 2 genes were efficiently used as probes to distinguish between HuIFN-alpha A and -alpha 2 genes. At 37 degrees C or 42 degrees C under aqueous conditions (0.9 M NaCl), hybridization between both probes and the alpha A and alpha 2 genes without any mismatch was strong, whereas the hybridization with one base mismatch (alpha A probe-alpha 2 gene and alpha 2 probe-alpha A gene) was very weak or negligible. Because the single base substitution of G in the alpha 2 gene for A in the alpha A gene provides an extra HinfI site in the alpha 2 gene at the center of the sequence hybridizing to the alpha 2 probe, digestion with HinfI restriction endonuclease caused complete loss of the hybridization between the alpha 2 probe and the alpha 2 gene. PvuII digestion provides 298-bp fragments hybridizing to the probes only from the alpha A and alpha 2 genes among the known HuIFN-alpha genes. Thus, with the use of these oligonucleotide probes in combination with PvuII and PvuII-HinfI restriction endonuclease digestion, the existence of the sequences corresponding to both IFN-alpha A and IFN-alpha 2 genes in human genomic DNAs was demonstrated. The results also surprisingly indicate that these genes, formerly considered alleles because of their essential identity (1 base pair difference in the coding sequence), are not likely to be alleles, but represent closely related distinct genes.