A polymerase chain reaction-based method for isolation of gene-specific sequences from the interferon-alpha gene cluster.
The interferon-alpha gene is a gene family of over 20 distinct genes having 80-95% homology with one another at a nucleotide level. Because of the high homology in the gene cluster, the available interferon-alpha gene probes can hybridize to multiple bands of different size on Southern blot analysis of restricted human genomic DNA. We used the polymerase chain reaction with the primers synthesized from Alu repetitive sequence and the conserved sequences of the interferon-alpha gene cluster to generate specific probes for individual interferon-alpha genes. The amplification products were subcloned into a plasmid vector and analyzed by DNA sequencing and Southern blotting of the restricted human placental DNA. One clone, which derived from interferon-alpha 14 gene, produced a single 5.2-kb band in Southern blots of the HindIII-restricted human placental DNA. This stands in contrast to the 10 bands of different size that were detected with a cDNA for the interferon-alpha I' gene. Our results indicate that a polymerase chain reaction-based method can be used to isolate gene-specific sequences from the interferon-alpha gene cluster. Since a variety of human cancers has been found to have the complete or partial deletion of the interferon-alpha gene cluster, the gene-specific probe generated by this method may aid in determining the breakpoints in the vicinity of the gene cluster.
Nobori
T
T
Department of Medicine, University of California, San Diego, La Jolla 92093-0663.
Reynolds
L
L
Orvis
L
L
Wu
D
D
Carson
D A
DA
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D016466:CHO Cells; D002899:Chromosomes, Human, Pair 9; D006224:Cricetinae; D004247:DNA / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D006822:Hybrid Cells; D016898:Interferon-alpha / Q000235:genetics*; D008969:Molecular Sequence Data; D005810:Multigene Family; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods*
D016898:Interferon-alpha; D004247:DNA