Further Characterization of HLA Typing Cell Lines at CCR-5 Receptor by a Gene Amplification and Detection Method

Abstract

The co-chemokine receptor 5 or CCR-5 is one of the major co-receptors for the macrophage-tropic (M-tropic) strain of HIV. The frequency of the 32 base pair (bp) deletion of CCR-5 is elevated in individuals who have had multiple exposures to HIV but remain uninfected. This is evidence suggests that the CCR-5 can act as a natural resistance to the infection. CCR-5 reduces the ability of an individual's cells to support an entry of the M-tropic virus, because the defective receptors are severely truncated. In this study, a typing method was designed specifically to detect the mutation that occurs on CCR-5 sequences. A gene amplification and detection method that involved polymerase chain reaction (PCR), restriction enzyme digestion, and gel electrophroresis was utilized. In order to establish this new amplification typing method, primers were assigned to yield a 73S-bp fragment spanning the site of the mutation, PCR condition optimized to increase the original of DNA product, and control Human Lymphocyte Antigens (HLAj) typing cell lines were tested to validate testing parameters. As a result, the functional data suggested that the designed reagents and method were found to be another effective way for detecting the mutation on cell lines, by defining the 32 bp deletion on the heterozygote mutant alleles, which furthered the characterization of HLA typing cell lines at the CCR-5 receptor. With this knowledge in understanding the mechanism of the resistance to HIV infection, it may provide additional information for inducing similar resistance in others and successfully combating AIDS.