Rapid detection of the CCR2-V64I, CCR5-A59029G and SDF1-G801A polymorphisms by tetra-primer PCR.
OBJECTIVE
To develop tetra-primer PCR assays for detection of the CCR2-V64I, CCR5-A59029G and SDF1-G801A polymorphisms associated with HIV pathogenesis.


METHODS
For each assay, two primers for the amplification of the gene locus are combined in one tube with two primers for the subsequent allele specific amplification (ASA). In the first set of cycles, pre-amplification of the gene region of interest is ensured by the gene specific primers. In the second set of cycles, lowering the annealing temperature allows ASA on the newly produced template.


RESULTS
Analysis of 90 DNA samples resulted in allele frequencies for CCR2-V64I, CCR5-A59029G and SDF1-G801A which are similar to other Caucasian cohorts. Furthermore, re-analysis of sequenced genomic DNA by tetra-primer PCR analysis (7-11 times) always showed identical results.


CONCLUSIONS
Our set of single-tube assays allows rapid and reproducible genotyping of the CCR2-V64I, CCR5-A59029G and SDF1-G801A polymorphisms. These inexpensive but accurate assays are valuable for screening these polymorphisms in cohorts of HIV-infected patients.
Hersberger
Martin
M
Institute of Clinical Chemistry, Division of Infectious Diseases and Hospital Hygiene, University Hospital of Zurich, Raemistrasse 100, CH-8091 Zurich, Switzerland. hmr@ikc.unizh.ch
Marti-Jaun
Jacqueline
J
Hänseler
Edgar
E
Speck
Roberto F
RF
D054377:Chemokine CXCL12; D019743:Chemokines, CXC / Q000235:genetics*; D017931:DNA Primers / Q000235:genetics*; D005787:Gene Frequency; D005820:Genetic Testing / Q000191:economics / Q000379:methods; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic / Q000235:genetics*; D054390:Receptors, CCR2; D019713:Receptors, CCR5 / Q000235:genetics*; D019707:Receptors, Chemokine / Q000235:genetics*; D015203:Reproducibility of Results; D013997:Time Factors; D044465:White People / Q000235:genetics
C515334:CCR2 protein, human; C515255:CXCL12 protein, human; D054377:Chemokine CXCL12; D019743:Chemokines, CXC; D017931:DNA Primers; D054390:Receptors, CCR2; D019713:Receptors, CCR5; D019707:Receptors, Chemokine