Abstract
Single nucleotide polymorphisms (SNPs) have important application value in the research of population genetics, hereditary diseases, tumors, and drug development. Conventional methods for detecting SNPs are typically based on PCR or DNA sequencing, which is time-consuming, costly, and requires complex instrumentation. In this study, we present a duplex probe-directed recombinase amplification (duplex-PDRA) assay that can perform real-time detection of two SNPs (rs6983267 and rs1447295) in four reactions in two tubes at 39°C within 30 min. The sensitivity of duplex-PDRA was 2×103-104 copies per reaction and no cross-reactivity was observed. A total of 382 clinical samples (179 prostate cancer patients and 203 controls) from northern China were collected and tested by duplex-PDRA assay and direct sequencing. The genotyping results were completely identical. In addition, the association analysis of two SNPs with prostate cancer risk and bone metastasis was conducted. We found that the TT genotype of rs6983267 (OR: 0.42; 95%CI: 0.23-0.78; P=0.005) decreased the risk of prostate cancer, while the CA genotype of rs1447295 (OR: 1.89; 95%CI: 1.20-2.96; P=0.005) increased the risk of prostate cancer. However, no association between the two SNPs (rs6983267 and rs1447295) and bone metastasis in prostate cancer was found in this study (P>0.05). In conclusion, the duplex-PDRA assay is an effective method for the simultaneous detection of two SNPs and shows great potential for widespread use in research and clinical settings.
Highlights
Single nucleotide polymorphism (SNP) is one of the most common forms of polymorphisms in the genome
The specificity results of duplex-probe-directed recombinase amplification (PDRA) showed that no cross-reactivity of duplex-PDRA assays was observed when up to 105 copies of recombinant plasmids were used as input templates per reaction
We found no significant difference in the genotyping distribution between rs6983267 and rs1447295 in patients with and without bone metastases in the prostate cancer (PCa) group (P40.05)
Summary
Single nucleotide polymorphism (SNP) is one of the most common forms of polymorphisms in the genome. In 2006, Amundadottir et al found in Iceland, Sweden, and other populations that multiple SNPs in the chromosome 8q24 region are closely related to the risk of prostate cancer (PCa) by a large genome-wide association study (GWAS) [3]. This correlation has been followed and confirmed in European Americans, African Americans and native Hawaiians, Spain, the Netherlands, Japan, and other peoples [4,5,6,7,8,9,10,11]. There is no relevant report on whether PCa bone metastasis is related to SNPs located on 8q24
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