| Autor: |
Timoteo, Vanessa Joy A., Nacis, Jacus S., Agarrado, Rod Erick L., Rodriguez, Marietta P., Dalmacio, Leslie Michelle M., Capanzana, Mario V. |
| Předmět: |
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| Zdroj: |
Philippine Journal of Science; 2019 Genomic issue, p283-296, 14p |
| Abstrakt: |
Cost-effective and high-throughput genotyping assays will enable the routine detection of single nucleotide polymorphisms (SNPs) in iron-related genes in the Philippines, where anemia due to iron deficiency remains a public health concern throughout the years. This paper presents the development of high-resolution melting (HRM) assays that were able to identify SNPs in HFE or homeostatic iron regulator gene (rs1800562, rs1799945); TF or transferrin gene (rs3811647, rs1799852); and TMPRSS6 or transmembrane serine protease 6 gene (rs4820268) in the Nutritional Genomics Laboratory of the Department of Science and Technology - Food and Nutrition Research Institute (DOST-FNRI NuGen Lab). Genomic DNA was extracted from 112 peripheral blood samples and SNP-genotyped using the Precision Melt Analysis? Software of Bio-Rad CFX96? Real-Time PCR (polymerase chain reaction) Detection System. Real-time amplification reactions were evaluated in terms of varying conditions in annealing temperature (Ta) and number of amplification cycles. HRM assays were then compared against that of the gold-standard sequencing technique using the validation parameters: sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). The ideal parameters of HRM assays that can genotype the target SNPs were determined. Genotype discrimination resulted in three genotypes of TF rs3811647, TF rs1799852, and TMPRSS6 rs4820268 with varying frequencies among samples used. Two genotypes were detected for HFE rs1799945, while only one genotype was obtained for HFE rs1800562. The confidence of genotype clusterings were at least 95%. All assays had 100% concordance with the sequencing results except for the TMPRSS6 rs4820268 assay, which achieved a sensitivity of 97%. Therefore, in line with the establishment of the DOST-FNRI NuGen Lab, HRM assays can be considered a reliable and affordable alternative to sequencing. Identifying genetically high-risk Filipinos in the diagnosis of anemia due to iron deficiency may be facilitated in the future. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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