A primer set and kit for detecting gene variants in glucose-6-phosphate dehydrogenase (G6PD) deficiency.

Abstract

This invention discloses a primer set and kit for detecting gene variant sites in glucose-6-phosphate dehydrogenase (G6PD) deficiency, belonging to the field of gene detection technology. The primer set includes multiplex PCR amplification primers shown in SEQ ID NO. 1–14 and single-base extension primers shown in SEQ ID NO. 15–35, capable of simultaneously detecting 21 G6PD pathogenic variant sites (including high-incidence sites in the Chinese population such as c.95A>G and c.202G>A). The kit also includes PCR amplification reaction reagents, SAP enzymatic digestion reaction reagents, extension reaction reagents, desalting resin, and a detection chip. The detection method employs “multiplex PCR amplification → SAP dephosphorylation → single-base extension → nucleic acid mass spectrometry detection,” automatically determining the genotype based on mass-to-charge ratio differences. This invention offers comprehensive site coverage (covering pathogenic mutations in over 99% of Chinese patients), high accuracy (100% consistent with Sanger sequencing results), ease of operation (automatic detection and result analysis), and suitability for small sample sizes. It can be applied to prenatal diagnosis and newborn genetic disease screening, and is suitable for large-scale promotion in hospitals at all levels, providing a reliable tool for rapid and accurate detection of G6PD deficiency.

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