Detection of high-molecular-weight glutenin subunit genes for 1Dx2 and 1Dx5 using loop-mediated isothermal amplification assay

High-molecular-weight glutenin subunits (HMW-GS) in wheat grain are the major determinants of dough elasticity and viscosity and thus of bread-making quality. PCR-based molecular markers designed based on DNA polymorphisms were used to analyze HMW-GS genes in wheat. The loop-mediated isothermal amplification (LAMP) assay is a simple and rapid method for specific detection of genomic DNA target sequences. In the present study, we designed a set of LAMP markers by targeting the unique sequences of 1Dx2 and 1Dx5 genes. The primers could effectively distinguish the 1Dx2 and 1Dx5 genes from other genes at the Glu-1 locus. The results were confirmed by agarose gel electrophoresis. For visualization, ethidium bromide was used, and fluorescence only appeared in the positive samples. Under optimal conditions, the detection could be finished in 1 h. Thirty-eight wheat cultivars with known HMW-GS were used to validate LAMP markers for 1Dx2 and 1Dx5 genes. Only DNA samples with target genes could be amplified, and the results could be read easily using this method. The tests using LAMP were easy to perform, rapid, and sensitive. Thus, the current study results have the potential to be a powerful tool for the detection of HMW-GS genes in wheat.