Development of a loop-mediated isothermal amplification marker for genotyping of the wheat Wx-B1 allele

Abstract

This study reports the development of a loop-mediated isothermal amplification (LAMP) reaction for genotyping of Wx-B1. The primer sets designed from the Wx-B1a and Wx-B1b sequences of wheat functioned most efficiently at 60 °C. In the LAMP reaction performed to detect Wx-B1a, DNA was amplified from the samples of individuals homozygous or heterozygous for Wx-B1a. However, DNA from individuals homozygous for Wx-B1b was not amplified. On the other hand, in the LAMP reaction performed to detect Wx-B1b, DNA was amplified from the samples of individuals homozygous or heterozygous for Wx-B1b. For easy genotyping, the NaOH extraction method and visualisation of DNA amplification by hydroxynaphthol blue were used. The results of genotyping by LAMP were consistent with those of genotyping by PCR using 32 F4 individuals of ‘Kinu-akari’ and ‘Syunyou’, which show the high reliability and usefulness of LAMP for marker-assisted selection.