To investigate kernel hardness, a compression test which is widely used to measure the hardness of individual kernels as a physical testing method was made simultaneously with the measurement of friabilin (15KDa) which is strongly associated with kernel hardness and was recently developed as a biochemical marker for evaluating kernel hardness in 79 Korean wheat varieties and experimental lines. With the scattered diagram based on the principal component analysis from the parameters of the compression test, 79 Korean wheat varieties were classified into three groups based on the principal component analysis. Since conventional methods required large amount of flour samples for analysis of friabilin due to the relatively small amount of friabilin in wheat kernels, those methods had limitations for quality prediction in wheat breeding programs. An extraction of friabilin from the starch of a single kernel through cesium chloride gradient centrifugation was successful in this experiment. Among 79 Korean wheat varieties and experimental lines 50 lines (63.3%) exhibited a friabilin band and 29 lines (36.7%) did not show a friabilin band. In this study, lines that contained high maximum force and the lower ratio of minimum force to maximum force showed the absence of the friabilin band. Identification of friabilin, which is the product of a major gene, could be applied in the screening procedures of kernel hardness. The single kernel analysis system for friabilin was found to be an easy, simple and effective screening method for early generation materials in a wheat breeding program for quality improvement.