Cereal Chem 61:129 - 135. | VIEW
Wheat Varietal Identification and Genetic Analysis by Reversed-Phase High-Performance Liquid Chromatography.
J. A. Bietz, T. Burnouf, L. A. Cobb, and J. S. Wall. Copyright 1984 by the American Association of Cereal Chemists, Inc.
Gliadins are a heterogeneous group of alcohol-soluble wheat endosperm storage proteins whose composition varies among wheat genotypes. Reversed-phase high-performance liquid chromatography (RP- HPLC) on large-spore columns gives high-resolution separations of gliadins from extracts of wheats, thereby permitting varietal identification and complementing other methods. Gliadins, extracted with 70% ethanol from single kernels or from bulk samples, are in jected onto a reversed-phase (C18) column having 300-Angstrom pores. The proteins are eluted with a gradient of increasing acetonitrile concentration, and are detected at 210 nm. Resulting chromatograms reveal approximately 30-40 components. Examples are presented that demonstrate that significant qualitative and quantitative differences occur between nearly all hard red winter, hard red spring, soft red winter, white, and durum wheats examined; only varieties having nearly identical pedigrees may be impossible to differentiate. Analysis of aneuploids permits determination of the chromosomal locations of genes that code proteins eluting in each chromatographic peak, thus establishing markers for the chromosomes. Analysis of phenotypic variants from a sample also permits detection of different genotypes, biotypes, or other off-types. RP-HPLC is fast, sensitive, exhibits good run- to-run reproducibility, has high resolving power, and is easily automated; the ability to accurately quantitate data may facilitate computer-assisted chromatographic comparisons. RP-HPLC of gliadins can be a valuable tool for indentification, selection, and comparison of genotypes in breeding and genetic studies.