నైరూప్య

Rapid Screening Methods for Wheat Lines with Modified Gliadin Compositions: MALDI-TOF-MS and RP-HPLC Comparison

Erick Straw

A complex set of flour proteins called wheat gliadins can cause celiac disease and severe food sensitivities. As a result, new wheat lines with lower immunogenic potential are being created using mutation breeding and biotechnology techniques. The creation of quick, high-throughput technologies that can be applied as a first step in choosing lines with altered gliadin content is essential to these efforts. In this study, we improved Reversed-Phase High-Performance Liquid Chromatography (RP-HPLC) and Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF-MS) methods for the isolation of gliadins from Triticum aestivum cv. Chinese Spring (CS). Using the entire set of gliadin gene sequences recently obtained from this cultivar as well as a collection of aneuploid lines, we assessed the quality of the generated profiles in CS. By using MALDI-TOF-MS, the gliadins were divided into 13 peaks. There were multiple peaks of-or-gliadins, which are plausible targets for efforts to reduce the immunogenicity of flour and include a lot of celiac disease epitopes. However, other peaks, including one with as many as 12 distinct gliadins, contained several- and -gliadins. In contrast, 28 gliadin peaks were obtained from the separation of proteins using RP-HPLC, including 13 peaks that contained gliadins and 8 peaks that included gliadins. Even while RP-HPLC was able to separate gliadins more well than MALDI-TOF-MS, it was not possible to connect specific peaks to specific protein sequences. MALDI-TOF-MS and RP-HPLC both successfully separated gliadins. Although MALDI-TOF-MS is quicker and may be advantageous in investigations that focus on particular gliadins, an efficient technique that can be used more widely to identify changes in gliadin composition is RP-HPLC.