Genetic characterization kernel polyphenol oxidases in wheat (Triticum spp.) and its cultivated and wild relatives

Authors: Beecher, Brian; FUERST, E - WASHINGTON STATE U

Submitted to: American Association of Cereal Chemists Meetings
Publication Type: Proceedings
Publication Acceptance Date: 7/1/2007
Publication Date: 11/15/2007
Citation: Beecher, B.S., Fuerst, E.P. 2007. Genetic characterization kernel polyphenol oxidases in wheat (Triticum spp.) and its cultivated and wild relatives. American Association of Cereal Chemists Meetings.

Interpretive Summary: Polyphenol oxidase (PPO, EC, 1.10.31) is a major cause of discoloring in raw dough containing wheat flour. PPO is a ubiquitous enzyme that occurs in many tissues of the wheat plant, including the outer layers of wheat kernels. High levels of flour PPO have been associated with diminished end-product color and brightness in a variety of Asian noodle products. This is because PPO catalyzes the oxidation of a number of endogenous phenolic substrates in flour, producing the dark pigmented products that diminish product quality. However, breadwheat is composed of the genomes of three grass species, and the genetic source(s) of PPO activity are not well understood. The aim of this study was to characterize the PPO genes and activity in wheat's diploid and tetraploid relatives.

Technical Abstract: Polyphenol oxidase (PPO, EC, 1.10.31) is a major cause of discoloring in raw dough containing wheat flour. PPO is a ubiquitous enzyme that occurs in many tissues of the wheat plant, including the outer layers of wheat kernels. High levels of flour PPO have been associated with diminished end-product color and brightness in a variety of Asian noodle products. This is because PPO catalyzes the oxidation of a number of endogenous phenolic substrates in flour, producing the dark pigmented products that diminish product quality. However, breadwheat is composed of the genomes of three grass species, and the genetic source(s) of PPO activity are not well understood. Tje aim of this study was to characterize the PPO genes and activity in wheat's diploid and tetraploid relatives. The L-DOPA colorimeteric assay was used to determine PPO activity levels in whole kernels. Northern and Western blot analysis was used to estimate PPO mRNA and protein levels in whole kernels respectively. Southern blot was used to estimate gene copy number in genomic DNA. PCR and DNA sequencing was used to confirm the presence of unique PPO sequences in each genotype. We also determined that each diploid wheat relative has active PPO enzyme expressed in the kernel. Analysis of genomic DNA indicates that the diploid wheat relatives have a few as two (A. speltoides and A tauchii) to as many as four (T. urratu) PPO genes that are wxpressed in developing kernels. However, we have also observed a substntial degree of variability in PPO protein and enzymatic activity levels between individual lines of the species A. speltoides and A. tauchii, indicating that a large amount of genetic variation exists to be exploited in reducing the PPO levels in breadwheat.