Genetic Variation And STS Molecular Markers For LOX Activity In Wheat

The lipoxygenase (LOX) in wheat grains has double effects to grains quality and flour quality. Low LOX activity even the deletion of LOX gene can effectively alleviate the oxidation of lipids, lightening the deterioration of grains and prolonging the storage life of grain. Additional, high LOX activity in grains can improve whiteness of flour. In this paper, the variation and distribution characteristic of LOX activity in wheat grains were studied to screen some germplasm with low or high LOX activity and advanced traits. In this basic, A STS marker for LOX activity was developed and validated. In addition, expression of LOX gene in different wheat varieties and in aging process was analyzed by fluorescence quantitative polymerase chain reaction(QPCR) techonolyg.The main results obtained in this study are summarized below.1. The results of genotypexenvironment interaction indicated the variation of genotype had greatest influence on LOX activity, while environment had minimal impact on it. And the effect of genotypex-environmentxplace interaction was more than genotype xplace interaction.109 wheat varieties were planted (in Hefei, Anhui province) in four years to determined LOX activity, and the result showed the variation of LOX activity of the experimental varieties in four crop seasons were significant and the coefficient of variation were all more than 25.0%, also there were highly significant correlation among LOX activity of four crop seasons, which indicated that LOX activity was mainly controlled by gene. In this study, RIL-4 population was constructed by "Lumai 22×Yangmai 9". the LOX activity of this population was varied discontinuously and showed skewed normal distribution characteristic, and that suggested that LOX activity of wheat be a quanitity trait and controlled by multi-genes.2. Baseed on two LOX genes reported in NCBI, some primers were designed to screen polymorphism of PCR products with 12 wheat varieties in which 6 with high LOX activity and the other 6 varieties with low LOX activity. The results of electro-phoresis showed the primer wp02 amplified a (bp), b(bp) and c(bp) fragments in 6 varieties with high LOX activity, while b and c fragments of PCR product in varieties with LOX activity. Using wp02 primer to test 212 varieties, the type with a, b and c fragments was detected in 174 varieties, in that there were 70.11% of total varieties had high LOX activity, but only 38 genotypes were amplified with a and c fragment and there were 78.95% of the total had low LOX activity. multiple comparisons revealed, there were extreme significance between abc fragments typ and ac fragment type. Chi square test of independence(X2=26.49**) suggested the marker was significant related with LOX activity. So, this marker could be used in MAS aimed at LOX activity. Using a set of nulli-tetrasomic lines of Chinese Spring, the marker wp-02 was located on chromosomes 4B.3. The analysis of geographical distribution showed the means and range of LOX activity were differed in different provinces. The means of LOX activity were higher in some varieties from Henan, Shangdong and Beijing, but lower in those fromforeign and Hebei province. The range of LOX activity was less in cultivars in Beijing than those in other provinces. In addition, using STS marker to detect the materials, the PCR products showed, the cultivars which amplified fragment with a, b and c were from Beijing, Shangdong and Henan, and implied gerplasms with high LOX activity is dominant in these areas. While the cultivars which amplified fragment with a and c were advanced in the foreign and the southen part of our country. In general, the cultivars with high LOX activity were in the majority in our country, in this study 10 varieties with low LOX activity and 10 with high activity had been screened.4.The LOX activity and carotenoids concerntration were determined spectrophotometrically in 63 wheat varieties. The results showed there were significant or highly significant variation of LOX activities and carotenoids concerntration and other color indexs both in fresh and stored whole flours. During storage, LOX activity was only significantly negatively correlated with carotenoids concerntration, whereas no significant relationship with the whole flour whiteness, brightness and yellowness. According to our results, the LOX activity of wheat grains could not evaluate the storage quality of wheat. From the study, there were not obvious effect of STS marker, that with a, b and c fragment, on aging index. But in some cultivars that implified a and c fragment(deletion of b fragment), the aging indexes were all low, especially in P206 and Wanmai44 which aging index were 3. So, to some extend, the deletion of b fragment was related to wheat seed storage quality.5. The results of QPCR showed that the expression amounts of LOX2 gene in 3 wheat cultivars were different, especially the amount of Huancheng3366 was highest. The ranking list of relative expression amounts of LOX gene in 3 cultivars was: Huncheng 3666>Luomaill>Annong0305. In the process of aging of seed, the expression amounts of LOX2 gene was induced by aging time. For all cultivars, following the increase of aging time, the expression amount of LOX2 gene presented same tendency:declined first, followed by a increase, and then declined again.