[show abstract][hide abstract] ABSTRACT: The α-gliadins from Crithopsis delileana (Schult) Roshev (2n=2x=14, KK) were investigated by Acid polyacrylamide gel electrophoresis (A-PAGE) analysis. It was indicated
that the electrophoresis mobility of gliadins from C.delileana had obvious difference with those from common wheat in α, γ and ω region. Using primers designed from published sequences
of α-gliadin genes, three α-gliadin genes were isolated from C. delileana, which were designated as gli-ka1, gli-ka2 and gli-ka3, respectively. Two in-frame stop codons were found in the coding sequences of gli-ka3, indicating that gli-ka3 could be a pseudogene. The gli-ka2 was a gliadin with an odd number of cysteines, resulting from a non-synonymous mutation. This change might lead to the interactive
behavior of gli-ka2. Three α-gliadin genes of C. delileana had the similar but not identical primary structures to the corresponding gene sequences from other wheat related species.
By the alignment of α-gliadin genes from Triticeae, phylogenetic analysis indicated that three α-gliadin genes of C. delileana clustered together with all α-gliadin genes from Ee genome of Lophopyrum elongatum by an interior paralleled branch.
–α-gliadin gene–Phylogenetic analysis–Triticeae
[show abstract][hide abstract] ABSTRACT: Salicylic acid (SA) is one of the key signal molecules in regulating plant resistance to diverse pathogens. In Arabidopsis thaliana, it is predominantly associated with resistance against biotrophic and hemibiotrophic pathogens, and triggering systemic acquired resistance. In contrast, the effect of SA on the defence efficiency of wheat against fusarium head blight (FHB) and its causal agent, Fusarium graminearum, is still poorly understood. Here we show that the F. graminearum mycelial growth and conidia germination were significantly inhibited, and eventually halted in the presence of increasing concentration of SA in both liquid and solid media. Addition of SA also significantly reduced the production of the mycotoxin deoxynivalenol (DON). However the inhibitory effect of SA required acidic growth conditions to be observed while basic conditions allowed F. graminearum to use SA as a carbon source. High performance liquid chromatography (HPLC) analysis confirmed the capacity of F. graminearum to metabolize SA. To better understand the effect of SA on F. graminearum mycelial growth, we have compared the expression profiles of SA-treated and untreated F. graminearum liquid cultures after 8 and 24 h of treatment, using an F. graminearum custom-commercial microarray. The microarray analysis suggested that F. graminearum can metabolize SA through either the catechol or gentisate pathways that are present in some fungal species. Inoculation of F. graminearum conidia in a SA-containing solution has led to reduced FHB symptoms in the very susceptible Triticum aestivum cv. Roblin. In contrast, no inhibition was observed when SA and conidia were inoculated sequentially. The expression patterns for the wheat PR1, NPR1, Pdf1.2, and PR4 genes, a group of indicator genes for the defence response, suggested that SA-induced resistance contributed little to the reduction of symptoms in our assay conditions. Our results demonstrate that, although F. graminearum has the capacity to metabolize SA, SA has a significant and direct impact on F. graminearum through a reduction in efficiency of germination and growth at higher concentrations.
[show abstract][hide abstract] ABSTRACT: To accurately quantify gene expression using quantitative PCR amplification, it is vital that one or more ideal internal control genes are used to normalize the samples to be compared. Ideally, the expression level of those internal control genes should vary as little as possible between tissues, developmental stages and environmental conditions. In this study, 32 candidate genes for internal control were obtained from the analysis of nine independent experiments which included 333 Affymetrix GeneChip Wheat Genome arrays. Expression levels of the selected genes were then evaluated by quantitative real-time PCR with cDNA samples from different tissues, stages of development and environmental conditions. Finally, fifteen novel internal control genes were selected and their respective expression profiles were compared using NormFinder, geNorm, Pearson correlation coefficients and the twofold-change method. The novel internal control genes from this study were compared with thirteen traditional ones for their expression stability. It was observed that seven of the novel internal control genes were better than the traditional ones in expression stability under all the tested cDNA samples. Among the traditional internal control genes, the elongation factor 1-alpha exhibited strong expression stability, whereas the 18S rRNA, Alpha-tubulin, Actin and GAPDH genes had very poor expression stability in the range of wheat samples tested. Therefore, the use of the novel internal control genes for normalization should improve the accuracy and validity of gene expression analysis.
[show abstract][hide abstract] ABSTRACT: Using the homologous primers designed from known sequences of low-molecular-weight glutenin subunits (LMW-GS), four LMW prolamine gene sequences, designated as LMW-GSK1 (EU283813), LMW-GSK2 (EU283814), LMW-GSK3 (EU283815) and LMW-GSK4 (EU283816), were isolated from wheat-related diploid species Crithopsis delileana, among which LMW-GSK3 and LMW-GSK4 were partial gene sequences. These LMW prolamine gene sequences from C. delileana had the similar typical primary structures to the known LMW glutenin subunit genes in wheat and its related species. The 5′ flanking region of the LMW-GSK1 contained two cis-elements EM (5′-TGTAAAGT-3′) and GLM (5′-GGCGAGTCAT-3′). The deduced amino-acid sequence of the LMW-GSK1 consisted of a conserved signal peptide with 20 residues, a N-terminal region with 15 residues, a repetitive domain with 80 residues and a C-terminal region with 182 residues. Several in-frame stop codons were found in the coding sequences of LMW-GSK2 and LMW-GSK4, indicating that the LMW-GSK2 and LMW-GSK4 could be the putative pseudogenes. Phylogenetic analysis indicated that LMW-GSK1, LMW-GSK2 and LMW-GSK4 were closely related to the LMW-GS genes from A, D and R genomes, while LMW-GSK3 could be clustered together with X03103 from H genome. It was the first time that the coding genes of LMW prolamines on K genome of C. delileana were characterized.
[show abstract][hide abstract] ABSTRACT: alpha-Amylase inhibitors are attractive candidates for the control of seed weevils, as these insects are highly dependent on starch as an energy source. In this study, we aimed to reveal the structure and diversity of dimeric alpha-amylase inhibitor genes in wild emmer wheat from Israel and to elucidate the relationship between the emmer wheat genes and ecological factors using single nucleotide polymorphism (SNP) markers. Another objective of this study was to find out whether there were any correlations between SNPs in functional protein-coding genes and the environment.
The influence of ecological factors on the genetic structure of dimeric alpha-amylase inhibitor genes was evaluated by specific SNP markers. A total of 244 dimeric alpha-amylase inhibitor genes were obtained from 13 accessions in 10 populations. Seventy-five polymorphic positions and 74 haplotypes were defined by sequence analysis. Sixteen out of the 75 SNP markers were designed to detect SNP variations in wild emmer wheat accessions from different populations in Israel. The proportion of polymorphic loci P (5%), the expected heterozygosity He, and Shannon's information index in the 16 populations were 0.887, 0.404, and 0.589, respectively. The populations of wild emmer wheat showed great diversity in gene loci both between and within populations. Based on the SNP marker data, the genetic distance of pair-wise comparisons of the 16 populations displayed a sharp genetic differentiation over long geographic distances. The values of P, He, and Shannon's information index were negatively correlated with three climatic moisture factors, whereas the same values were positively correlated by Spearman rank correlation coefficients' analysis with some of the other ecological factors.
The populations of wild emmer wheat showed a wide range of diversity in dimeric alpha-amylase inhibitors, both between and within populations. We suggested that SNP markers are useful for the estimation of genetic diversity of functional genes in wild emmer wheat. These results show significant correlations between SNPs in the alpha-amylase inhibitor genes and ecological factors affecting diversity. Ecological factors, singly or in combination, explained a significant proportion of the variations in the SNPs, and the SNPs could be classified into several categories as ecogeographical predictors. It was suggested that the SNPs in the alpha-amylase inhibitor genes have been subjected to natural selection, and ecological factors had an important evolutionary influence on gene differentiation at specific loci.