Isolation and characterization of the rye Waxy gene.

Complete genomic and cDNA sequences of the Waxy gene encoding granule-bound starch synthase I (GBSSI) were isolated from the rye genome and characterized. The full-length rye Waxy genomic DNA and cDNA are 2767 bp and 1815 bp, respectively. The genomic sequence has 11 exons interrupted by 10 introns. The rye Waxy gene is GC-rich, with a higher GC frequency in the coding region, especially in the third position of the codons. Exon regions of the rye Waxy gene are more conserved than intron regions when compared with the homologous sequences of other cereals. The mature rye GBSSI proteins share more than 95% sequence identity with their homologs in wheat and barley. A phylogenetic tree based on sequence comparisons of available plant GBSSI proteins shows the evolutionary relationship among Waxy genes from rye and other plant genomes. The identification of the rye Waxy gene will enable the manipulation of starch metabolism in rye and triticale.

[Integrating genetic and gene expression data: methods and applications of eQTL mapping].

The availability of high-throughput genotyping technologies and microarray assays has allowed researchers to investigate genetic variations that influence levels of gene expression. Expression Quantitative Trait Locus (eQTL) mapping methods have been used to identify the genetic basis of gene expression. Similar to traditional QTL studies, the main goal of eQTL is to identify the genomic locations to which the expression traits are linked. Although microarrays provide the expression data of thousands of transcripts, standard QTL mapping methods, which are able to handle at most tens of traits, cannot be applied directly. As a result, it is necessary to consider the statistical principles involved in the design and analysis of these experiments. In this paper, we reviewed individual selection, experimental design of microarray, normalization of gene expression data, mapping methods, and explaining of results and proposed potential methodological problems for such analyses. Finally, we discussed the applications of this integrative genomic approach to estimate heritability of transcripts, identify candidate genes, construct gene networks, and understand interactions between genes, genes and environments.

[Comparative analysis of genetic diversity revealed by genomic-sSR, EST-SSR and pedigree in wheat (Triticum asetivum L.)].

Two SSR molecular markers, genomic-SSR and EST (expressed sequence tagged)-SSR, were used to measure the genetic diversity among 18 accessions of common wheat with known pedigrees, which were collected from winter wheat production region in Northern China. In addtion, the genetic diversity revealed by pedigree, EST-SSR and genomic-SSR was also compared. The results showed that the average number of alleles per genomic-SSR locus is 3.34, which is higher than that of EST-SSR (2.31), indicating that genomic-SSR is more polymorphic than EST-SSR. Genomic-SSR and EST-SSR were used to calculate the genetic distance (GD) in different materials. The mean GD value of EST-SSR for the 18 wheat genotypes is 0.3996,which is lower than that of genomic-SSR (0.5458). At the locus level, the mean GD calculated using pedigree information is higher (0. 9716) than that of genomic-SSR and EST-SSR markers (0.5458 and 0.3996). Therefore, although polymorphism of EST-SSR is low as compared to genomic SSR,it provides a more accurate evaluation of genetic relationship, especially when accessions are very closely related in pedigree. The strategy for improving the genetic diversity of wheat was also discussed.

[Genetic diversity of D-genome revealed by SSR markers in synthesized hexaploid wheat introduced from CIMMYT].

Simple sequence repeat (SSR) molecular marker was used to measure the genetic diversity of D-genome in 26 synthesized hexaploid wheat (AABBDD) introduced from CIMMYT. Twenty-three D-genome specific SSR primers were selected for PCR amplification, among which 22 primers can detect polymorphism. A total of 92 alleles were identified at 23 loci using the above SSR primers, with an average of 4 alleles per locus. The 92 alleles were used to calculate Nei's similarity index (GS) and the genetic distance (GD). It was also found that the mean genetic distance between 26 synthesized hexaploid wheat was 0.4955, which was obviously high. From the above results, it can be indicated that the genetic variation of D-genome in synthesized hexaploid wheat was abundant and could be used to improve the genetic diversity in wheat breeding. Interestingly, synthesized hexaploid wheat 17 and 18 shared the same D-genome donor, but three of 23 detected SSR loci were polymorphic between the two materials. Therefore, during the period of allopolyploidization, there was genetic differentiation in repeat region of donor genome.

[Genetic diversity revealed by ISSR molecular marker in common wheat, spelt, compactum and progeny of recurrent selection].

It is important to estimate the genetic diversity between the parents for improving the heterosis of hybrid wheat. In this study, ISSR(inter-simple sequence repeat) marker was used to measure the genetic diversity within and among common wheat (Triticum aestivum L.), spelt (Triticum spelta L.), compactum (Triticum compactum Host.) and progeny of foreign wheat-based recurrent selection, and the possibility of establishing the new heterotic group was also assessed. Forty seven genotypes were used for ISSR analysis, which included 14 common wheat, 10 spelt wheat, 11 compactum and 12 progeny of recurrent selection. Eleven of 33 ISSR primers that can produce distinguishable bands were selected for PCR amplification. A total of 238 bands were amplified, among which 207 (87%) bands were polymorphic. The polymorphic bands amplified by each primer ranged from 11 to 38, with an averaged of 18.8. The percentage of polymorphic band (80.3%) in common wheat was higher than that in progeny of recurrent selection (78.7%), spelt (75.0%) and compactum (74.9%). The 238 polymorphic products were used to calculate Nei's similarity index (GS) and the genetic distance (GD). It was found that the mean genetic distance between different wheat types (0.3115-0.3442) was obviously higher than that within common wheat (0.2743), spelt (0.2351), compactum (0.2622). In addition, progeny of recurrent selection also showed much higher genetic distance with other three wheat types (0.3217, 0.3256, 0.3198). The cluster analysis was performed based on the genetic distance (GD) matrix by using UPGMA method. Common wheat, spelt, compactum and progeny of recurrent selection were classified into four different groups. In this study, ISSR marker was firstly used to assess genetic diversity among common wheat, spelt, compactum and progeny of recurrent selection, and can differentiate the wheat cultivars (lines) that selected from the same cross combination. It was concluded that spelt, compactum and progeny of recurrent selection can be used to diversify the genetic basis for hybrid wheat breeding and improve heterosis. It is possible to establish the wheat heterotic group by ISSR marker.

Relationship between differences of gene expression in early developing seeds of hybrid versus parents and heterosis in wheat.

In order to understand molecular basis of heterosis, mRNA differential display was used to analyze the differences in gene expression between seeds of 18 reciprocal hybrids and their 6 parents at 6th day after pollination. The relationship between gene expression patterns and heterosis was determined. Only bands that can be repeated in duplicate PCR were used for analysis so as to reduce false positive bands. Among the total of 2,025 bands displayed, 1,386 bands (68.43%) were reproducible. Eight patterns (fifteen kinds) of gene expression were observed, which include: (1) bands occurring in only one parent (two kinds); (2) bands observed in both hybrids and one parent (two kinds); (3) bands detected in parents and one hybrid (two kinds); (4) bands displayed in only one hybrid (two kinds); (5) bands revealed in one hybrid and its corresponding female (two kinds) or male (two kinds) parent; (6) bands visualized in only both hybrids (one kind); (7) bands occurring in only parents (one kind); (8) bands observed in parents and both hybrids (one kind). Our results indicate that differences of gene expression between hybrids and their parents are very obvious. The percentages of bands observed in only both hybrids and in only both parents are lower. The analysis shows that bands observed in parents and both hybrids are not correlated with all nine agronomic traits, which indicates differentially expressed genes are mainly responsible for the observed heterosis. At least one pattern of differential gene expression is significantly correlated with nine agronomic traits. Silenced bands in hybrid (which include bands occurring in only one parent, bands detected in both parents and one hybrid and bands in only parents) and bands present in one hybrid and its corresponding female or male parent are likely to play important roles in heterosis. These results suggest that early seed development could be closely related to heterosis.

A wheat lipid transfer protein 3 could enhance the basal thermotolerance and oxidative stress resistance of Arabidopsis.

Wheat (Triticum aestivum L.) is one of the major grain crops, and heat stress adversely affects wheat production in many regions of the world. Previously, we found a heat-responsive gene named Lipid Transfer Protein 3 (TaLTP3) in wheat. TaLTP3 was deduced to be regulated by cold, ABA, MeJA, Auxin and oxidative stress according to cis-acting motifs in its promoter sequences. In this study, we show that TaLTP3 is responsive to prolonged water deficit, salt or ABA treatment in wheat seedlings. Also, TaLTP3 accumulation was observed after the plant suffered from heat stress both at the seedling and the grain-filling stages. TaLTP3 protein was localized in the cell membrane and cytoplasm of tobacco epidermal cells. Overexpression of TaLTP3 in yeast imparted tolerance to heat stress compared to cells expressing the vector alone. Most importantly, transgenic Arabidopsis plants engineered to overexpress TaLTP3 showed higher thermotolerance than control plants at the seedling stage. Further investigation indicated that transgenic lines decreased H2O2 accumulation and membrane injury under heat stress. Taken together, our results demonstrate that TaLTP3 confers heat stress tolerance possibly through reactive oxygen species (ROS) scavenging.