Two classes of isocitrate lyase genes are expressed during late embryogeny and postgermination in Brassica napus L.

Deletion of the SLT2 gene of Saccharomyces cerevisiae, which codes for a homologue of MAP (mitogen-activated) protein kinases, causes an autolytic lethal phenotype in cells grown at 37° C. The gene encodes domains characteristic of protein kinases, which include a lysine (at position 54) that lies 19 residues from a glycine-rich cluster, considered to be the putative ATP binding site. The ability of three mutant alleles of SLT2 generated by site-directed mutagenesis, namely E54 (glutamic acid), R54 (arginine) and F54 (phenylalanine), to complement slt2 mutants was tested. All three failed to complement the autolytic phenotype and were unable to restore growth and viability of cells. A strain obtained by transplacement of slt2-F54 also behaved as a thermosensitive autolytic mutant. By immunoprecipitation with polyclonal antibodies raised against Slt2 protein expressed in Escherichia coli, it was possible to confirm that alteration of the lysine-54 residue did not affect the stability of the protein, thus allowing us to conclude that activity of the Slt2 protein kinase is critically required for growth and morphogenesis of S. cerevisiae at 37° C. A significant fraction of the mutant cell population lysed at 24° C and the cells displayed a characteristic alteration of the surface consisting of a typical depression in an area of the cell wall. At 37° C, the cell surface was clearly disorganized.     

Identification and mapping of two brown planthopper resistance genes in rice

The brown planthopper (BPH) is one of the most serious insect pests of rice. In this study, we conducted a molecular marker-based genetic analysis of the BPH resistance of ’B5’, a highly resistant line that derived its resistant genes from the wild rice Oryza officinalis. Insect resistance was evaluated using 250 F₃ families from a cross between ’B5’ and ’Minghui 63’, based on which the resistance of each F₂ plant was inferred. Two bulks were made by mixing, respectively, DNA samples from highly resistant plants and highly susceptible plants selected from the F₂ population. The bulks were surveyed for restriction fragment length polymorphism using probes representing all 12 chromosomes at regular intervals. The survey revealed two genomic regions on chromosome 3 and chromosome 4 respectively that contained genes for BPH resistance. The existence of the two loci were further assessed by QTL (quantitative trait locus) analysis, which resolved these two loci to a 14.3-cM interval on chromosome 3 and a 0.4-cM interval on chromosome 4. Comparison of the chromosomal locations and reactions to BPH biotypes indicated that these two genes are different from at least nine of the ten previously identified BPH resistance genes. Both of the genes had large effects on BPH resistance and the two loci acted essentially independent of each other in determining t he resistance. These two genes may be a useful BPH resistance resource for rice breeding programs. Received: 6 March 2000 / Accepted: 28 July 2000     

Molecular characterisation and genetic mapping of candidate genes for qualitative disease resistance in perennial ryegrass (Lolium perenne L.)

Background  

Qualitative pathogen resistance in both dicotyledenous and monocotyledonous plants has been attributed to the action of resistance (R) genes, including those encoding nucleotide binding site – leucine rich repeat (NBS-LRR) proteins and receptor-like kinase enzymes. This study describes the large-scale isolation and characterisation of candidate R genes from perennial ryegrass. The analysis was based on the availability of an expressed sequence tag (EST) resource and a functionally-integrated bioinformatics database.     

Transcriptional profiles of primary metabolism and signal transduction-related genes in response to water stress in field-grown sunflower genotypes using a thematic cDNA microarray

A sunflower cDNA microarray containing about 800 clones covering major metabolic and signal transduction pathways was used to study gene expression profiles in leaves and embryos of drought-tolerant and -sensitive genotypes subjected to water-deficit stress under field conditions. Using two-step ANOVA normalization and analysis models, we identified 409 differentially expressed genes among genotypes, water treatment and organs. The majority of the cDNA clones differentially expressed under water stress was found to display opposite gene expression profiles in drought-tolerant genotype compared to drought-sensitive genotype. These dissimilarities suggest that the difference between tolerant and non-tolerant plants seems to be associated with changes in qualitative but not quantitative mRNA expression. Comparing leaves and embryos, 82 cDNA clones showing organ-specific variation in gene expression levels were identified in response to water stress across genotypes. Genes related to amino acids and carbohydrates metabolisms, and signal transduction were induced in embryos and repressed in leaves; suggesting that vegetative and reproductive organs respond differentially to water stress. Adaptive mechanisms controlling water deficit tolerance are proposed and discussed.     

Identification and linkage mapping of complementary recessive genes causing hybrid breakdown in an intraspecific rice cross

One outcome of hybrid breakdown is poor growth, which we observed as a reduction in the number of panicles per plant and in culm length in an F₂ population derived from a cross between the genetically divergent rice (Oryza sativa L.) cultivars ‘Sasanishiki’ (japonica) and ‘Habataki’ (indica). Quantitative trait locus (QTL) analysis of the two traits and two-way ANOVA of the detected QTLs suggested that the poor growth was due mainly to an epistatic interaction between genes at QTLs located on chromosomes 2 and 11. The poor growth was likely to result when a plant was homozygous for the ‘Habataki’ allele at the QTL on chromosome 2 and homozygous for the ‘Sasanishiki’ allele at the QTL on chromosome 11. The results suggest that the poor growth found in the F₂ population was due to hybrid breakdown of a set of complementary genes. To test this hypothesis and determine the precise chromosomal location of the genes causing the hybrid breakdown, we performed genetic analyses using a chromosome segment substitution line, in which a part of chromosome 2 from ‘Habataki’ was substituted into the genetic background of ‘Sasanishiki’. The segregation patterns of poor growth in plants suggested that both of the genes underlying the hybrid breakdown were recessive. The gene on chromosome 2, designated hybrid breakdown 2 (hbd2), was mapped between simple sequence repeat markers RM3515 and RM3730. The gene on chromosome 11, hbd3, was mapped between RM5824 and RM1341. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Identification of candidate genes in Arabidopsis and Populus cell wall biosynthesis using text-mining, co-expression network analysis and comparative genomics

Populus is an important bioenergy crop for bioethanol production. A greater understanding of cell wall biosynthesis processes is critical in reducing biomass recalcitrance, a major hindrance in efficient generation of biofuels from lignocellulosic biomass. Here, we report the identification of candidate cell wall biosynthesis genes through the development and application of a novel bioinformatics pipeline. As a first step, via text-mining of PubMed publications, we obtained 121 Arabidopsis genes that had the experimental evidence supporting their involvement in cell wall biosynthesis or remodeling. The 121 genes were then used as bait genes to query an Arabidopsis co-expression database, and additional genes were identified as neighbors of the bait genes in the network, increasing the number of genes to 548. The 548 Arabidopsis genes were then used to re-query the Arabidopsis co-expression database and re-construct a network that captured additional network neighbors, expanding to a total of 694 genes. The 694 Arabidopsis genes were computationally divided into 22 clusters. Queries of the Populus genome using the Arabidopsis genes revealed 817 Populus orthologs. Functional analysis of gene ontology and tissue-specific gene expression indicated that these Arabidopsis and Populus genes are high likelihood candidates for functional characterization in relation to cell wall biosynthesis.     

Identification of quantitative trait loci and associated candidate genes for low-temperature tolerance in cold-hardy winter wheat

Low-temperature (LT) tolerance is an important economic trait in winter wheat (Triticum aestivum L.) that determines the plants’ ability to cope with below freezing temperatures. Essential elements of the LT tolerance mechanism are associated with the winter growth habit controlled by the vernalization loci (Vrn-1) on the group 5 chromosomes. To identify genomic regions, which in addition to vrn-1 determine the level of LT tolerance in hexaploid wheat, two doubled haploid (DH) mapping populations were produced using parents with winter growth habit (vrn-A1, vrn-B1, and vrn-D1) but showing different LT tolerance levels. A total of 107 DH lines were analyzed by genetic mapping to produce a consensus map of 2,873 cM. The LT tolerance levels for the Norstar (LT₅₀=−20.7°C) × Winter Manitou (LT₅₀=−14.3°C) mapping population ranged from −12.0 to −22.0°C. Single marker analysis and interval mapping of phenotyped lines revealed a major quantitative trait locus (QTL) on chromosome 5A and a weaker QTL on chromosome 1D. The 5A QTL located 46 cM proximal to the vrn-A1 locus explained 40% of the LT tolerance variance. Two C-repeat Binding Factor (CBF) genes expressed during cold acclimation in Norstar were located at the peak of the 5A QTL.     

Identification and characterization of genes induced for anthocyanin synthesis and chlorophyll degradation in regenerated torenia shoots using suppression subtractive hybridization, cDNA microarrays, and RNAi techniques

Anthocyanin synthesis and chlorophyll degradation in regenerated torenia (Torenia fournieri Linden ex Fourn.) shoots induced by osmotic stress with 7% sucrose were examined to identify the genes regulating the underlying molecular mechanism. To achieve this, suppression subtractive hybridization was performed to enrich the cDNAs of genes induced in anthocyanin-synthesizing and chlorophyll-degrading regenerated shoots. The nucleotide sequences of 1,388 random cDNAs were determined, and these were used in the preparation of cDNA microarrays for high-throughput screening. From 1,056 cDNAs analyzed in the microarrays, 116 nonredundant genes were identified, which were up regulated by 7% sucrose to induce anthocyanin synthesis and chlorophyll degradation in regenerated shoots. Of these, eight genes were selected and RNAi transformants prepared, six of which exhibited anthocyanin synthesis inhibition and/or chlorophyll degradation in their leaf discs. Notably, the RNAi transformants of the glucose 6-phosphate/phosphate translocator gene displayed inhibition both of anthocyanin synthesis and chlorophyll degradation in both leaf discs and regenerated shoots. There was also less accumulation of anthocyanin in the petals, and flowering time was shortened. The genes we identified as being up-regulated in the regenerated torenia shoots may help further elucidate the molecular mechanism underlying the induction of anthocyanin synthesis and chlorophyll degradation.