Tissue-specific gene expression in soybean (Glycine max) detected by cDNA microarray analysis

We have constructed cDNA microarrays for soybean (Glycine max L. Merrill), containing approximately 4,100 Unigene ESTs derived from axenic roots, to evaluate their application and utility for functional genomics of organ differentiation in legumes. We assessed microarray technology by conducting studies to evaluate the accuracy of microarray data and have found them to be both reliable and reproducible in repeat hybridisations. Several ESTs showed high levels (>50 fold) of differential expression in either root or shoot tissue of soybean. A small number of physiologically interesting, and differentially expressed sequences found by microarray analysis were verified by both quantitative real-time RT-PCR and Northern blot analysis. There was a linear correlation (r(2) = 0.99, over 5 orders of magnitude) between microarray and quantitative real-time RT-PCR data. Microarray analysis of soybean has enormous potential not only for the discovery of new genes involved in tissue differentiation and function, but also to study the expression of previously characterised genes, gene networks and gene interactions in wild-type, mutant or transgenic; plants.

DAF yields a cloned marker linked to the soybean (Glycine max) supernodulation nts-1 locus

We report a further characterization of the genomic region containing the soybean supernodulation gene NTS-1. We performed a search for new markers linked to NTS-1 by combining DNA amplification fingerprinting (DAF) and bulked segregant analysis (BSA). The search resulted in one cloned polymorphism (B44-456) linked in trans, 8.5cM from the locus. Southern hybridization showed duplication of the B44-456 sequence in the soybean genome. Additionally, a DNA database search revealed one Arabidopsis thaliana genomic clone from chromosome I possessing 62% homology to the B44-456 marker. A relatively low number of polymorphisms were identified by several PCR marker technologies for this soybean genomic region, providing an additional support for its highly conserved and/or duplicated organization.

Fast neutron mutagenesis of soybean (Glycine soja L.) produces a supernodulating mutant containing a large deletion in linkage group H

We describe for the first time the application of fast neutron mutagenesis to the genetic dissection of root nodulation in legumes. We demonstrate the utility of chromosomal deletion mutations through production of a soybean supernodulation mutant FN37 that lacks the internal autoregulation of nodulation mechanism. After inoculation with microsymbiont Bradyrhizobium japonicum, FN37 forms at least 10 times more nodules than the wild type G. soja parent and has a phenotype identical to that of chemically induced allelic mutants nts382 and nts1007 (NTS-1 locus). Reciprocal grafting of shoots and roots confirmed systemic shoot control of the FN37 nodulation phenotype. RFLP/PCR marker pUTG132a and AFLP marker UQC-IS1 which are tightly linked to NTS-1 allowed the isolation of BAC contigs delineating both ends of the deletion. The genetic/physical distance ratio in the NTS-1 region is 279 kb/cM. The deletion is estimated to be about 460 kb based on the absence of markers and bacterial artificial chromosomes (BAC) ends as well as genetic and physical mapping. Deletion break points were determined physically and placed within flanking BAC contigs.

Quantitative trait loci for agronomic and seed quality traits in an F-2 and F-4 : 6 soybean population

Molecular breeding is becoming more practical as better technology emerges. The use of molecular markers in plant breeding for indirect selection of important traits can favorably impact breeding efficiency. The purpose of this research is to identify quantitative trait loci (QTL) on molecular linkage groups (MLG) which are associated with seed protein concentration, seed oil concentration, seed size, plant height, lodging, and maturity, in a population from a cross between the soybean cultivars 'Essex' and 'Williams.' DNA was extracted from F-2 generation soybean leaves and amplified via polymerase chain reaction (PCR) using simple sequence repeat (SSR) markers. Markers that were polymorphic between the parents were analyzed against phenotypic trait data from the F-2 and F-4:6 generation. For the F-2 population, significant additive QTL were Satt540 (MLG M, maturity, r(2)=0.11; height, r(2)=0.04, seed size, r(2)=0.061, Satt373 (MLG L, seed size, r(2)=0.04; height, r(2)=0.14), Satt50 (MLG A1, maturity r(2)=0.07), Satt14 (MLG D2, oil, r(2)=0.05), and Satt251 (protein r(2)=0.03, oil, r(2)=0.04). Significant dominant QTL for the F-2 population were Satt540 (MLG M, height, r(2)=0.04; seed size, r(2)=0.06) and Satt14 (MLG D2, oil, r(2)=0.05). In the F-4:6 generation significant additive QTL were Satt239 (MLG I, height, r(2)=0.02 at Knoxville, TN and r(2)=0.03 at Springfield, TN), Satt14 (MLG D2, seed size, r(2)=0.14 at Knoxville, TN), Satt373 (MLG L, protein, r(2)=0.04 at Knoxville, TN) and Satt251 (MLG B I, lodging r(2)=0.04 at Springfield, TN). Averaged over both environments in the F-4:6 generation, significant additive QTL were identified as Satt251 (MLG B 1, protein, r(2)=0.03), and Satt239 (MLG 1, height, r(2)=0.03). The results found in this study indicate that selections based solely on these QTL would produce limited gains (based on low r(2) values). Few QTL were detected to be stable across environments. Further research to identify stable QTL over environments is needed to make marker-assisted approaches more widely adopted by soybean breeders.

Lack of mycorrhizal autoregulation and phytohormonal changes in the supernodulating soybean mutant nts1007

Autoregulatory mechanisms have been reported in the rhizobial and the mycorrhizal symbiosis. Autoregulation means that already existing nodules or an existing root colonization by an arbuscular mycorrhizal fungus systemically suppress subsequent nodule formation/root colonization in other parts of the root system. Mutants of some legumes lost their ability to autoregulate the nodule number and thus display a supernodulating phenotype. On studying the effect of pre-inoculation of one side of a split-root system with an arbuscular mycorrhizal fungus on subsequent mycorrhization in the second side of the split-root system of a wild-type soybean (Glycine max L.) cv. Bragg and its supernodulating mutant nts1007, we observed a clear suppressional effect in the wild-type, whereas further root colonization in the split-root system of the mutant nts1007 was not suppressed. These data strongly indicate that the mechanisms involved in supernodulation also affect mycorrhization and support the hypothesis that the autoregulation in the rhizobial and the mycorrhizal symbiosis is controlled in a similar manner. The accumulation patterns of the plant hormones IAA, ABA and Jasmonic acid (JA) in non-inoculated control plants and split-root systems of inoculated plants with one mycorrhizal side of the split-root system and one non-mycorrhizal side, indicate an involvement of IAA in the autoregulation of mycorrhization. Mycorrhizal colonization of soybeans also resulted in a strong induction of ABA and JA levels, but on the basis of our data the role of these two phytohormones in mycorrhizal autoregulation is questionable.

Adaptive responses of wild mungbean (Vigna radiata ssp sublobata) to photo-thermal environment. I. Phenology

The phenology of 11 diverse accessions of wild mungbean was observed under natural and artificial photoperiod - temperature conditions, in order to examine whether genotypic differences might be attributed to adaptive responses to photo-thermal conditions. There was large variation in phenological response among accessions and across environments, much of which was due to differences in the duration of the pre-flowering phase. Accessions that flowered earlier tended to flower for longer, apart from 2 earlier flowering, inland Australian lines that were also earlier maturing. The patterns of response in time from sowing to flowering over environment were consistent with quantitative short-day photoperiodic adaptation, a conclusion supported by the effects of artificial day-length extension and by 'goodness of fit' of the observed responses to standard models relating rate of development to photoperiod and temperature. The fitted models indicated that rate of development towards flowering was hastened by warmer temperatures, and delayed by longer day lengths, with differential sensitivity between accessions to both factors. The models also suggested that photoperiod was more important for accessions collected closer to the equator, which were generally later flowering as a consequence. Conversely, temperature was relatively more important in lines from higher latitudes. Modelling also suggested that the period from first flowering to maturity was sensitive to photoperiod and temperature. Again, longer days appeared to prolong growth and delay maturity. However, cooler temperatures accelerated rather than slowed maturity, by suppressing further vegetative growth. The variation observed indicated that there is considerable scope for using the wild population to broaden the adaptation of cultivated mungbean. In particular, the unusual response of a late-flowering, photoperiod-insensitive accession warrants further study to establish whether the wild population contains a unique 'long juvenile' trait analogous to that being used for improving phenological adaptation in soybean.

Responses of sugarcane, maize, and soybean to phosphorus and vesicular-arbuscular mycorrhizal fungi

The presence of vesicular-arbuscular mycorrhizal (VAM) fungi in long-term cane-growing fields associated with yield decline led to the supposition that VAM fungi may be responsible for the poor yields. A glasshouse trial was established to test the effectiveness of a species of VAM fungi, Glomus clarum, extracted from one of these North Queensland fields on the growth of sugarcane (Saccharum interspecific hybrid), maize (Zea mays), and soybean (Glycine max) for 6 phosphorus (P) rates (0, 2.7, 8.2, 25, 74, 222 mg/kg). For maize and soybean plants that received VAM (+ VAM), root colonisation was associated with enhanced P uptake, improved dry weight (DW) production, and higher index tissue-P concentrations than those without VAM (-VAM). By comparing DW responses of maize and soybean for different P rates, savings in fertiliser P of up to 160 and 213 kg/ha, respectively, were realised. Sugarcane plants were generally less responsive. Apart from a 30% DW increase with VAM when 2.7 mg P/kg was added, DW of +VAM plants was equivalent to, or worse than in the case of 222 mg P/kg, DW of -VAM plants. For all 3 host species, colonisation was least at the highest P application, presumably from excessive P within the plant tissue. Critical P concentrations for the 3 host species were below those reported elsewhere, and for soybean and sugarcane, the critical concentration for +VAM plants was lower than that of -VAM plants. There are 3 implications that arise from this study. First, VAM fungi present in cane-growing soils can promote the growth of maize and soybean, which are potential rotation crops, over a range of P levels. Second, the mycorrhizal strain taken from this site did not generally contribute to a yield decline in sugarcane plants. Third, application of P fertiliser is not necessary for sugarcane when acid-extractable P is