Genetic analysis of tolerance to Boron toxicity in the legume Medicago truncatula

Background: Medicago truncatula Gaertn. (barrel medic) is cultivated as a pasture legume for its high protein content and ability to improve soils through nitrogen fixation. Toxic concentrations of the micronutrient Boron (B) in agricultural soils hamper the production of cereal and leguminous crops. In cereals, the genetic analysis of B tolerance has led to the development of molecular selection tools to introgress and maintain the B tolerance trait in breeding lines. There is a comparable need for selection tools in legumes that grow on these toxic soils, often in rotation with cereals. Results: Genetic variation for B tolerance in Medicago truncatula was utilised to generate two F2 populations from crosses between tolerant and intolerant parents. Phenotyping under B stress revealed a close correlation between B tolerance and biomass production and a segregation ratio explained by a single dominant locus. M. truncatula homologues of the Arabidopsis major intrinsic protein (MIP) gene AtNIP5;1 and the efflux-type transporter gene AtBOR1, both known for B transport, were identified and nearby molecular markers screened across F2 lines to verify linkage with the B-tolerant phenotype. Most (95%) of the phenotypic variation could be explained by the SSR markers h2_6e22a and h2_21b19a, which flank a cluster of five predicted MIP genes on chromosome 4. Three CAPS markers (MtBtol-1,-2,-3) were developed to dissect the region further. Expression analysis of the five predicted MIPs indicated that only MtNIP3 was expressed when leaf tissue and roots were assessed. MtNIP3 showed low and equal expression in the roots of tolerant and intolerant lines but a 4-fold higher expression level in the leaves of B-tolerant cultivars. The expression profile correlates closely with the B concentration measured in the leaves and roots of tolerant and intolerant plants. Whereas no significant difference in B concentration exists between roots of tolerant and intolerant plants, the B concentration in the leaves of tolerant plants is less than half that of intolerant plants, which further supports MtNIP3 as the best candidate for the tolerance trait-defining gene in Medicago truncatula. Conclusion: The close linkage of the MtNIP3 locus to B toxicity tolerance provides a source of molecular selection tools to pasture breeding programs. The economical importance of the locus warrants further investigation of the individual members of the MIP gene cluster in other pasture and in grain legumes.Paul Bogacki, David M Peck, Ramakrishnan M Nair, Jake Howie and Klaus H Oldac

Improving the Feeding Value of Dryland Lucerne in Australia

Lucerne (Medicago sativa L.) is the most widely grown perennial legume species in southern Australia. Within Australian farming systems it plays an important role in the provision of high-quality feed for livestock, nitrogen fixation and dewatering soils to reduce watertable recharge and dryland salinity (Cocks 2001). The majority of lucerne varieties have been developed for the areas with high rainfall or supplementary irrigation. The new challenge is to develop lucerne cultivars specifically for dryland mixed farming systems in temperate and mediterranean climate zones (Humphries and Auricht, 2001). Persistence in these environments and feeding value to sheep are critical selection traits. In this paper we compare nutritive traits of 35 commercial and experimental accessions of lucerne, sampled during the vegetative phase, and test the hypothesis that there will be significant differences between the accessions for in vitro dry matter digestibility (DMD), crude protein (CP), acid detergent fibre (ADF), neutral detergent fibre (NDF) and hemicellulose

Genetic analysis of tolerance to the root lesion nematode Pratylenchus neglectus in the legume Medicago littoralis

BACKGROUND: The nematode Pratylenchus neglectus has a wide host range and is able to feed on the root systems of cereals, oilseeds, grain and pasture legumes. Under the Mediterranean low rainfall environments of Australia, annual Medicago pasture legumes are used in rotation with cereals to fix atmospheric nitrogen and improve soil parameters. Considerable efforts are being made in breeding programs to improve resistance and tolerance to Pratylenchus neglectus in the major crops wheat and barley, which makes it vital to develop appropriate selection tools in medics. RESULTS: A strong source of tolerance to root damage by the root lesion nematode (RLN) Pratylenchus neglectus had previously been identified in line RH-1 (strand medic, M. littoralis). Using RH-1, we have developed a single seed descent (SSD) population of 138 lines by crossing it to the intolerant cultivar Herald. After inoculation, RLN-associated root damage clearly segregated in the population. Genetic analysis was performed by constructing a genetic map using simple sequence repeat (SSR) and gene-based SNP markers. A highly significant quantitative trait locus (QTL), QPnTolMl.1, was identified explaining 49% of the phenotypic variation in the SSD population. All SSRs and gene-based markers in the QTL region were derived from chromosome 1 of the sequenced genome of the closely related species M. truncatula. Gene-based markers were validated in advanced breeding lines derived from the RH-1 parent and also a second RLN tolerance source, RH-2 (M. truncatula ssp. tricycla). Comparative analysis to sequenced legume genomes showed that the physical QTL interval exists as a synteny block in Lotus japonicus, common bean, soybean and chickpea. Furthermore, using the sequenced genome information of M. truncatula, the QTL interval contains 55 genes out of which five are discussed as potential candidate genes responsible for the mapped tolerance. CONCLUSION: The closely linked set of SNP-based PCR markers is directly applicable to select for two different sources of RLN tolerance in breeding programs. Moreover, genome sequence information has allowed proposing candidate genes for further functional analysis and nominates QPnTolMl.1 as a target locus for RLN tolerance in economically important grain legumes, e.g. chickpea.Klaus H Oldach, David M Peck, Ramakrishnan M Nair, Maria Sokolova, John Harris, Paul Bogacki, and Ross Ballar

Classification and identification of Pfiesteria and Pfiesteria-like species.

Dinoflagellates can be classified both botanically and zoologically; however, they are typically put in the botanical division Pyrrhophyta. As a group they appear most related to the protistan ciliates and apicomplexans at the ultrastructure level. Within the Pyrrhophyta are both unarmored and armored forms of the dominant, motile flagellated stage. Unarmored dinoflagellates do not have thecal or wall plates arranged in specific series, whereas armored species have plates that vary in thickness but are specific in number and arrangement. In armored dinoflagellates, the plate pattern and tabulation is a diagnostic character at the family, subfamily, and even genus levels. In most cases, the molecular characterization of dinoflagellates confirms the taxonomy on the basis of external morphology; this has been demonstrated for several groups. Together, both genetic and morphological criteria are becoming increasingly important for the characterization, separation, and identification of dinoflagellates species. Pfiesteria and Pfiesteria-like species are thinly armored forms with motile dinospore stages characterized by their distinct plate formulae. Pfiesteria piscicida is the best-known member of the genus; however, there is at least one other species. Other genetically and morphologically related genera, now grouped under the common names of "Lucy," "Shepherd's crook," and cryptoperidiniopsoid, are being studied and described in separate works. All these other heterotrophic dinoflagellate groups, many of which are thought to be benign, co-occur in estuarine waters where Pfiesteria has been found

Overexpression of defense response genes in transgenic wheat enhances resistance to Fusarium head blight

Fusarium head blight (FHB) of wheat, caused by Fusarium graminearum and other Fusarium species, is a major disease problem for wheat production worldwide. To combat this problem, large-scale breeding efforts have been established. Although progress has been made through standard breeding approaches, the level of resistance attained is insufficient to withstand epidemic conditions. Genetic engineering provides an alternative approach to enhance the level of resistance. Many defense response genes are induced in wheat during F. graminearum infection and may play a role in reducing FHB. The objectives of this study were (1) to develop transgenic wheat overexpressing the defense response genes α-1-purothionin, thaumatin-like protein 1 (tlp-1), and β-1,3-glucanase; and (2) to test the resultant transgenic wheat lines against F. graminearum infection under greenhouse and field conditions. Using the wheat cultivar Bobwhite, we developed one, two, and four lines carrying the α-1-purothionin, tlp-1, and β-1,3-glucanase transgenes, respectively, that had statistically significant reductions in FHB severity in greenhouse evaluations. We tested these seven transgenic lines under field conditions for percent FHB disease severity, deoxynivalenol (DON) mycotoxin accumulation, and percent visually scabby kernels (VSK). Six of the seven lines differed from the nontransgenic parental Bobwhite line for at least one of the disease traits. A β-1,3-glucanase transgenic line had enhanced resistance, showing lower FHB severity, DON concentration, and percent VSK compared to Bobwhite. Taken together, the results showed that overexpression of defense response genes in wheat could enhance the FHB resistance in both greenhouse and field conditions

Transriptomics, metabolomics and proteomics: Integration of latest technologies for improving future wheat productivity

No abstract availabl

Characterization of resistance to pratylenchus thornei (Nematoda) in wheat (Triticum aestivum): attraction, penetration, motility, and reproduction

Lines from a cross between two wheat (Triticum aestivum) cultivars with contrasting resistance phenotypes to Pratylenchus thornei (Nematoda) were investigated to determine the stage at which resistance occurs. Host resistance was examined at nematode attraction to and penetration of roots and nematode motility, maturation, and reproduction within roots. There was no significant difference in the rate at which P. thornei was attracted toward or penetrated resistant or susceptible roots. However, suppression of migration, juvenile maturation, and reproduction in and near resistant roots was evident, suggesting that resistance acts post penetration. No preferential root penetration zone was observed in contrast to other studies. The inhibitory compounds from resistant wheat plants appeared to be constitutively expressed and water soluble because nematode migration was suppressed in roots and root exudates of unchallenged seedlings. The effects of these compounds were reversible and affected P. thornei but not P. neglectus. Apart from migration, nematode multiplication was greatly inhibited by resistance because only a few juveniles (10%) developed past stage three in roots of resistant compared with susceptible plants. Earlier in the life cycle, egg deposition and hatch of P. thornei were also significantly reduced in resistant roots and root exudates, suggesting the presence of hatching inhibitors.Katherine Joanne Linsell, Ian Timothy Riley, Kerrie Ann Davies, and Klaus Herbert Oldac

New insights into the infection process of Rhynchosporium secalis in barley using GFP

Through the use of a Rhynchosporium secalis isolate transformed with the green fluorescent protein gene and LASER scanning confocal microscopy (LSCM), fungal development during the R. secalis/barley interaction was analysed. Each infection stage was investigated from 0.5h to 14 days post-inoculation (p.i.) with extensive sampling within the first 48 h p.i. Early germination events were observed that had not been previously described. A specific time of germination was noted, with germ tube formation appearing as early as 1h p.i. Conidia were observed within anticlinal grooves of epidermal cells and the formation of bubbles within these pectin-rich regions was observed within 24h p.i. The study reports R. secalis pectinase production and suggests degradation of these pectin-rich regions. Reactive oxygen species were present during early penetration, 3h p.i. and co-localised with fungal development. LSCM allowed the visualisation of fungal growth deep within tissues at the later stage of the infection.Katherine J. Linsell, Felicity J. Keiper, Angus Forgan and Klaus H. Oldachhttp://www.elsevier.com/wps/find/journaldescription.cws_home/622835/description#descriptio

Analysis of high pI alpha-Amy-1 gene family members expressed in late maturity alpha-amylase in wheat (Triticum aestivum L.)

Late maturity α-amylase (LMA) is a genetic defect involving the synthesis of high pI isozymes of α-amylase encoded by α-Amy-1 genes during the later stages of grain development. The aims of this investigation were to determine both the number of expressed α-Amy-1 genes and their relative transcript abundance. Sub-cloning and sequencing of expressed high pI α-amylase genes in developing wheat seeds revealed three insertion/deletion patterns in the 3' untranslated region and numerous single nucleotide polymorphisms at the 3' end of α-Amy-1. The genetic variations defined 36 α-Amy-1 gene sequences that were expressed on the onset of LMA in doubled haploid progenies (SpM25, SpM52 and SpM127) derived from the cross Spica (LMA)/Maringa (non-LMA). Five isoelectric point groups were predicted based on the translated partial coding sequences. The potential application of quantitative real-time RT-PCR in screening wheat genotypes for LMA is discussed.Cong-Rong Cheng, Klaus Oldach, Kolumbina Mrva, Daryl Mare