QTL analysis of ergot resistance in sorghum

Sorghum ergot, caused predominantly by Claviceps africana Frederickson, Mantle, de Milliano, is a significant threat to the sorghum industry worldwide. The objectives of this study were firstly, to identify molecular markers linked to ergot resistance and to two pollen traits, pollen quantity (PQ) and pollen viability (PV), and secondly, to assess the relationship between the two pollen traits and ergot resistance in sorghum. A genetic linkage map of sorghum RIL population R931945-2-2 x IS 8525 (resistance source) was constructed using 303 markers including 36 SSR, 117 AFLP™, 148 DArT™ and two morphological trait loci. Composite interval mapping identified nine, five, and four QTL linked to molecular markers for percentage ergot infection (PCERGOT), PQ and PV, respectively, at a LOD >2.0. Co-location/linkage of QTL were identified on four chromosomes while other QTL for the three traits mapped independently, indicating that both pollen and non pollen-based mechanisms of ergot resistance were operating in this sorghum population. Of the nine QTL identified for PCERGOT, five were identified using the overall data set while four were specific to the group data sets defined by temperature and humidity. QTL identified on SBI-02 and SBI-06 were further validated in additional populations. This is the first report of QTL associated with ergot resistance in sorghum. The markers reported herein could be used for marker-assisted selection for this important disease of sorghum.

Identification of QTL for sugar-related traits in a sweet x grain sorghum (Sorghum bicolor L. Moench) recombinant inbred population

QTL for stem sugar-related and other agronomic traits were identified in a converted sweet (R9188) × grain (R9403463-2-1) sorghum population. QTL analyses were conducted using phenotypic data for 11 traits measured in two field experiments and a genetic map comprising 228 SSR and AFLP markers grouped into 16 linkage groups, of which 11 could be assigned to the 10 sorghum chromosomes (SBI-01 to SBI-10). QTL were identified for all traits and were generally co-located to five locations (SBI-01, SBI-03, SBI-05, SBI-06 and SBI-10). QTL alleles from R9188 were detected for increased sucrose content and sugar content on SBI-01, SBI-05 and SBI-06. R9188 also contributed QTL alleles for increased Brix on SBI-05 and SBI-06, and increased sugar content on SBI-03. QTL alleles from R9403463-2-1 were found for increased sucrose content and sucrose yield on SBI-10, and increased glucose content on SBI-07. QTL alleles for increased height, later flowering and greater total dry matter yield were located on SBI-01 of R9403463-2-1, and SBI-06 of R9188. QTL alleles for increased grain yield from both R9403463-2-1 and R9188 were found on SBI-03. As an increase in stem sugars is an important objective in sweet sorghum breeding, the QTL identified in this study could be further investigated for use in marker-assisted selection of sweet sorghum.

Congruence in QTL for adventitious rooting in Pinus elliottii x Pinus caribaea hybrids resolves between and within-species effects

Targeting between-species effects for improvement in synthetic hybrid populations derived from outcrossing parental tree species may be one way to increase the efficacy and predictability of hybrid breeding. We present a comparative analysis of the quantitative trait loci (QTL) which resolved between from within-species effects for adventitious rooting in two populations of hybrids between Pinus elliottii and P. caribaea, an outbred F1 (n=287) and an inbred-like F2 family (n=357). Most small to moderate effect QTL (each explaining 2-5% of phenotypic variation, PV) were congruent (3 out of 4 QTL in each family) and therefore considered within-species effects as they segregated in both families. A single large effect QTL (40% PV) was detected uniquely in the F2 family and assumed to be due to a between-species effect, resulting from a genetic locus with contrasting alleles in each parental species. Oligogenic as opposed to polygenic architecture was supported in both families (60% and 20% PV explained by 4 QTL in the F 2 and F1 respectively). The importance of adventitious rooting for adaptation to survive water-logged environments was thought in part to explain oligogenic architecture of what is believed to be a complex trait controlled by many hundreds of genes.

Construction of microsatellite linkage maps for Corymbia

The genus Corymbia is closely related to the genus Eucalyptus, and like Eucalyptus contains tree species that are important for sub-tropical forestry. Corymbia's close relationship with Eucalyptus suggests genetic studies in Corymbia should benefit from transfer of genetic information from its more intensively studied relatives. Here we report a genetic map for Corymbia spp. based on microsatellite markers identified de novo in Corymbia sp or transferred from Eucalyptus. A framework consensus map was generated from an outbred F 2 population (n = 90) created by crossing two unrelated Corymbia torelliana x C. citriodora subsp. variegata F1 trees. The map had a total length of 367 cM (Kosambi) and was composed of 46 microsatellite markers distributed across 13 linkage groups (LOD 3). A high proportion of Eucalyptus microsatellites (90%) transferred to Corymbia. Comparative analysis between the Corymbia map and a published Eucalyptus map identified eight homeologous linkage groups in Corymbia with 13 markers mapping on one or both maps. Further comparative analysis was limited by low power to detect linkage due to low genome coverage in Corymbia, however, there was no convincing evidence for chromosomal structural differences because instances of non-synteny were associated with large distances on the Eucalyptus map. Segregation distortion was primarily restricted to a single linkage group and due to a deficit of hybrid genotypes, suggesting that hybrid inviability was one factor shaping the genetic composition of the F2 population in this inter-subgeneric hybrid. The conservation of microsatellite loci and synteny between Corymbia and Eucalyptus suggests there will be substantial value in exchanging information between the two groups.

Mapping species differences for adventitious rooting in a Corymbia torelliana x Corymbia citriodora subspecies variegata hybrid

Quantitative trait loci (QTL) detection was carried out for adventitious rooting and associated propagation traits in a second-generation outbred Corymbia torelliana x Corymbia citriodora subspecies variegata hybrid family (n=186). The parental species of this cross are divergent in their capacity to develop roots adventitiously on stem cuttings and their propensity to form lignotubers. For the ten traits studied, there was one or two QTL detected, with some QTL explaining large amounts of phenotypic variation (e.g. 66% for one QTL for percentage rooting), suggesting that major effects influence rooting in this cross. Collocation of QTL for many strongly genetically correlated rooting traits to a single region on linkage group 12 suggested pleiotropy. A three locus model was most parsimonious for linkage group 12, however, as differences in QTL position and lower genetic correlations suggested separate loci for each of the traits of shoot production and root initiation. Species differences were thought to be the major source of phenotypic variation for some rooting rate and root quality traits because of the major QTL effects and up to 59-fold larger homospecific deviations (attributed to species differences) relative to heterospecific deviations (attributed to standing variation within species) evident at some QTL for these traits. A large homospecific/heterospecific ratio at major QTL suggested that the gene action evident in one cross may be indicative of gene action more broadly in hybrids between these species for some traits.

High subsoil chloride concentrations reduce soil water extraction and crop yield on Vertosols in north-eastern Australia

Salinity, sodicity, acidity, and phytotoxic levels of chloride (Cl) in subsoils are major constraints to crop production in many soils of north-eastern Australia because they reduce the ability of crop roots to extract water and nutrients from the soil. The complex interactions and correlations among soil properties result in multi-colinearity between soil properties and crop yield that makes it difficult to determine which constraint is the major limitation. We used ridge-regression analysis to overcome colinearity to evaluate the contribution of soil factors and water supply to the variation in the yields of 5 winter crops on soils with various levels and combinations of subsoil constraints in the region. Subsoil constraints measured were soil Cl, electrical conductivity of the saturation extract (ECse), and exchangeable sodium percentage (ESP). The ridge regression procedure selected several of the variables used in a descriptive model, which included in-crop rainfall, plant-available soil water at sowing in the 0.90-1.10 m soil layer, and soil Cl in the 0.90-1.10 m soil layer, and accounted for 77-85% of the variation in the grain yields of the 5 winter crops. Inclusion of ESP of the top soil (0.0-0.10 m soil layer) marginally increased the descriptive capability of the models for bread wheat, barley and durum wheat. Subsoil Cl concentration was found to be an effective substitute for subsoil water extraction. The estimates of the critical levels of subsoil Cl for a 10% reduction in the grain yield were 492 mg cl/kg for chickpea, 662 mg Cl/kg for durum wheat, 854 mg Cl/kg for bread wheat, 980 mg Cl/kg for canola, and 1012 mg Cl/kg for barley, thus suggesting that chickpea and durum wheat were more sensitive to subsoil Cl than bread wheat, barley, and canola.

Subsoil constraints in Vertosols: Crop water use nutrient concentration, and grain yields of bread wheat, durum wheat, barley, chickpea and canola

Single or multiple factors implicated in subsoil constraints including salinity, sodicity, and phytotoxic concentrations of chloride (Cl) are present in many Vertosols including those occurring in Queensland, Australia. The variable distribution and the complex interactions that exist between these constraints limit the agronomic or management options available to manage the soil with these subsoil constraints. The identification of crops and cultivars adapted to these adverse subsoil conditions and/or able to exploit subsoil water may be an option to maintain productivity of these soils. We evaluated relative performance of 5 winter crop species, in terms of grain yields, nutrient concentration, and ability to extract soil water, grown on soils with various levels and combinations of subsoil constraints in 19 field experiments over 2 years. Subsoil constraints were measured by levels of soil Cl, electrical conductivity of the saturation extract (ECse), and exchangeable sodium percentage (ESP). Increasing levels of subsoil constraints significantly decreased maximum depth of water extraction, grain yield, and plant-available water capacity for all the 5 crops and more so for chickpea and durum wheat than bread wheat, barley, or canola. Increasing soil Cl levels had a greater restricting effect on water availability than did ECse and ESP. We developed empirical relationships between soil Cl, ECse, and ESP and crop lower limit (CLL) for estimating subsoil water extraction by 5 winter crops. However, the presence of gypsum influenced the ability to predict CLL based on the levels of ECse. Stronger relationships between apparent unused plant-available water (CLL - LL15; LL15 is lower limit at -1.5 MPa) and soil Cl concentrations than ESP or ECse suggested that the presence of high Cl in these soils most likely inhibited the subsoil water extraction by the crops. This was supported by increased sodium (Na) and Cl concentration with a corresponding decrease in calcium (Ca) and potassium (K) in young mature leaf of bread wheat, durum wheat, and chickpea with increasing levels of subsoil constraints. Of the 2 ions, Na and Cl, the latter appears to be more damaging than the former, resulting in plant dieback and reduced grain yields.

Confirmation of QTL mapping and marker validation for partial seedling resistance to crown rot in wheat line '2-49'

We have tested the efficacy of putative microsatellite single sequence repeat (SSR) markers, previously identified in a 2-49 (Gluyas Early/Gala) × Janz doubled haploid wheat (Triticum aestivum) population, as being linked to partial seedling resistance to crown rot disease caused by Fusarium pseudograminearum. The quantitative trait loci (QTLs) delineated by these markers have been tested for linkage to resistance in an independent Gluyas Early × Janz doubled haploid population. The presence of a major QTL on chromosome 1DL (QCr.usq-1D1) and a minor QTL on chromosome 2BS (QCr.usq-2B1) was confirmed. However, a putative minor QTL on chromosome 2A was not confirmed. The QTL on 1D was inherited from Gluyas Early, a direct parent of 2-49, whereas the 2B QTL was inherited from Janz. Three other putative QTLs identified in 2-49 × Janz (on 1AL, 4BL, and 7BS) were inherited by 2-49 from Gala and were not able to be confirmed in this study. The screening of SSR markers on a small sample of elite wheat genotypes indicated that not all of the most tightly linked SSR markers flanking the major QTLs on 1D and 1A were polymorphic in all backgrounds, indicating the need for additional flanking markers when backcrossing into some elite pedigrees. Comparison of SSR haplotypes with those of other genotypes exhibiting partial crown rot resistance suggests that additional, novel sources of crown rot resistance are available.

Epithiospecifier protein activity in broccoli: The link between terminal alkenyl glucosinolates and sulphoraphane nitrile

The chemical nature of the hydrolysis products from the glucosinolate-myrosinase system depends on the presence or absence of supplementary proteins, such as epithiospecifier proteins (ESPs). ESPs (non-catalytic cofactors of myrosinase) promote the formation of epithionitriles from terminal alkenyl glucosinolates and as recent evidence suggests, simple nitriles at the expense of isothiocyanates. The ratio of ESP activity to myrosinase activity is crucial in determining the proportion of these nitriles produced on hydrolysis. Sulphoraphane, a major isothiocyanate produced in broccoli seedlings, has been found to be a potent inducer of phase 2 detoxification enzymes. However, ESP may also support the formation of the non-inductive sulphoraphane nitrile. Our objective was to monitor changes in ESP activity during the development of broccoli seedlings and link these activity changes with myrosinase activity, the level of terminal alkenyl glucosinolates and sulphoraphane nitrile formed. Here, for the first time, we show ESP activity increases up to day 2 after germination before decreasing again to seed activity levels at day 5. These activity changes paralleled changes in myrosinase activity and terminal alkenyl glucosinolate content. There is a significant relationship between ESP activity and the formation of sulforaphane nitrile in broccoli seedlings. The significance of these findings for the health benefits conferred by eating broccoli seedlings is briefly discussed.

Flour yield QTLs in three Australian doubled haploid wheat populations

Flour yield quantitative trait loci (QTLs) were identified in 3 Australian doubled haploid populations, Sunco × Tasman, CD87 × Katepwa, and Cranbrook × Halberd. Trial data from 3 to 4 sites or years were available for each population. QTLs were identified on chromosomes 2BS, 4B, 5AL, and 6BL in the Sunco × Tasman population, on chromosomes 4B, 5AS, and 6DL in the CD87 × Katepwa population, and on chromosomes 4DS, 5DS, and 7AS in the Cranbrook × Halberd population. In the Sunco × Tasman cross the highest genetic variance was detected with the QTL on chromosome 2B (31.3%), in the CD87 × Katepwa cross with the QTL on chromosome 4B (23.8%), and in the Cranbrook × Halberd cross with the QTL on chromosome 5D (18%). Only one QTL occurred in a similar location in more than one population, indicating the complexity of the flour yield character across different backgrounds.

Mapping quantitative trait loci for resistance to Pratylenchus thornei from synthetic hexaploid wheat in the International Triticeae Mapping Initiative (ITMI) population

Root-lesion nematode (Pratylenchus thornei) is a serious pathogen of wheat in many countries. The International Triticeae Mapping Initiative (ITMI) population of recombinant inbred lines (RILs) was assessed for resistance to P. thornei to determine the chromosome locations of the resistance genes. The ITMI population is derived from a cross between the resistant synthetic hexaploid wheat W-7984 and a susceptible bread wheat cultivar Opata 85. Two years of phenotypic data for resistance to P. thornei were obtained in replicated glasshouse trials. Quantitative trait locus (QTL) analysis was performed using available segregation and map data for 114 RILs. A QTL on chromosome 6DS showed consistent effects for reduced nematode numbers (partial resistance) across years and accounted for 11% and 23% of the phenotypic variation. A second QTL for P. thornei resistance on chromosome 2BS accounted for an additional 19% and 5%. Restriction fragment length polymorphism (RFLP) and simple sequence repeat (SSR) markers associated with the QTLs are physically located in regions rich in major genes at the distal ends of the short chromosome arms of 6D and 2B. SSR markers with potential for marker-assisted selection of P. thornei resistance effective in different genetic backgrounds have been identified.

Sorghum stay-green QTL individually reduce post-flowering drought-induced leaf senescence

Sorghum is an important source of food, feed, and biofuel, especially in the semi-arid tropics because this cereal is well adapted to harsh, drought-prone environments. Post-flowering drought adaptation in sorghum is associated with the stay-green phenotype. Alleles that contribute to this complex trait have been mapped to four major QTL, Stg1-Stg4, using a population derived from BTx642 and RTx7000. Near-isogenic RTx7000 lines containing BTx642 DNA spanning one or more of the four stay-green QTL were constructed. The size and location of BTx642 DNA regions in each RTx7000 NIL were analysed using 62 DNA markers spanning the four stay-green QTL. RTx7000 NILs were identified that contained BTx642 DNA completely or partially spanning Stg1, Stg2, Stg3, or Stg4. NILs were also identified that contained sub-portions of each QTL and various combinations of the four major stay-green QTL. Physiological analysis of four RTx7000 NILs containing only Stg1, Stg2, Stg3, or Stg4 showed that BTx642 alleles in each of these loci could contribute to the stay-green phenotype. RTx7000 NILs containing BTx642 DNA corresponding to Stg2 retained more green leaf area at maturity under terminal drought conditions than RTx7000 or the other RTx7000 NILs. Under post-anthesis water deficit, a trend for delayed onset of leaf senescence compared with RTx7000 was also exhibited by the Stg2, Stg3, and Stg4 NILs, while significantly lower rates of leaf senescence in relation to RTx7000 were displayed by all of the Stg NILs to varying degrees, but particularly by the Stg2 NIL. Greener leaves at anthesis relative to RTx7000, indicated by higher SPAD values, were exhibited by the Stg1 and Stg4 NILs. The RTx7000 NILs created in this study provide the starting point for in-depth analysis of stay-green physiology, interaction among stay-green QTL and map-based cloning of the genes that underlie this trait.

A marker-assisted backcross approach for developing submergence-tolerant rice cultivars

Submergence stress regularly affects 15 million hectares or more of rainfed lowland rice areas in South and Southeast Asia. A major QTL on chromosome 9, Sub1, has provided the opportunity to apply marker assisted backcrossing (MAB) to develop submergence tolerant versions of rice cultivars that are widely grown in the region. In the present study, molecular markers that were tightly linked with Sub1, flanking Sub1, and unlinked to Sub1 were used to apply foreground, recombinant, and background selection, respectively, in backcrosses between a submergence-tolerant donor and the widely grown recurrent parent Swarna. By the BC2F2 generation a submergence tolerant plant was identified that possessed Swarna type simple sequence repeat (SSR) alleles on all fragments analyzed except the tip segment of rice chromosome 9 that possessed the Sub1 locus. A BC3F2 double recombinant plant was identified that was homozygous for all Swarna type alleles except for an approximately 2.3-3.4 Mb region surrounding the Sub1 locus. The results showed that the mega variety Swarna could be efficiently converted to a submergence tolerant variety in three backcross generations, involving a time of two to three years. Polymorphic markers for foreground and recombinant selection were identified for four other mega varieties to develop a wider range of submergence tolerant varieties to meet the needs of farmers in the flood-prone regions. This approach demonstrates the effective use of marker assisted selection for a major QTL in a molecular breeding program.

Identification of quantitative trait loci for resistance to two species of root-lesion nematode (Pratylenchus thornei and P. neglectus) in wheat

Pratylenchus thornei and P. neglectus are two species of root-lesion nematode that cause substantial yield losses in wheat. No commercially available wheat variety has resistance to both species. A doubled-haploid population developed from a cross between the synthetic hexaploid wheat line CPI133872 and the bread wheat Janz was used to locate and tag quantitative trait loci (QTLs) associated with resistance to both P. thornei and P. neglectus. Wheat plants were inoculated with both species of nematode in independent replicated glasshouse trials repeated over 2 years. Known locations of wheat microsatellite markers were used to construct a framework map. After an initial single-marker analysis to detect marker-trait linkages, chromosome regions associated with putative QTLs were targetted with microsatellite markers to increase map density in the chromosome regions of interest. In total, 148 wheat microsatellite markers and 21 amplified fragment length polymorphism markers were mapped. The codominant microsatellite marker Xbarc183 on the distal end of chromosome 6DS was allelic for resistance to both P. thornei and P. neglectus. The QTL were designated QRlnt.lrc-6D.1 and QRlnn.lrc-6D.1, for the 2 traits, respectively. The allele inherited from CPI133872 explained 22.0-24.2% of the phenotypic variation for P. thornei resistance, and the allele inherited from Janz accounted for 11.3-14.0% of the phenotypic variation for P. neglectus resistance. Composite interval mapping identified markers that flank a second major QTL on chromosome 6DL (QRlnt.lrc-6D.2) that explained 8.3-13.4% of the phenotypic variation for P. thornei resistance. An additional major QTL associated with P. neglectus resistance was detected on chromosome 4DS (QRlnn.lrc-4D.1) and explained a further 10.3-15.4% of the phenotypic variation. The identification and tagging of nematode resistance genes with molecular markers will allow appropriate allele combinations to be selected, which will aid the successful breeding of wheat with dual nematode resistance.

Simple sequence repeat markers associated with three quantitative trait loci for black point resistance can be used to enrich selection populations in bread wheat

Black point in wheat has the potential to cost the Australian industry $A30.4 million a year. It is difficult and expensive to screen for resistance, so the aim of this study was to validate 3 previously identified quantitative trait loci (QTLs) for black point resistance on chromosomes 2B, 4A, and 3D of the wheat variety Sunco. Black point resistance data and simple sequence repeat (SSR) markers, linked to the resistance QTLs and suited to high-throughput assay, were analysed in the doubled haploid population, Batavia (susceptible) × Pelsart (resistant). Sunco and Pelsart both have Cook in their pedigree and both have the Triticum timopheevii translocation on 2B. SSR markers identified for the 3 genetic regions were gwm319 (2B, T. timopheevii translocation), wmc048 (4AS), and gwm341 (3DS). Gwm319 and wmc048 were associated with black point resistance in the validation population. Gwm341 may have an epistatic influence on the trait because when resistance alleles were present at both gwm319 and wmc048, the Batavia-derived allele at gwm341 was associated with a higher proportion of resistant lines. Data are presented showing the level of enrichment achieved for black point resistance, using 1, 2, or 3 of these molecular markers, and the number of associated discarded resistant lines. The level of population enrichment was found to be 1.83-fold with 6 of 17 resistant lines discarded when gwm319 and wmc048 were both used for selection. Interactions among the 3 QTLs appear complex and other genetic and epigenetic factors influence susceptibility to black point. Polymorphism was assessed for these markers within potential breeding material. This indicated that alternative markers to wmc048 may be required for some parental combinations. Based on these results, marker-assisted selection for the major black point resistance QTLs can increase the rate of genetic gain by improving the selection efficiency and may facilitate stacking of black point resistances from different sources.