Functional markers delimiting a <Emphasis Type="Italic">Medicago</Emphasis> orthologue of pea symbiotic gene <Emphasis Type="Italic">NOD3</Emphasis>

Symbiotic gene mutated in the pea (Pisum sativum L.) line RisfixC is a determinant of the number of symbiotic root nodules. In parallel to a sharp increase in nodule number, its mutational inactivation brings about the insensitivity of nodulation to the ambient nitrate level (Nts trait). Using the established localization to the SYM2-NOD3 region of the pea linkage group I, functional PCR markers were developed for the orthologous region on the chromosome 5 of the model species Medicago truncatula. Owing to the conservation of the binding regions of the designed primers, pea orthologues were successfully amplified with 60% of the primer pairs tested. When applied to a mapping pea population from the cross of the line RisfixC x Afghanistan L1268 (sym2), the new markers allowed to localize the supernodulation mutation within 2.5 cM confidence interval in the pea genome. The placement of the functional markers on the M. truncatula chromosome 5 confined the orthologous gene location to eight overlapping BACs spanning approximately 710 kbp (positions 37,755,678–38,467,472). The narrowed list of the annotated Medicago genes in combination with the published data on their symbiotic and nitrate regulation can be used for the candidate gene identification, together with the requirements imposed by the known function in nodule number initiation and nitrate sensing. In addition, the new markers are applicable for tracking the RisfixC allele in breeding programmes aimed at the improvement of symbiotic performance.     

Genetic and cytological analysis of a new spontaneous male sterility in radish (<Emphasis Type="Italic">Raphanus sativus</Emphasis> L.)

A male sterile plant appeared in the radish breeding program at the Hubei Academy of Agricultural Sciences, Hubei, China. In its progeny, a two-type (half of plants male sterile, the other half male fertile) line 01GAB was established. An F₂ population of 260 plants from a cross of male-sterile 01GAB and a male fertile line 9802H segregated for male fertility in a 3:1 ratio indicating that fertility was restored by a single dominant gene, here designated RsMs. A PCR-based DNA marker specific to the male fertility Rfob gene in 9802H was absent in 01GAB. Linkage analysis placed the RsMs locus 10.7 cM away from the Rfo locus. In an F₂ population of hybrids between 01GAB and male fertile 9802B, a co-dominant DNA marker for the RSultr3.2A (a radish sulfate transporter gene) locus was linked to the RsMs locus at 1.5 cM suggesting that fertility restoration in 01GAB was located in the region with known male sterility restorers in radish. However, no maintainer for the 01GAB source of male sterility has been identified so far. Cytological observations have shown that the abnormalities in male sterile anthers first appeared in tapetum at the tetrad stage, followed by a hypertrophy of the tapetal cells at the vacuolate microspore period. These results suggest that male sterility in 01GAB is likely to be genetic in nature, or it may represent a new type of the cytoplasmic male sterility.     

Genotype × environment interactions in populations possessing <Emphasis Type="Italic">Ga1</Emphasis>-<Emphasis Type="Italic">s</Emphasis> and <Emphasis Type="Italic">ga1</Emphasis> alleles for cross incompatibility in maize

Use of cross incompatibility in corn (Zea mays L.) by the Ga1-s allele may reduce cross-fertilization in specialty and conventional organic corn with pollen from genetically-modified (GM) corn. For effective use, information about environment, and genotype × environment effects on cross-fertilization by ga1 as well as heritability of cross incompatibility in maize is necessary. Our objective was to obtain this information. Four population pairs differing in their genotype at ga1 were evaluated for cross incompatibility with ga1 pollen in different environments. Populations were derived by crossing the recurrent parents B116, PHG35, ARZM16035:S19, and (CHZM05015:Mo17)Mo17 to Ga1-s donor parent Mo508W/Mo506W. Two replicates of each treatment were grown in the center of 952 m² fields planted with purple corn as an adventitious source of ga1/ga1 pollen. Open pollination was allowed and the amount of cross-fertilization estimated by averaging the percentage of purple seeds. Environment and genotype × environment effects were not significant. Contrasts to evaluate differences in cross-fertilization between Ga1-s and ga1 populations revealed that mean percentages of cross-fertilization in Ga1-s populations of B116, ARZM16035:S19, and (CHZM05015:Mo17)Mo17 were significantly lower than in ga1 populations. The estimated broad-sense heritability on an entry-mean basis for cross incompatibility was 0.81. Results suggest differences in genotype at ga1 played a major role in cross-fertilization of populations differing in their genotype at the ga1 locus. Incompatibility may be selected effectively over different environments and the Ga1-s system may be of value to reduce cross-fertilization with GM corn pollen.     

Characterization and molecular mapping of EMS-induced brittle culm mutants of diploid wheat (<Emphasis Type="Italic">Triticum monococcum</Emphasis> L.)

Diploid wheat (Triticum monococcum L, A^mA^m) is an ideal material for induced mutations which can be easily characterized and transferred to polyploid wheats. The EMS-induced brittle culm mutants, brc1, brc2, and brc3 used in the present investigation, were isolated from T. monococcum. All the brittle mutants had brittle roots, leaves, leaf sheaths, culms, and spikes, and were also susceptible to lodging. The mutants had 47–57% reduced α-cellulose in the secondary cell walls than that of T. monococcum indicating that all of them had defective synthesis of cellulose. All the mutants were monogenic recessive. Bulk segregation analysis of the mutants, using A^m genome anchored SSR markers in their F ₂ populations with T. boeoticum, located the mutants, brc1, brc2, and brc3 on chromosome 6A, 3A, and 1A of T. monococcum, respectively. Molecular analysis of the putatively linked markers showed that brc1 mapped on chromosome 6AS between Xbarc37 and Xbarc113 markers, brc2 on chromosome 3AL between Xcfd62 and Xcfa2170 markers whereas brc3 mapped on chromosome 1AL between Xgwm135 and Xwmc470 markers. Isolation and mapping of three different brittle culm mutants in wheat for the first time shows that there might be many more genes in wheat which affect synthesis and deposition of cellulose.     

Improved <Emphasis Type="Italic">S</Emphasis>-genotyping and new incompatibility groups in Japanese plum

The selection of cross-compatible cultivars is essential to ensure fruit set in self-incompatible species like Japanese plum and thus the S-genotype must be determined in order to establish incompatibility groups. In this study an improved Japanese plum S-genotyping method, based in polymerase chain reaction and capillary electrophoresis detection of intron polymorphisms of S-locus genes, S-RNase and SFB, has been assayed and validated in a wide sample of cultivars. This method allows a more precise determination of amplified fragment sizes and therefore a better differentiation of self-incompatibility alleles. The assayed methodology was proven effective in the detection of 13 different S-alleles of S-RNases and SFBs and was used to S-genotype 105 Japanese plum cultivars, 32 of which are described by first time in this work. Analysed cultivars were assigned into 11 incompatibility groups and two new incompatibility groups (XX and XXI) were identified, increasing to 21 the number of incompatibility groups described in this crop.     

Assessment of the uniformity and stability of grapevine cultivars using a set of microsatellite markers

Solidity of microsatellite markers is a key issue for varietal identification, especially when they are used for legal purposes, what includes their probable future use in the distinctness, uniformity and stability testing of new varieties needed for the granting of Plant Breeders’ Rights. Nine grapevine microsatellites (VVS2, VVMD5, VVMD27, VVMD28, ssrVrZAG29, ssrVrZAG62, ssrVrZAG67, ssrVrZAG83 and ssrVrZAG112), which had previously demonstrated its capacity to discriminate any grapevine variety, have been assessed to evaluate its uniformity and stability. 19 varieties were selected, representative of a high diversity for morphological, agronomical, cultural and historical aspects, as well as for microsatellite allele variability. Then, for each variety, uniformity and stability were evaluated through the analysis of 50 plants from each of three different plots, and five plants from each of seven additional plots. Material from 4,137 plants of 229 plots of the 19 varieties was sampled in seven countries. Of 3,654 plants analyzed with the set of nine microsatellites, 3,299 were of the right variety and used for the survey. An average of 172 individual values was studied for each allele of each microsatellite of each variety, and none differences were detected that could not be explained as technical variations, with the exception of several putative chimeras in two varieties. Of the total of 171 variety x microsatellite combinations, only in one combination (‘Merlot’ x VVMD27) the number of off-types exceeded the threshold allowed. The remaining 170 combinations have been found uniform and stable according to internationally accepted rules.     

Genetic analysis and genotype × environment (G × E) for grey leaf spot disease resistance in elite African maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) germplasm

Maize grey leaf spot (GLS) disease remains an important foliar disease in sub-Saharan Africa accounting for more than 25% yield losses in maize. Information on inheritance of GLS resistance of germplasm adapted to African environments is required in new sources being identified. Therefore, hybrids generated from a 10 × 10 half-diallel mating of tropical advanced maize inbred lines were evaluated in six environments to determine combining ability, genotype × environment interaction (G × E) and the impact of GLS disease on grain yield. General combining ability effects were highly significant and accounted for 72 and 68% of the variation for GLS resistance and grain yield, respectively. Significant specific combining ability effects associated with reduced disease levels were observed in some hybrids when one parent was resistant, and these may be exploited in developing single cross maize hybrids. Regression analysis showed a 260–320 kg ha^?1 decrease in maize grain yield per each increase in GLS disease severity score, and significant associations (r = ?0.31 to ?0.60) were observed between grain yield and GLS severity scores. This showed the potential of GLS disease to reduce yield in susceptible varieties grown under favourable disease conditions, without control measures. Genotype and genotype × environment biplots and correlation analysis indicated that the significant G × E observed was not due to changes in hybrid ranking, implying absence of a significant crossover interaction. Therefore, predominance of additive gene effects imply that breeding progress for GLS disease resistance would be made through selection and this could be achieved at a few hot-spot sites, such as Baynesfield and Cedara locations in South Africa, and still deploy the resistant germplasm to other environments in which they are adapted.     

Mineral bioavailability in grains of Pakistani bread wheat declines from old to current cultivars

Estimating variation in grain mineral concentration and bioavailability in relation to grain yield and the year of cultivar release is important for breeding wheat with increased content of bioavailable minerals. The grain yield and yield components, grain phytate concentration, and concentration and bioavailability of minerals (zinc Zn, iron Fe and calcium Ca) in wheat grains were estimated in 40 wheat cultivars released in Punjab (Pakistan) during the last five decades. Mean grain Zn and Ca concentrations in current-cultivars were significantly lower (≥14%) than in obsolete cultivars released during the Green Revolution (1965–1976). Much of this variation was related to increased grain weight in current-cultivars. There was a positive correlation among minerals (r = 0.39 or higher, n = 40) and minerals with phytate in wheat grains (r = 0.38 or higher, n = 40). The tested cultivars varied widely in grain yield and grain phytate-to-mineral molar ratios (phytate:mineral). Compared to obsolete cultivars, the current-cultivars had a higher phytate:mineral ratio in grains, indicating poor bioavailability of minerals to humans. The study revealed a non-significant relationship between grain yield and phytate:mineral ratios in grains. Therefore, breeding for lower phytate:mineral ratios in wheat grains can ensure increased mineral bioavailability without significant reduction in the yield potential. Future breeding should be focused on developing new genotypes suitable for mineral biofortification and with increased mineral bioavailability in grains.     

Detecting epistatic effects associated with cotton traits by a modified MDR approach

Genetic expression of a trait is complicated and it is usually associated with many genes including their interactions (epistasis) and genotype-by-environment interactions. Genetic mapping currently focuses primarily on additive models or marginal genetic effects due to the complexity of epistatic effects. Thus, there exists a need to appropriately identify favorable epistatic effects for important biological traits. Several multifactor dimensionality reduction (MDR) based methods are important resources to identify high-order gene–gene interactions. These methods are mainly focused on human genetic studies. Many traits in plant systems are not only quantitatively inherited but also are often measured in repeated field plots under multiple environments. In this study, we proposed a mixed model based MDR approach, which is suitable for inclusion of various fixed and random effects. This approach was used to analyze a cotton data set that included eight agronomic and fiber traits and 20 DNA markers. The results revealed high order epistatic effects were detected for most of these traits using this modified MDR approach.     

Mapping and marker-assisted breeding of a gene allelic to the major Asian rice gall midge resistance gene <Emphasis Type="Italic">Gm8</Emphasis>

Host plant resistance is the preferred management strategy for Asian rice gall midge (Orseolia oryzae), a serious pest in many rice-growing countries. Identification of simple sequence repeat (SSR) markers that are tightly linked to pest resistance genes can accelerate development of gene pyramids for durable/multiple resistance. Based on conventional and molecular allelism tests, we report herein that rice genotype Aganni possesses Gm8 gene, conferring hypersensitive independent (HR– type) resistance to gall midge biotypes GMB1, GMB2, GMB3, GMB4, and GMB4M. The gene Gm8 was mapped to chromosome 8 within a 400-kbp region, and the SSR markers RM22685 and RM22709 flank the gene closely. Using these closely linked flanking markers, nine other gall midge-resistant genotypes were identified as carrying the same gene Gm8. Through marker-assisted selection, Gm8 has been introgressed into an elite bacterial blight-resistant cultivar, Improved Samba-Mahsuri (IS).