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W244: Climate change poses major challenge for global wheat production and thus food security. Drought, heat, salinity and recurrent rust, septoria and fusarium epidemic waves are among the most common abiotic and biotic stresses limiting wheat crop productivity and sustainability worldwide. If adequately characterized, natural variants present in underutilized plant genetic resources (PGRs) are potentially valuable for providing improvements on resilience to abiotic stress and durable resistance to plant pathogens. In this context, Golden-standard reference genomes as well as characterized PGRs are instrumental. Both Triticum aestivum and Triticum turgidum ssp. durum share BB and AA genomes inherited from wild and domesticated emmer throughout complex steps of domestication, spread and migration events from the Fertile Crescent to diverse environments and agro-ecological conditions. With the aim of facilitating germplasm characterization and use, we assembled two comprehensive and complementary tetraploid collections: i) the Global Durum Wheat Panel (GDP) and the Tetraploid wheat Global Collection (TGC, Maccaferri et al. Nature Genetics 2019). GDP was established through a cooperative effort in the frame of the Wheat Initiative. GDP, currently maintained by ICARDA, was assembled by bringing together the durum wheat cultivated germplasm from more than 50 countries worldwide, including ca. 500 cultivars and 400 landraces, pre-breeeding lines and emmer. The Tetraploid wheat Global Collection (TGC), of 1,856 single-seed descent derived-genotypes, represents 11 tetraploid BBAA wheat taxa covering the whole distribution range. The Illumina 90K wheat SNP array was used to characterize both collections. Ca. 20,000 unique, non-redundant, single Mendelian SNP markers that were both genetically and physically mapped were use to obtain a haplo-based map of germplasm. We provide a detailed dissection of the huge reservoir of genetic diversity available in these two tetraploid genetic resources and examples of successful utilization of these resource to conduct GWAS for traits related to improvement of wheat sustainability and adaptation to climate change effects.

Among the proxy traits involved in the adaptive response of wheat to drought and other abiotic stresses, information on the QTLome governing osmotic adjustment (OA) and chlorophyll fluorescence imaging (CFI) is very limited, mainly due to the difficulty to measure these traits on a scale suitable for GWAS. In this study, OA and CFI were measured in plots of 248 elite accessions of durum wheat grown (two replicates) under a fully automated Lemnatec Field Scanalyzer (LFS) and exposed to a progressively severe drought treatment starting 5 days prior to flowering which lasted for two weeks. Leaf relative water content (RWC), chlorophyll content (SPAD) and leaf rolling (LR) were also measured. CFI was measured (i) in the dark in four time series using the LFS camera and (ii) during daytime using a light-induced fluorescence transient (LIFT) sensor mounted on a manually pushed cart. The high variability in OA and CFI among the durum accessions resulted in high to medium repeatability (h2 = 72.3 and 54.6%, respectively). At the end of the drought treatment (mean leaf RWC = 62.1%), OA and leaf RWC were positively correlated (r = 0.78). Association mapping using flowering time as covariate revealed 15 QTLs for OA (global R2 = 63.6%) as well as eight major QTL hotspots on chromosome arms 1BL, 2BL, 4AL, 5AL, 6AL, 6BL and 7BS where a higher OA capacity was always positively associated with leaf RWC, SPAD and negatively associated with LR, hence indicating a beneficial effect of OA on the water status of the plant. Additionally, the comparative analysis with previous field trials showed concurrent effects for five of these hotspots on normalized difference vegetation index (NDVI), thousand kernel weight (TKW) and/or grain yield (GY), hence supporting OA as a valuable proxy for marker-assisted selection aimed at enhancing drought resistance in wheat.

Triticum turgidum genetic resources are a wide reservoir of diversity, valuable for both durum wheat and common wheat pre-breeding, as highlighted by the success of synthetic wheats. However, a comprehensive analysis of these resources is lacking. As part of the Svevo durum wheat genome sequencing, we assembled a Global Tetraploid wheat Collection (GTC) of 1,856 accessions from 11 tetraploid wheat taxa, including wild and domesticated emmer (WEW and DEW) durum landraces (DWL) and modern durum wheat cultivars (DWC). We used the iSelect 90K SNP array anchored to the Svevo genome to investigate population structure and selection/demography signatures. We traced: i) the inheritance of haplotype blocks from either the North-eastern or Southern Levantine Fertile Crescent (Turkey) WEW populations to whole DEW germplasm, with a 0.65/0.35 overall inheritance ration, ii) the two subsequent independent but similar star-like dispersal patterns associated to DEW and DWL evolution, with six main populations each. Durum wheat most probably originated from the Southern-levant DEW. Ethiopian emmer and durum, T. turanicum and T. carthlicum were the most differentiated with minimal contribution to modern durum. Modern durum originated mostly from the North-African and Transcaucasian DWL populations. WEW, with the highest genome-wide diversity, provided the reference for assessing the diversity reductions associated to domestication and breeding. Numerous strong diversity depletions signals were observed primarily for the WEW-to-DEW transition. These signals progressively consolidated through domestication and breeding. Specific DEW-to-DWL signals were also observed. Diversity reduction index (DRI), Fst, haplotype-based XP-EHH and hapFLK and spatial pattern of site frequency spectrum (XP-CLR) metrics were considered. Frequently two or more indexes occurred in overlapping regions (selection clusters). In total, 104 pericentromeric (average size of 107.7 Mb) and 350 non-pericentromeric (average size of 11.4 Mb) clusters were identified. WEW-to-DEW and DEW-to-DWL transitions mostly involved extended pericentromeric regions tagged by DRI and Fst, while the DEW-to-DWL and DWL-to-DWC transitions were mostly associated to numerous XP-EHH and XP-CLR signals. We carried out a comprehensive projection of the published tetraploid QTLome and investigated the genome-wide QTL distribution and co-location between meta-QTLs and demography/selection signals. The usefulness of the GTC to elucidate the evolutionary patterns associated to loci, haplotype blocks and causative sequence variants for traits of breeding interest like cadmium-cumulation in grains, grain size and disease resistance loci are reported.

The genome of modern durum wheat (DW) cultivar Svevo has been assembled based on a combination of whole genome shotgun sequencing (270X), NRGene deNovoMagic assembler, high-resolution genetic mapping obtained from the cross between Svevo DW and Zavitan wild emmer wheat (WEW) and scaffold ordering based on chromosome conformation capture sequencing (Hi-C). The assembly consisted of 9.96 Gb of ordered sequences with 66,559 high-confidence (HC) genes. We used this resource to investigate the genetic diversity and ancestry of tetraploid wheat germplasm. iSelect 90K SNP array was used to genotype a global collection of 1,858 non-redundant accessions covering the whole range of tetraploid genetic resources from WEW, cultivated emmer (CEW), durum landraces (DWL) and modern durum cultivars (DWC). We performed a whole-genome scan for population genetic structure, selective sweeps together with the tetraploid QTLome projection. Average whole-genome genetic diversity were pWEW= 0.285, pCEW = 0.254, pDWL = 0.201, pDWC = 0.192, with an overall WEW-DWC decrease in diversity equal to 32.6%. Diversity depletions were more relevant in peri-centromeric regions (pWEW_C = 0.269, pDWC_C = 0.151) as compared to the highly-recombinogenic distal regions (pWEW_R = 0.287, pDWC_R = 0.250). From WEW to DWC, 68 chromosome regions were subjected to diversity depletion, affecting up to 38% of the genome in total: 19 of these were associated to WEW-CEW transition, 41 to CEW-DWL and 8 to DWL-DWC. The gene content of these regions is being explored in relation to known QTL content and haplotype analysis. Overall, the analysis pointed out the chromosome regions subjected to strong selective sweeps during the domestication and breeding selection, on one side, and those regions that would benefit from targeted genetic diversity restoration on the other side.

PubMedID: 30962619 The domestication of wild emmer wheat led to the selection of modern durum wheat, grown mainly for pasta production. We describe the 10.45 gigabase (Gb) assembly of the genome of durum wheat cultivar Svevo. The assembly enabled genome-wide genetic diversity analyses revealing the changes imposed by thousands of years of empirical selection and breeding. Regions exhibiting strong signatures of genetic divergence associated with domestication and breeding were widespread in the genome with several major diversity losses in the pericentromeric regions. A locus on chromosome 5B carries a gene encoding a metal transporter (TdHMA3-B1) with a non-functional variant causing high accumulation of cadmium in grain. The high-cadmium allele, widespread among durum cultivars but undetected in wild emmer accessions, increased in frequency from domesticated emmer to modern durum wheat. The rapid cloning of TdHMA3-B1 rescues a wild beneficial allele and demonstrates the practical use of the Svevo genome for wheat improvement. © 2019, The Author(s), under exclusive licence to Springer Nature America, Inc. Western Grains Research Foundation Ministry of Agriculture - Saskatchewan Israel Science Foundation: 1137/17 031A536 FP7 Food, Agriculture and Fisheries, Biotechnology: DROPS ID244347 Natural Sciences and Engineering Research Council of Canada InterOmics PB05 Genome Prairie United States-Israel Binational Science Foundation: 2015409 Genome Canada Saskatchewan Canola Development Commission 2819103915 PIR01_00017 3060-21000-038-00-D We acknowledge the funding support of: the Italian Ministry of Education and Research with projects CNR Flagship InterOmics PB05 (L.M., A.C., G.S.), PON ELIXIR CNR-BiOmics PIR01_00017 (L.M., M.G., M.Mo.) and PON ISCOCEM (P.D.); CREA project Interomics (L.C.); Fondazione in rete per la ricerca agroalimentare AGER project From Seed to Pasta (R.T.); FP7-KBBE Project DROPS ID244347 (R.T.); Genome Canada (A.G.S., C.P.); the Western Grain Research Foundation (A.G.S., C.P.); the Manitoba Wheat and Barley Commission (A.G.S., C.P.); the Saskatchewan Wheat Development Commission (A.G.S., C.P.); the Alberta Wheat Development Commission (A.G.S., C.P.); the Saskatchewan Ministry of Agriculture (A.G.S., C.P.); the administrative support of Genome Prairie (A.G.S., C.P.); Canadian Triticum Applied Genomics -CTAG2-(A.G.S., C.P.); Binational Science Foundation grant no. 2015409 (I.H., A.D.); Israel Science Foundation grant no. 1137/17 (A.D.); USDA-Agricultural Research Service Current Research Information System project 3060-21000-038-00-D (J.D.F., S.S.X.); German Federal Ministry of Food and Agriculture grant no. 2819103915 (N.S., K.F.X.M.); German Ministry of Education and Research grant no. 031A536 (K.F.X.M.); and Natural Sciences and Engineering Council of Canada grant nos. SPG 336119-06 and RGPIN 92787 (G.J.T., C.P.). The authors are grateful to E. Elias (North Dakota State University) for providing nine DW cultivars, included in the Global Tetraploid Wheat Collection and E. Scarpella (University of Alberta, Edmonton, Canada) for assistance with confocal microscopy.

Producing better and healthier food is an unprecedented challenge that the food-supply chain will face, particularly in view of the negative impact of climate change on food security and safety. Among the different options available to meet these challenges, the recent advances in genome sequencing provide unparalleled opportunities to identify beneficial alleles at the genes controlling food properties and leverage this information for tailoring the genetic make-up of the crops that feed mankind. Breeding programmes aiming at selecting more nutritious and safer crops increasingly benefit from the application of marker-assisted selection and genomic selection. Cloning the genes underscoring food quality paves the way for the targeted use of gene editing to fully harness the opportunities offered by native and artificially induced genetic variability.

Given the importance of early cold tolerance in maize (Zea mays L.), we previously conducted a divergent selection for low (L) and high (H) cold tolerance at germination. The source population was the F2 of B73 × IABO78 single cross. Selection was first conducted as recurrent selection (four cycles) and then in inbreeding (three generations). Ten lines were finally selected for L and ten for H; these lines were then evaluated to investigate: (i) their field cold tolerance in the early autotrophic phase and the relationship with the cold tolerance expressed at emergence and in the heterotrophic phase; (ii) the inheritance of these cold tolerance traits; (iii) their agronomic performance as lines per se and as crosses; (iv) their SNP-based genetic characterization. For all cold tolerance traits, the H group of lines was superior to the L group; selection responses were often symmetric, suggesting the prevalence of additive effects. The relationship among cold tolerance traits was appreciable, suggesting a common genetic basis. Differences among testcrosses were largely due to general combining ability effects and the relationship between line per se and testcross performance was high, confirming the prevalence of additive effects. Correlated responses were also obtained for agronomic traits, the H lines being superior to L lines both per se and as crosses. For SNP-marker loci, the B73 allelic frequency was greater in the H group (61.7 %) than in L group (39.7 %). Across field trials, an H line excelled for all traits, thus proving to deserve further investigations.

Durum wheat (Triticum turgidum subsp. durum) is a key crop worldwide, and yet, its improvement and adaptation to emerging environmental threats is made difficult by the limited amount of allelic variation included in its elite pool. New allelic diversity may provide novel loci to international crop breeding through quantitative trait loci (QTL) mapping in unexplored material. Here, we report the extensive molecular and phenotypic characterization of hundreds of Ethiopian durum wheat landraces and several Ethiopian improved lines. We test 81 587 markers scoring 30 155 single nucleotide polymorphisms and use them to survey the diversity, structure, and genome-specific variation in the panel. We show the uniqueness of Ethiopian germplasm using a siding collection of Mediterranean durum wheat accessions. We phenotype the Ethiopian panel for ten agronomic traits in two highly diversified Ethiopian environments for two consecutive years and use this information to conduct a genome-wide association study. We identify several loci underpinning agronomic traits of interest, both confirming loci already reported and describing new promising genomic regions. These loci may be efficiently targeted with molecular markers already available to conduct marker-assisted selection in Ethiopian and international wheat. We show that Ethiopian durum wheat represents an important and mostly unexplored source of durum wheat diversity. The panel analysed in this study allows the accumulation of QTL mapping experiments, providing the initial step for a quantitative, methodical exploitation of untapped diversity in producing a better wheat. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Divergently selected genotypes can be used for detecting the genomic regions affecting the selected trait (selection signature). Moreover, the genetic distances (GDs) among divergently selected lines can be correlated with the agronomic performances of the crosses among them. Using as source the maize F2 of B73 × IABO78, we previously conducted four cycles of divergent recurrent selection and three cycles of divergent selection in inbreeding for cold tolerance at germination. We finally obtained 10 lines selected for low (L) and 10 lines selected for high (H) cold tolerance, which exhibited a notable divergence for both the selected and associated traits. Herein, we investigated the 20 lines and the 28 single diallel crosses among eight random lines (four L and four H); the main objectives were to identify the putative regions controlling the selected and associated traits and to study the relationships between crosses performances and GDs among their parental lines. Allele frequencies at 932 recombination blocks based on 19,220 polymorphic SNPs were obtained for the two lines’ groups; the F ST calculated across sliding windows indicated 18 regions highly divergent between groups. The increasing alleles for cold tolerance were contributed by both parents, consistently with the transgressive segregations previously found. Several regions associated to DG also affected various agronomic traits. The cross performances showed some relationships with the genetic distances among parental lines for traits affected by dominance, provided that all crosses were considered, while these relationships vanished when only L × H crosses were examined.

Low-temperature stress is considered as the major abiotic constraint limiting plant’s growth and the potential land cultivation. Crop adaptation to limiting temperature is thus an important breeding objective because it determines yield stability in environment-friendly cultivation practices. Conventional breeding methods had limited success in improving the cold tolerance of important crop plants because of the complexity of stress tolerance traits, low genetic variance, and lack of efficient selection criteria. The knowledge of physiology, of genetics, and of the DNA technology has improved substantially nowadays, and these advancements will allow the breeder to predict the breeding value of best genotypes by using physiology, genetics, and molecular information. The perspective for selecting more effectively cold-tolerant crops will involve efficient genotyping, reliable phenotyping and envirotyping, and adequate statistical models.

Maize originated in tropical areas and improving cold tolerance is an important breeding objective for cultivation in high latitudes. We review the main limitations in understanding and improving cold tolerance in maize and the contribution of genomics in dissecting the genetic basis of the trait and selecting better genotypes. Physiological analyses revealed that non-optimal temperature exerts detrimental effects on a multitude of metabolic functions at different growing stages, each under the control of independent gene sets. Loci controlling cold tolerance at different growing stages have been investigated by means of linkage mapping or genome-wide association, revealing that no major genes are responsible for the trait. This finding was confirmed in transcriptomic studies that always revealed multiple candidates, and a large amount of data is being collected that altogether will make it possible to obtain a more coherent picture of response to cold. To harness the increasing body of information available from the maize genome sequence and gene expression data, new bioinformatics tools will be helpful for integrating the big-data obtained from the large-scale genomics and phenomics experiments. With the enhancement of knowledge, plant science is shifting its focus from “explanatory” to “predictive” and from a plant breeding perspective the focus will be predicting the breeding value of the best genotypes by using molecular information. The future strategies for selection of cold tolerance will involve intensive genotyping, high-precision phenotyping and advanced statistical analyses to predict the optimal genotypes for more time- and cost-efficient breeding strategies.

Bo23 (Reg. No. GP-599, PI 677346) is a maize (Zea mays L.) inbred line developed at the Department of Agricultural Sciences, University of Bologna (Bologna, Italy), using as source the F2 of the cross B73 × IABO78. B73 is a well-known inbred from a US Corn Belt germplasm, and IABO78 is an inbred from Italian germplasm not previously used in breeding. Bo23 was released in 2016 due to its high cold tolerance and agronomic performance when sown early. The breeding procedure leading to Bo23 consisted of four cycles of full-sib recurrent selection and three cycles of selection in inbreeding for germination at low temperature. In the evaluation of selection responses, a line coded as H102 (now Bo23) excelled for cold tolerance in the field and for agronomic traits. Therefore, Bo23 was further evaluated in seven trials conducted in early sowing in the southern Po Valley (Italy). The line again exhibited outstanding performance for cold tolerance and agronomic traits, both per se and as testcross. Bo23 was superior to B73 for most traits, indicating that its performance is also due to alleles of IABO78. This implies that the use of Bo23 in breeding for cold tolerance and agronomic traits can also contribute the beneficial genetic variation from its Italian parent.

Previous studies revealed that defoliation can bring about differential responses in maize (Zea mays L.) genotypes for cold tolerance. This research was conducted to study the inheritance of the responses to defoliation for germination at low temperature and for seedling traits in early-sown field trials. Six inbred lines were crossed according to the complete diallel scheme, thus producing 36 geno- types. At milk ripening stage, half of the plants of each genotype were completely defoliated (D) whereas the other half were not (ND). Two experiments were conducted for two years, one in the germinator and one in the field. The response to defoliation was evaluated as (D–ND). Across the two years, the D treatment caused an average kernel weight decrease of - 56 mg ( - 23.1 % as referred to ND), an increase of 2.3 % for germination at 9 C (G9) and a reduction of - 0.3 d for the average time of germination (ATG); in contrast, the defoliation effect was negligible for germination at 25 C. In the field, the defoliation effects were more notable in the first (colder) year and led to an increase of 2.0 % for field emergence (FE) and to a decrease of - 2.7 g for seedling fresh weight (SFW). The genotypic variation for the (D–ND) response was significant for additive, dominance and reciprocal effects for G9, ATG, FE and SFW. There was consistency among lines across traits for additive effects, with Lo1016 and Os420 showing the best and the worst effect, respectively.

In a previous study on a maize (Zea mays L.) population of recombinant inbreds derived from B73 × H99, we identified several quantitative trait loci (QTL) for agronomic traits with high dominance-additive ratio. Then, for four of these QTL, we developed families of near-isogenic lines (NILs) homozygous either for the QTL allele from B73 (BB) or from H99 (HH); for two of these QTL, the NILs' families were produced in two different genetic backgrounds. The present study was conducted to: (1) characterize these QTL for agronomic traits and (2) verify whether their effects were influenced by the genetic background, inbreeding level and plant density (PD). The six NILs' families were tested across 3 years and in three experiments at different inbreeding levels as NILs per se and their reciprocal crosses (Experiment 1), NILs crossed to related inbreds B73 and H99 (Experiment 2) and NILs crossed to four unrelated inbreds (Experiment 3). Experiment 2 was conducted at two PDs (4.5 and 9.0 plants m(-2)). Results of Experiments 1 and 2 confirmed previous findings as to QTL effects, with dominance-additive ratio superior to 1 for several traits; as a tendency, dominance effects were more pronounced in Experiment 1. The QTL effects were also confirmed in Experiment 3. The interactions involving QTL effects, families and PD were generally negligible, suggesting a certain stability of the QTL. Results emphasize the importance of dominance effects for these QTL, suggesting that they might deserve further studies, using the NILs' families and their crosses as base materials.

Background Maize (Zea mays) is a globally produced crop with broad genetic and phenotypic variation. New tools that improve our understanding of the genetic basis of quantitative traits are needed to guide predictive crop breeding. We have produced the first balanced multi-parental population in maize, a tool that provides high diversity and dense recombination events to allow routine quantitative trait loci (QTL) mapping in maize. Results We produced 1,636 MAGIC maize recombinant inbred lines derived from eight genetically diverse founder lines. The characterization of 529 MAGIC maize lines shows that the population is a balanced, evenly differentiated mosaic of the eight founders, with mapping power and resolution strengthened by high minor allele frequencies and a fast decay of linkage disequilibrium. We show how MAGIC maize may find strong candidate genes by incorporating genome sequencing and transcriptomics data. We discuss three QTL for grain yield and three for flowering time, reporting candidate genes. Power simulations show that subsets of MAGIC maize might achieve high-power and high-definition QTL mapping. Conclusions We demonstrate MAGIC maize’s value in identifying the genetic bases of complex traits of agronomic relevance. The design of MAGIC maize allows the accumulation of sequencing and transcriptomics layers to guide the identification of candidate genes for a number of maize traits at different developmental stages. The characterization of the full MAGIC maize population will lead to higher power and definition in QTL mapping, and lay the basis for improved understanding of maize phenotypes, heterosis included. MAGIC maize is available to researchers. Electronic supplementary material The online version of this article (doi:10.1186/s13059-015-0716-z) contains supplementary material, which is available to authorized users.

Summary Multiparental cross designs for mapping quantitative trait loci (QTL) provide an efficient alternative to biparental populations because of their broader genetic basis and potentially higher mapping resolution. We describe the development and deployment of a recombinant inbred line (RIL) population in durum wheat (Triticum turgidum ssp. durum) obtained by crossing four elite cultivars. A linkage map spanning 2664 cM and including 7594 single nucleotide polymorphisms (SNPs) was produced by genotyping 338 RILs. QTL analysis was carried out by both interval mapping on founder haplotype probabilities and SNP bi-allelic tests for heading date and maturity date, plant height and grain yield from four field experiments. Sixteen QTL were identified across environments and detection methods, including two yield QTL on chromosomes 2BL and 7AS, with the former mapped independently from the photoperiod response gene Ppd-B1, while the latter overlapped with the vernalization locus VRN-A3. Additionally, 21 QTL with environment-specific effects were found. Our results indicated a prevalence of environment-specific QTL with relatively small effect on the control of grain yield. For all traits, functionally different QTL alleles in terms of direction and size of genetic effect were distributed among parents. We showed that QTL results based on founder haplotypes closely matched functional alleles at known heading date loci. Despite the four founders, only 2.1 different functional haplotypes were estimated per QTL, on average. This durum wheat population provides a mapping resource for detailed genetic dissection of agronomic traits in an elite background typical of breeding programmes.

Defoliation during maize (Zea mays L.) kernel development has been observed to induce tolerance to cold of germinating seeds in responsive genotypes. The objectives of this study were to evaluate the response to defoliation of immature embryo and mature seed germinability at cold and to verify if the response was influenced by the developmental stage at which the treatment was applied. In three environments, six inbred lines (B73, IABO78, Lo1016, Lo964, Mo17, Os420) were defoliated (D) approximately 20 days after pollination (DAP) or not defoliated (ND). Immature embryos were excised three days after defoliation and germinated in vitro at 9 or 25 ∘C. At maturation, kernel germination was tested at the same temperatures. Defoliation improved cold tolerance and mean time to germination (MTG) at 9 ∘C of both embryos and kernels of Lo1016. To study the effect of kernel developmental stage on response to defoliation, plants of B73, Lo1016 and Lo964 were defoliated at 15, 18, 21, 24, 27, 30, 33, 36, and 39 DAP, or not defoliated. At the same DAP, immature grains were analyzed for dry weight, water and abscisic acid (ABA) content. In Lo1016, low amounts of kernel ABA were detected at all stages, while in Lo964 and B73 ABA increased during development. Lo1016 mature kernels showed an improvement of cold tolerance due to defoliation at all times, while the other genotypes did not. In conclusion inbred lines showed variability for mature seed and immature embryo tolerance to cold at germination and for the ability to acquire tolerance after defoliation.

Pollen genotype selection for genes expressed in both the haploid and diploid phases of the plant life cycle can lead to correlated responses detectable in the sporophyte. A pollen selection was conducted in two genetic backgrounds of maize (Zea mays L.) for chlorsulfuron resistance, conferred by the XA17 allele. Plants of two backcross (BC) lines segregating 1 (heterozygote, resistant) : 1 (homozygote, susceptible) for chlorsulfuron resistance were used as pollen donor. Selection treatment was applied during microspore development, and tassels were cut about 10 days before anthesis and placed into a liquid medium with or without 40 mg l–1 of chlorsulfuron. Pollen was used to fertilize an unrelated susceptible genotype (tester). The resulting testcrosses (TC) were evaluated in the greenhouse by spraying seedlings with chlorsulfuron at 23 g ha–1. Non-selected TC progenies derived from heterozygous BC plants showed a proportion of resistant and susceptible plants close to the expected 1:1 ratio, while non-selected TC progenies derived from homozygous BC plants showed susceptible plants only. Selected TC progenies derived from heterozygous BC plants showed a frequency of resistant plants ranging from 89% to 100%. BC plants homozygous for the susceptible allele, subjected to selection treatment, gave poor seed set, and no resistant plants were found in their TC progenies. Resistant TC plants obtained through pollen genotype selection were selfed, and the proportion of resistant plants was close to 3:1 in all selfed families, in accordance with the hypothesis that all of them inherited the XA17 allele through selection. In this study, pollen genotype selection was extremely effective, and its effect persisted in the second sporophytic generation.

Soil carryover of the herbicide trifluralin can injureZea mays.Therefore, the development of resistant hybrids can be an important breeding objective. This research was conducted to study the genetic variability for trifluralin resistance inZ. mays, the effects of genes controlling resistance, and the seed lipid content of resistant (R) and susceptible (S) inbreds. Twenty inbreds were tested under greenhouse conditions at three trifluralin rates (0, 12.5, and 125 g ai ha−1). Lo1067 was the most resistant, and A632 was the most susceptible inbred. Hybrids among R and S inbreds were tested, along with their parents, under greenhouse conditions (using the same three rates) and in the field (at 0, 0.4, and 0.8 kg ai ha−1). Under both greenhouse and field conditions, inbreds R were more resistant than S. Hybrids R × R were more resistant than S × S, indicating that additive effects were important. Hybrids R × S and S × R did not significantly differ, indicating that reciprocal effects were not important. On average, hybrids R × S and S × R were intermediate between R × R and S × S, suggesting that nonadditive effects were negligible. The difference between the mean across hybrids and the mean across parents (further estimating the importance of nonadditive effects) was significant only for parameters investigated in the greenhouse. Greenhouse data were correlated with field data, but the coefficients of determination were < 50%. The ability to predict hybrid resistance on the basis of parental mean was higher in the greenhouse (r2= 0.78) than in the field (r2= 0.47). R and S inbreds also differed in seed lipid content, but correlations were negligible with greenhouse and field data. Data indicated the presence of genetic variability for trifluralin resistance, that additive effects were prevailing, and that the resistance level was not related to seed lipid content.

Full-sib family recurrent selection for grain yield was conducted for four cycles in maize (Zea mays L.). The source population was the F 2 of A632 (with red cob) × Mu195 (with white cob). Objectives of this research were to evaluate the change in allelic frequency at the P1 gene (determining cob color), to assess whether this change was due to random drift only, and to detect whether allelic frequencies were correlated with grain yield responses. The source and the four selected populations were crossed to two inbred lines with white cob (recessive trait). The frequency of red cob allele increased with selection following a highly significant linear trend (b = 0.066 cycle -1 ), suggesting the effect of directional selection. Correlation between allelic frequency and grain yield was significant for populations as S2 generation (r = 0.938), and as testcrosses (r = 0.886), and could be ascribed to tight linkage, to pleiotropy, or both.

Background Despite the crucial role that heterosis has played in crop improvement, its genetic and molecular bases are still elusive. Several types of structured populations were used to discover the genetic architecture underlying complex phenotypes, and several QTL related to heterosis were detected. However, such analyses generally lacked the statistical power required for the detailed characterization of individual QTL. Currently, QTL introgression into near-isogenic materials is considered the most effective strategy to this end, despite such materials inevitably contain a variable, unknown and undesired proportion of non-isogenic genome. An introgression program based on residual heterozygous lines allowed us to develop five pairs of maize (Zea mays L.) near-isogenic lines (NILs) suitable for the fine characterization of three major heterotic QTL previously detected. Here we describe the results of the detailed genomic characterization of these NILs that we undertook to establish their genotypic structure, to verify the presence of the expected genotypes within target QTL regions, and to determine the extent and location of residual non-isogenic genomic regions. Results The SNP genotyping approach allowed us to determine the parent-of-origin allele for 14,937 polymorphic SNPs and to describe in detail the genotypic structure of all NILs. The correct introgression was confirmed for all target QTL in the respective NIL and several non-isogenic regions were detected genome-wide. Possible linkage drag effects associated to the specific introgressed regions were observed. The extent and position of other non-isogenic regions varied among NIL pairs, probably deriving from random segregating sections still present at the separation of lineages within pairs. Conclusions The results of this work strongly suggest that the actual isogenicity and the genotypic architecture of near-isogenic materials should be monitored both during the introgression procedure and on the final materials as a paramount requisite for a successful mendelization of target QTL. The information here gathered on the genotypic structure of NILs will be integrated in future experimental programs aimed at the fine mapping and isolation of major heterotic QTL, a crucial step towards the understanding of the molecular bases of heterosis in maize.

Recent advances in high-throughput sequencing technologies have triggered a shift toward single-nucleotide polymorphism (SNP) markers. A systematic bias can be introduced if SNPs are ascertained in a small panel of genotypes and then used for characterizing a larger population (ascertainment bias). With the objective of evaluating a potential ascertainment bias of the Illumina MaizeSNP50 array with respect to elite European maize dent and flint inbred lines, we compared the genetic diversity among these materials based on 731 amplified fragment length polymorphisms (AFLPs), 186 simple sequence repeats (SSRs), 41,434 SNPs of the MaizeSNP50 array (SNP-A), and two subsets of it, i.e., 30,068 Panzea (SNP-P) and 11,366 Syngenta markers (SNP-S). We evaluated the bias effects on major allele frequency, allele number, gene diversity, modified Roger's distance (MRD), and on molecular variance (AMOVA). We revealed ascertainment bias in SNP-A, compared to AFLPs and SSRs. It affected especially European flint lines analyzed with markers (SNP-S) specifically developed to maximize differences among North American dent germplasm. The bias affected all genetic parameters, but did not substantially alter the relative distances between inbred lines within groups. For these reasons, we conclude that the SNP markers of the MaizeSNP50 array can be employed for breeding purposes in the investigated material. However, attention should be paid in case of comparisons between genotypes belonging to different heterotic groups. In this case, it is advisable to prefer a marker subset with potentially low ascertainment bias, like in our case the SNP-P marker set.

The early genotypes of maize (Zea mays L.) selected for the northern Corn Belt regions can perform poorly when grown as a delayed-sowing crop in warmer regions. Two early synthetics, A and B, were developed and two cycles of a modified reciprocal recurrent selection were conducted in a delayed-sowing crop by using A632 as a tester for Synthetic A and W117 as a tester for Synthetic B. Selection was based on a multitrait index involving grain yield, percent erect plants, and grain moisture. This study was conducted in three environments to evaluate the effects of selection. The source and selected populations of the two synthetics were tested per se and in hybrid combination to each other, to their inbred tester, and to the unrelated hybrid, A188 × W64A. The gain in the cross between the two synthetics was 4.9% cycle -1 (P≤0.01) for the performance index (calculated similarly to the selection index), and this was mainly due to the gain in grain yield (12.2% cycle -1 ). However, undesired increases were found for grain moisture and silking date. The gains for performance index and grain yield were due to a balanced contribution of both changes per cycle of populations per se and of heterosis. Synthetic B contributed more than Synthetic A to the gain in the population cross for performance index, grain yield, and to the loss of earliness. Population crosses of both synthetics with corresponding testers and A188 × W64A showed a positive response for performance index and grain yield, and a loss of earliness for Synthetic B. The results indicated that the selection procedure was effective in improving the performance index of the two synthetics both per se and in hybrid combinations

Improving cold tolerance in maize (Zea mays L.) is an important breeding objective, allowing early sowings which result in many agronomic advantages. Using as source the F(2) population of B73 × IABO78 single cross, we previously conducted four cycles of divergent recurrent selection for high (H) and low (L) cold tolerance level, evaluated as the difference (DG) between germination at 9.5 °C and at 25 °C in the germinator. Then, we pursued the divergent selection in inbreeding from S(1) to S(4). This research was conducted to study (1) the direct response to selection (by testing ten S(4) L and ten S(4) H lines), (2) the trait inheritance (in a complete diallel scheme involving four L and four H lines), (3) the associated responses for cold tolerance in the field (at early and delayed sowings) and (4) the responses for other traits, by testing the ten L and the ten H lines at usual sowing. Selection was effective, leading to appreciable and symmetric responses for DG. Variation among crosses was mainly due to additive effects and the ability to predict hybrid DG based on parental lines DG was appreciable. Associated responses for cold tolerance traits in the field were noticeable, though the relationship between DG and these traits was not outstanding. High tolerance was also associated with early flowering, short plants, less leaves, low kernel moisture, red and thin cob, and flint kernels. These divergently selected lines can represent valuable materials for undertaking basic studies and breeding works concerning cold tolerance.

The objectives of this research were to determine if genes controlling the reaction to the herbicide acetochlor in maize (Zea mays L.) are active during both the haploid and the diploid phases of the life cycle and if pollen selection can be utilized for improving sporophytic resistance. Pollen of eight inbred lines, previously characterized through sporophytic analysis for the level of tolerance to acetochlor, showed a differential reaction to the herbicide forin vitro tube length; moreover, such pollen reactions proved to be significantly correlated (r =0.786(*),df=6) with those of the sporophytes producing the pollen. Pollen analysis of two inbred lines (i.e. Mo17, tolerant, and B79, susceptible) and their single cross showed that thein vitro pollen-tube length reaction of the hybrid was intermediate between those of two parents. An experiment on pollen selection was then performed by growing tassels of Mo17xB79 in the presence of the herbicide. Pollen obtained from treated tassels showed a greater tolerance to acetochlor, assessed asin vitro tube length reaction, than pollen obtained from control tassels. Moreover, the backcross [B79 (Mo17xB79)] sporophytic population obtained using pollen from the treated tassels was more tolerant (as indicated by the fresh weight of plants grown in the presence of the herbicide) than was the control backcross population. The two populations did not differ when grown without the herbicide. These findings indicate that genes controlling the reaction to acetochlor in maize have haplodiploid expression; consequently, pollen selection can be applied for improving plant tolerance.

In order to assess the efficiency of male gametophytic selection (MGS) for crop improvement, pollen selection for tolerance to herbicide was applied in maize. The experiment was designed to test the parallel reactivity to Alachlor of pollen and plants grown in controlled conditions or in the field, the response to pollen selection in the sporophytic progeny, the response to a second cycle of MGS, and the transmission of the selected trait to the following generations. The results demonstrated that pollen assay can be used to predict Alachlor tolerance under field conditions and to monitor the response to selection. A positive response to selection applied to pollen in the sporophytic progeny was obtained in diverse genetic backgrounds, indicating that the technique can be generally included in standard breeding programs; the analysis of the data produced in a second selection cycle indicated that the selected trait is maintained in the next generation.

The proteins performing the activity of the inner membrane anion channel (IMAC) and its plant counterpart (PIMAC) are still unknown. Lurin et al. (Biochem J 348: 291-295, 2000) indicated that a chloride channel (CLC) protein corresponds to PIMAC activity in tobacco seedling mitochondria. In this study, we investigated: (i) the presence of a CLC protein in maize seedling mitochondria; (ii) the involvement of this protein in plant cold tolerance; and (iii) its possible role in PIMAC activity. We validated the presence of a CLC protein (ZmCLCc) in maize mitochondria by immunoassay using a polyclonal antibody against its C-terminus. The differential expression of the ZmCLCc protein in mitochondria was measured in seedlings of maize populations divergently selected for cold tolerance and grown at different temperatures. The ZmCLCc protein level was higher in cold stressed than in non-stressed growing conditions. Moreover, the ZmCLCc level showed a direct relationship with the cold sensitivity level of the populations under both growing conditions, suggesting that selection for cold tolerance induced a constitutive change of the ZmCLCc protein amount in mitochondria. The anti-ZmCLCc antibody inhibited (about 60%) the channel-mediated anion translocations by PIMAC, whereas the same antibody did not affect the free diffusion of potassium thiocyanide through the inner mitochondrial membrane. For this reason, we conclude that the mitochondrial ZmCLCc protein can perform the PIMAC activity in maize seedlings.

Genomics approaches and platforms offer novel opportunities to identify and clone the loci that control root architecture in maize. Mutants for root features have been described and in some cases the relevant genes have been cloned. A number of studies have also described quantitative trait loci (QTLs) that provide access to valuable genetic diversity for the morpho-physiological features that characterize root functionality. Nonetheless, although a number of major QTLs have been identified, none of these QTLs has been cloned so far, mainly due to the difficulty to accurately phenotype the target traits in a sufficiently large number of plants. It is foreseeable that QTL cloning will be facilitated by the adoption of high-throughput phenomics platforms as well as by the information made available through the sequencing of the maize genome and the profiling of the transcriptome, proteome, and metabolome, all of which will contribute to the identification of plausible candidate genes. Allele mining in germplasm and mutant collections through forward- and reverse-genetics approaches, coupled with marker-assisted backcrossing and/or genetic engineering, will further facilitate the introgression of novel genetic variation in elite materials. New QTL-based modeling approaches will improve our capacity to understand genotype × environment interaction at varying water and nutrient regimes, thus contributing to define the most promising “molecular” ideotypes. This notwithstanding, a sizeable impact of marker-assisted selection and other genomics approaches in tailoring the ideal root architecture in maize will only be possible through a deeper knowledge of root functions under a broad range of growing conditions and an integration with conventional breeding methodologies.

The activity of the plant inner membrane mitochondrial anion channel (PIMAC) is involved in metabolite shuttles and mitochondrial volume changes and could have a role in plant temperature tolerance. Our objectives were to in- vestigate (i) the occurrence and (ii) the temperature de- pendence of anion fluxes through PIMAC in mitochondria isolated from seedlings of three maize populations differing in terms of cold tolerance; and (iii) the relationships between the PIMAC activity kinetics and the level of cold tolerance. Populations were the source population (C0) and two populations divergently selected for high (C4H) and low (C4L) cold tolerance. Such divergently selected popula- tions are expected to share most of their genes, with the main exception of those genes controlling cold tolerance. Arrhenius plots of PIMAC chloride fluxes showed a linear temperature dependence when seedlings were grown at 25 or 14C, whereas a non-linear temperature dependence was found when seedlings were grown at 5C, with or with- out acclimation at 14C. The activation energy and other thermodynamic parameters of PIMAC activity varied depending on temperature treatments during seedling growth. When seedlings were grown at 14 and 5C with acclimation, PIMAC activity of the C4H population increased, while that of C4L declined, as compared with the activities of seedlings grown at 25C. These symmetric responses indicate that PIMAC activity changes are asso- ciated with genetically determined differences in the cold tolerance level of the investigated populations. We conclude that anion fluxes by PIMAC depend upon changes on growth temperature and are differentially related to the tolerance level of the tested populations.

As a preliminary step to using gametophytic selection to obtain genotypes of maize, Zea mays L., differing for abscisic acid (ABA) content and/or sensitivity, the influence of ABA on the in vitro germination and tube growth of maize pollen and the presence of genetic variability for these traits was investigated. In a first experiment, pollen samples of three genotypes were germinated on substrates containing increasing amounts of ABA. Low ABA concentration (0.5 νM) significantly increased pollen germination and tube growth, while 500 νM ABA significantly decreased both traits, thus indicating that ABA can influence the growth of maize pollen in vitro. In a second experiment, eleven inbred lines were screened for the in vitro reactivity of their microgametophytes to ABA. The lines differed significantly for the response (% of control) of tube length, but not of pollen germination to ABA.

Four cycles of divergent full-sib recurrent selection for the ability to germinate at low temperature were conducted in a maize (Zea mays L.) F2 population. The selection criterion was the high (H) or the low (L) value in algebraic terms of the difference (DG) between germination percentage at 9.5°C (G9.5) detected 19 days after sowing and germination percentage at 25°C (G 25) seven days after sowing; both traits were evaluated in a controlled environment (germinator). Direct and correlated responses estimated during the course of selection were in accordance with those evaluated at the end. Selection for H led to populations with higher DG values, while the reverse was noted for L; differences between H and L populations increased in successive selection cycles, though divergence tended to level off. Selection for H also resulted in higher G 9.5 (day 19), shorter germination time and more flinty kernels, while selection for L led to responses in the opposite direction as well as to a lower G 9.5 detected 37 days after sowing (i.e. at the end of germination). In contrast, responses were negligible for G 25 and varied erratically from one cycle to another for kernel weight.

Although heterosis is widely exploited in agriculture, a clear understanding of its genetic bases is still elusive. This work describes the development of maize recombinant near-isogenic lines (NILs) for the mendelization of six heterotic QTL previously identified based on a maize (Zea mays L.) RIL population. The efficient and inexpensive strategy adopted to generate sets of NILs starting from QTL-specific residual heterozygous lines (RHLs) is described and validated. In particular, we produced nine pairs of recombinant NILs for all six QTL starting from RHLs F(4:5) originally obtained during the production of the RIL population mentioned above. Whenever possible, two different NIL pairs were generated for each QTL. The efficiency of this procedure was tested by analyzing two segregating populations for two of the selected heterotic QTL for plant height, yield per plant and ears per plant. Both additive and dominant effects were observed, consistently with the presence of the QTL within the introgressed regions. Refinement of QTL detection was consistent with previous observations in terms of effects and position of the considered QTL. The genetic material developed in this work represents the starting point for QTL fine mapping aimed at understanding the genetic bases of hybrid vigor in maize.

The evaluation of recombinant inbred lines (RILs) per se can be biased by inbreeding depression in case of allogamous species. To overcome this drawback, RILs can be evaluated in combination with testers; however, testers can carry dominant alleles at the quantitative trait loci (QTL), thus hampering their detection. This study was conducted on the maize (Zea mays L.) population of 142 RILs derived from the single cross B73 × H99 to evaluate the role of different testers in affecting: (1) QTL detection, (2) the estimates of their effects, and (3) the consistency of such estimates across testers. Testcrosses (TCs) were produced by crossing RILs with inbred testers B73 [TC(B)], H99 [TC(H)], and Mo17 [TC(M)]. TCs were field tested in three environments. TC(B) mean was higher than TC(H) mean for all traits, while TC(M) mean was the highest for plant vigor traits and grain yield. As to the number of detected QTL, tester Mo17 was superior to H99 and B73 for traits with prevailing additive effects. Several overlaps among the QTL were detected in two or all the three TC populations with QTL effects being almost always consistent (same sign). For traits with prevailing dominance–overdominance effects, as grain yield, the poor performing tester H99 was clearly the most effective; fewer overlaps were found and some of them were inconsistent (different sign). Epistatic interactions were of minor importance. In conclusion, the three testers proved to affect QTL detection and estimation of their effects, especially for traits showing high dominance levels.

The variation of response to acetochlor was studied in a two-year experiment carried out by subjecting 18 maize (Zea mays L.) inbred lines to three herbicide rates (0, 2.5 and 5 l a.i./ha). In both years some inbred lines consistently exhibited an evident susceptibility, with symptoms consisting of the seedling curling up below the soil surface and causing impaired field emergence. The results were poor plant density and lower grain yield in comparison to control. In contrast, other lines showed a satisfactory level of tolerance. Then, to gather data on the inheritance of response to acetochlor, four tolerant inbreds (T) and four susceptible inbreds (S) were crossed to obtain four T×T, four S×S, four S×T and the corresponding four T×S two-way hybrids. These hybrids were studied together with parental lines by applying the same herbicide rates used in the previous trial. The S×S hybrids showed susceptibility to the herbicide and the T×T were tolerant, whereas the S×T and the T× S hybrids showed a tolerance very close to that of the T×T hybrids. No difference was found between S×T and the corresponding T×S hybrids as to herbicide response. On average, the 16 hybrids exhibited greater tolerance than the eight parental lines, with each hybrid group being more tolerant than its parental line group. These results indicate that tolerance to acetochlor is prevailingly dominant, that action of extranuclear genes should be ruled out, and that the level of plant vigour can affect herbicide reactiveness.

The exploitation of heterosis is one of the most outstanding advancements in plant breeding, although its genetic basis is not well understood yet. This research was conducted on the materials arising from the maize single cross B73 × H99 to study heterosis by procedures of classical genetic and quantitative trait loci (QTL) analyses. Materials were the basic generations, the derived 142 recombinant inbred lines (RILs), and the three testcross populations obtained by crossing the 142 RILs to each parent and their F1. For seedling weight (SW), number of kernels per plant (NK), and grain yield (GY), heterosis was >100% and the average degree of dominance was >1. Epistasis was significant for SW and NK but not for GY. Several QTL were identified and in most cases they were in the additive–dominance range for traits with low heterosis and mostly in the dominance–overdominance range for plant height (PH), SW, NK, and GY. Only a few QTL with digenic epistasis were identified. The importance of dominance effects was confirmed by highly significant correlations between heterozygosity level and phenotypic performance, especially for GY. Some chromosome regions presented overlaps of overdominant QTL for SW, PH, NK, and GY, suggesting pleiotropic effects on overall plant vigor.

Molecular Breeding, 16 (4), ISSN:1380-3743, ISSN:1572-9788

Rhizobacteria (2808) were isolated on Pseudomonas-selective S1 medium from two maize inbred lines and from their hybrid at three plant growth stages. Positive phl D hybridization was found for 364 of them. The PhlD+ isolates were significantly more numerous in the rhizosphere of the hyrid than in those of parental lines. Furthermore, the frequency of PhlD+ was significantly higher for the hybrid at the flowering stage. An amplified rDNA restriction analysis showed that the hybrid genotype also increases the genetic diversity of PhlD+ populations when compared with its inbred parent lines, and this could be an effect of heterosis. Influence of the hybrid on the frequency and diversity of the bacterial PhlD+ population varied along the plant growth stage.

Theoretical and Applied Genetics, 109 (3), ISSN:0040-5752, ISSN:1432-2242

Maize pollen was exposed to the herbicide Chlorsulfuron (CS), and segregation for tolerance was observed. The resulting plant generation exhibited significantly greater tolerance to CS than other (control) progeny. Since the increase in tolerance occurred after only one generation of pollen exposure, this result demonstrates that pollen selection can be used to develop herbicide-resistant crop species, even when the species are not amenable to cell culture. It also suggests a possible mechanism for the rapid evolution of herbicide tolerance in weeds.

In order to evaluate the response at both the gametophytic and sporophytic level of a selection based on the pollen competitive ability and to compare its effects with those obtainable from a conventional sporophytic procedure, three recurrent selection plans were developed in maize starting from the same F2 population. Two gametophytic recurrent selection procedures at high (GSH) and low (GSL) selection intensity were performed by utilizing, to advance the populations, kernels taken from the base (GSH) or apex (GSL) of ears obtained from pair-crosses of randomly chosen plants. The third scheme was a sporophytic full-sib recurrent selection procedure (SS); the only selection criterion was the machine-harvestable grain yield of the families. In a sixyear period of selection, six cycles of both GSH and GSL and three cycles of SS were performed. The source and the selected populations (16 entries) were tested for pollen performance and for sporophytic traits. The selection cycles advanced through GSH showed a progressive increase, as compared to GSL, in pollen tube length measured at 4 h of in vitro culture. The SS cycles response was intermediate at 4 h whereas at 2 h it exceeded both GSH and GSL. A slight decrease in pollen diameter was evidenced in populations advanced with GSL procedure. The SS selection caused a marked increase for grain yield, lateness, leaves per plant and plant height. No response was shown by gametophytic selection for grain yield. The GSH procedure, however, led to an increase in kernel weight and to a decrease in kernel moisture, leaf number and plant height, as compared to GSL. Though gametophytic selection showed limited effects on sporophytic traits, it can be considered an efficient tool to supplement conventional sporophytic selection.

In higher plants pollen grain fertilization ability is a rather complex trait which depends on the rate of germination (i.e. germinability and-or germination time) and on the rate of tube growth. The expression of such components is in turn largely affected by the genotype of the stylar tissue, even in regard to compatible pollinations (for a review, see Heslop-Harrison, 1987).

Pollen selection can lead to correlated responses in the subsequent sporophytes because many genes are expressed in both haploid and diploid phases. This study was conducted to evaluate the effects of pollen selection for tolerance to the herbicide alachlor [2-chloro-N-(2,6-diethylphenyl)-N-(methoxymethyl) acetamide] in maize (Zea mays L.). Tassels of the single cross hybrid K55 (tolerant) x Ky226 (susceptible) were grown in absence (control) and in presence (selected) of alachlor (1 g L -1 ). Mature pollen from this process was used to make backcrosses to Ky226 (susceptibility being recessive). The selected population outperformed the control population (88 vs. 81%; P ≤ 0.01) for percentage of normal seedlings (i.e., not injured by alachlor) when compared in a field trial treated with the herbicide (5 kg a.i. ha -1 ). Pollen of plants grown without alachlor was analyzed in vitro; in presence of the herbicide in the medium (50 mg L -1 ) the selected population exceeded the control population for pollen tube length (334 vs. 308 μm; P ≤ 0.01). Two cycles of pollen selection were also conducted in the single cross hybrid K55 x H99 (susceptible) following the backcross procedure to the susceptible parent. Seeds of control and selected populations were exposed to alachlor (1 g L -1 ), and seedlings were grown in the greenhouse. For percentage of normal seedlings, we obtained a gain (16%) in the first cycle and an additional gain (10%) in the second cycle. The results indicate that genes controlling alachlor tolerance in maize are expressed in both haploid and diploid phases and that pollen selection can be effectively applied to improve the trait.

The first step to QTL cloning is the discovery of genetic loci linked to phenotypes of interest. Increasing the power, definition and speed by which such genetic loci are pinpointed on the genome is crucial for the success of this challenging task. New breeding designs emerged to address this need by increasing diversity and recombination events in QTL mapping panels. We discuss the MAGIC maize, a 1,636 recombinant inbred lines mapping panel we developed by intercrossing eight diverse Zea mays inbred lines in a balanced scheme. Each MAGIC recombinant inbred line is a unique mosaic of the diversity of the original founders, the population altogether collecting more than a hundred thousand recombination events. The increased diversity in elevated minor allele frequency permits the MAGIC maize to bridge the flexibility of association mapping approaches to the power of linkage mapping, speeding up the discovery of candidates for cloning. The MAGIC maize leverages transcriptomics data and full sequencing of the founder lines to reach single-gene definition. We genotyped and phenotyped a subset of the MAGIC maize and used it to show the general features of the population, identifying candidate genes for complex phenotypes. Mapping power simulations showed that the MAGIC maize may allow efficient QTL mapping even when few hundred lines are phenotyped, favoring its use in controlled conditions experiments. We discuss the perspectives that the MAGIC maize discloses in maize QTL mapping and cloning, and the further steps to exploit this resource, including the production of recombinant intercrosses for the study of heterosis.