QTL mapping and candidate genes for resistance to Fusarium ear rot and fumonisin contamination in maize.

BACKGROUND: Fusarium verticillioides is a common maize pathogen causing ear rot (FER) and contamination of the grains with the fumonisin B1 (FB1) mycotoxin. Resistance to FER and FB1 contamination are quantitative traits, affected by environmental conditions, and completely resistant maize genotypes to the pathogen are so far unknown. In order to uncover genomic regions associated to reduced FER and FB1 contamination and identify molecular markers for assisted selection, an F2:3 population of 188 progenies was developed crossing CO441 (resistant) and CO354 (susceptible) genotypes. FER severity and FB1 contamination content were evaluated over 2 years and sowing dates (early and late) in ears artificially inoculated with F. verticillioides by the use of either side-needle or toothpick inoculation techniques. RESULTS: Weather conditions significantly changed in the two phenotyping seasons and FER and FB1 content distribution significantly differed in the F3 progenies according to the year and the sowing time. Significant positive correlations (P < 0.01) were detected between FER and FB1 contamination, ranging from 0.72 to 0.81. A low positive correlation was determined between FB1 contamination and silking time (DTS). A genetic map was generated for the cross, based on 41 microsatellite markers and 342 single nucleotide polymorphisms (SNPs) derived from Genotyping-by-Sequencing (GBS). QTL analyses revealed 15 QTLs for FER, 17 QTLs for FB1 contamination and nine QTLs for DTS. Eight QTLs located on linkage group (LG) 1, 2, 3, 6, 7 and 9 were in common between FER and FB1, making possible the selection of genotypes with both low disease severity and low fumonisin contamination. Moreover, five QTLs on LGs 1, 2, 4, 5 and 9 located close to previously reported QTLs for resistance to other mycotoxigenic fungi. Finally, 24 candidate genes for resistance to F. verticillioides are proposed combining previous transcriptomic data with QTL mapping. CONCLUSIONS: This study identified a set of QTLs and candidate genes that could accelerate breeding for resistance of maize lines showing reduced disease severity and low mycotoxin contamination determined by F. verticillioides.

Genetic properties of the MAGIC maize population: a new platform for high definition QTL mapping in Zea mays.

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.

A unique mutation in a MYB gene cosegregates with the nectarine phenotype in peach.

Nectarines play a key role in peach industry; the fuzzless skin has implications for consumer acceptance. The peach/nectarine (G/g) trait was described as monogenic and previously mapped on chromosome 5. Here, the position of the G locus was delimited within a 1.1 cM interval (635 kb) based on linkage analysis of an F2 progeny from the cross 'Contender' (C, peach) x 'Ambra' (A, nectarine). Careful inspection of the genes annotated in the corresponding genomic sequence (Peach v1.0), coupled with variant discovery, led to the identification of MYB gene PpeMYB25 as a candidate for trichome formation on fruit skin. Analysis of genomic re-sequencing data from five peach/nectarine accessions pointed to the insertion of a LTR retroelement in exon 3 of the PpeMYB25 gene as the cause of the recessive glabrous phenotype. A functional marker (indelG) developed on the LTR insertion cosegregated with the trait in the CxA F2 progeny and was validated on a broad panel of genotypes, including all known putative donors of the nectarine trait. This marker was shown to efficiently discriminate between peach and nectarine plants, indicating that a unique mutational event gave rise to the nectarine trait and providing a useful diagnostic tool for early seedling selection in peach breeding programs.

Fine mapping and identification of a candidate gene for a major locus controlling maturity date in peach.

BACKGROUND: Maturity date (MD) is a crucial factor for marketing of fresh fruit, especially those with limited shelf-life such as peach (Prunus persica L. Batsch): selection of several cultivars with differing MD would be advantageous to cover and extend the marketing season. Aims of this work were the fine mapping and identification of candidate genes for the major maturity date locus previously identified on peach linkage group 4. To improve genetic resolution of the target locus two F2 populations derived from the crosses Contender x Ambra (CxA, 306 individuals) and PI91459 (NJ Weeping) x Bounty (WxBy, 103 individuals) were genotyped with the Sequenom and 9K Illumina Peach Chip SNP platforms, respectively. RESULTS: Recombinant individuals from the WxBy F2 population allowed the localisation of maturity date locus to a 220 kb region of the peach genome. Among the 25 annotated genes within this interval, functional classification identified ppa007577m and ppa008301m as the most likely candidates, both encoding transcription factors of the NAC (NAM/ATAF1, 2/CUC2) family. Re-sequencing of the four parents and comparison with the reference genome sequence uncovered a deletion of 232 bp in the upstream region of ppa007577m that is homozygous in NJ Weeping and heterozygous in Ambra, Bounty and the WxBy F1 parent. However, this variation did not segregate in the CxA F2 population being the CxA F1 parent homozygous for the reference allele. The second gene was thus examined as a candidate for maturity date. Re-sequencing of ppa008301m, showed an in-frame insertion of 9 bp in the last exon that co-segregated with the maturity date locus in both CxA and WxBy F2 populations. CONCLUSIONS: Using two different segregating populations, the map position of the maturity date locus was refined from 3.56 Mb to 220 kb. A sequence variant in the NAC gene ppa008301m was shown to co-segregate with the maturity date locus, suggesting this gene as a candidate controlling ripening time in peach. If confirmed on other genetic materials, this variant may be used for marker-assisted breeding of new cultivars with differing maturity date.

Extensive genomic characterization of a set of near-isogenic lines for heterotic QTL in maize (Zea mays L.).

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.

Characterization of heterotic quantitative trait loci in maize by evaluation of near-isogenic lines and their crosses at two competition levels.

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.

QTL detection in maize testcross progenies as affected by related and unrelated testers.

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 x 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.

Recombinant near-isogenic lines: a resource for the mendelization of heterotic QTL in maize.

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.

Classical genetic and quantitative trait loci analyses of heterosis in a maize hybrid between two elite inbred lines.

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 x 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 F(1). 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.

Origins, genetic organization and transcription of a family of non-autonomous helitron elements in maize.

Helitron transposable elements carrying gene fragments were recently discovered in maize. These elements are frequently specific to certain maize lineages. Here we report evidence supporting the involvement of helitrons in the rapid evolution of the maize genome, in particular in the multiplication of related genic fragments across the genome. We describe a family of four closely related, non-autonomous maize helitrons and their insertion sites at four non-allelic genetic loci across the maize genome: two specific to the B73 inbred, and two to the Mo17 inbred. We propose the phylogeny of this helitron family and provide an approximate timeline of their genomic insertions. One of these elements, the Mo17-specific helitron on chromosome 1 (bin 1.07), is transcriptionally active, probably as a result of insertion in the vicinity of a promoter. Significantly, it produces an alternatively spliced and chimeric transcript joining together genic segments of different chromosomal origin contained within the helitron. This transcript potentially encodes up to four open reading frames. During the course of evolution, transcribed helitrons containing multiple gene fragments may occasionally give rise to new genes with novel biochemical functions by a combinatorial assembly of exons. Thus helitrons not only constantly reshape the genomic organization of maize and profoundly affect its genetic diversity, but also may be involved in the evolution of gene function.

Gene duplication and exon shuffling by helitron-like transposons generate intraspecies diversity in maize.

We report a whole-genome comparison of gene content in allelic BAC contigs from two maize inbred lines. Genic content polymorphisms involve as many as 10,000 sequences and are mainly generated by DNA insertions. The termini of eight of the nine genic insertions that we analyzed shared the structural hallmarks of helitron rolling-circle transposons. DNA segments defined by helitron termini contained multiple gene-derived fragments and had a structure typical of nonautonomous helitron-like transposons. Closely related insertions were found in multiple genomic locations. Some of these produced transcripts containing segments of different genes, supporting the idea that these transposition events have a role in exon shuffling and the evolution of new proteins. We identified putative autonomous helitron elements and found evidence for their transcription. Helitrons in maize seem to continually produce new nonautonomous elements responsible for the duplicative insertion of gene segments into new locations and for the unprecedented genic diversity. The maize genome is in constant flux, as transposable elements continue to change both the genic and nongenic fractions of the genome, profoundly affecting genetic diversity.