The agronomic performance of wheat doubled haploid lines derived from wheat x maize crosses

Summary The agronomic performance of 9 doubled haploid (DH) lines of Chinese Spring, 6 DH lines of Hope, 14 DH lines of the single chromosome substitution line Chinese Spring (Hope 5 A) and their respective parents was analyzed under field conditions. Seventeen Chinese Spring DH lines derived from wheat x Hordeum bulbosum crosses were also included for comparison. No significant variation was detected in either population of Chinese Spring DH lines and neither DH population differed from its parent. The Hope DH lines differed significantly for tiller biomass, spikelet number per ear, ear grain weight and 50-grain weight. However, all the variation could be attributed to the poor performance of only one line. Chinese Spring (Hope 5 A) DH lines showed significant variation for ear emergence time, but this was probably due to genetic heterogeneity in the parental stock. Overall, the results suggest that most DH lines produced by the wheat x maize method resemble their wheat parent, and that the variation induced in DH production is likely to be similar to that found in DHs from wheat x Hordeum bulbosum crosses.     

RFLP mapping of Brassica napus using doubled haploid lines

The combined use of doubled haploid lines and molecular markers can provide new genetic information for use in breeding programs. An F₁-derived doubled haploid (DH) population of Brassica napus obtained from a cross between an annual canola cultivar (Stellar) and a biennial rapeseed (Major) was used to construct a linkage map of 132 restriction fragment length polymorphism loci. The marker loci were arranged into 22 linkage groups and six pairs of linked loci covering 1016 cM. The DH map was compared to a partial map constructed with a common set of markers for an F₂ population derived from the same F₁ plant, and the overall maps were not significantly different. Comparisons of maps in Brassica species suggest that less recombination occurs in B. napus (n = 19) than expected from the combined map distances of the two hypothesized diploid progenitors, B. oleracea (n = 9) and B. rapa (n=10). A high percentage (32%) of segregating marker loci were duplicated in the DH map, and conserved linkage arrangements of some duplicated loci indicated possible intergenome homoeology in the amphidiploid or intragenome duplications from the diploid progenitors. Deviation from Mendelian segregation ratios (P < 0.05) was observed for 30% of the marker loci in the DH population and for 24% in the F₂ population. Deviation towards each parent occurred at equal frequencies in both populations and marker loci that showed deviation clustered in specific linkage groups. The DH lines and molecular marker map generated for this study can be used to map loci for agronomic traits segregating in this population. Present address Embrapa/Cenargen, C.P. 0.2372, CEP 70.770, Brasilia DF, Brazil     

Genotypic and phenotypic characterization of isogenic doubled haploid exotic introgression lines in maize

We characterized the genotypic and phenotypic variation in cell wall digestibility (CWD) and other agronomic traits of 50 backcross 1 generation doubled haploid (BC1DH) lines developed from the Germplasm Enhancement of Maize project. These lines were generated by introgressing 31 exotic unadapted maize races into PHZ51 and PHB47, temperate inbred lines with expired Plant Variety Protection. The 50 BC1DH lines and five check lines were genotyped with 199 single nucleotide polymorphism markers distributed across the genome. We identified, on average, 11.8% of markers with exotic donor parent alleles. This likely underestimates the proportion of donor introgressions, since we cannot discriminate monomorphic alleles from donor and recurrent parents. The potential roles of natural selection and the doubled haploid process in favouring selection of the recurrent genome are discussed. Although the proportion of donor parent genome was underestimated, donor fragments evaluated across the 50 BC1DH lines covered 92.9% of the recurrent parent genome. The evaluation of BC1DH lines for CWD revealed promising lines with CWD not differing significantly (???=?0.05) from forage quality lines used as checks. The introgression of exotic genome segments, however, was generally associated with higher ears, lodging, and late flowering. Even with limited power, our association analysis revealed quantitative trait polymorphisms associated with CWD, flowering date, and lodging.     

High-density linkage mapping of vitamin E content in maize grain

Vitamin E refers to eight distinct compounds collectively known as tocochromanols and can be further divided into two classes, tocotrienols and tocopherols. Tocochromanols are the major lipid-soluble antioxidants in maize (Zea mays L.) grain. Enhancing vitamin E content of maize through plant breeding has important implications for human and animal nutrition. Four inbred lines exhibiting unique variation for tocochromanol compounds were chosen from the Goodman maize diversity panel to construct two biparental mapping populations (N6xNC296 and E2558xCo125). The N6xNC296 population was developed to analyze segregation for α-tocopherol and α-tocotrienol content. The E2558WxCo125 population was developed to analyze segregation for the ratio of total tocotrienols to tocopherols. The tocochromanol variation in two replicates of each population was quantified using liquid chromatography-diode array detection. Using high-density linkage mapping, novel quantitative trait loci (QTL) in the N6xNC296 population were mapped using tocopherol ratio traits. These QTL contain the candidate gene homogentisate phytyltransferase (ZmVTE2) within the respective support intervals. This locus was not mapped in a previous genome-wide association study that analyzed tocochromanols in the Goodman diversity panel. Transgressive segregation was observed for γ- and α-tocochromanols in these populations, which facilitated QTL identification. These QTL and transgressive segregant families can be used in selection programs for vitamin E enhancement in maize. This work illustrates the complementary nature of biparental mapping populations and genome-wide association studies to further characterize genetic variation of tocochromanol content in maize grain.     

Comparative mapping of the Oregon Wolfe Barley using doubled haploid lines derived from female and male gametes

The Oregon Wolfe Barley mapping population is a resource for genetics research and instruction. Prior reports are based on a population of doubled haploid (DH) lines developed by the Hordeum bulbosum (H.b.) method, which samples female gametes. We developed new DH lines from the same cross using anther culture (A.C.), which samples male gametes. Linkage maps were generated in each of the two subpopulations using the same 1,328 single nucleotide polymorphism markers. The linkage maps based on DH lines derived from the products of megasporogeneis and microsporogenesis revealed minor differences in terms of estimated recombination rates. There were no differences in locus ordering. There was greater segregation distortion in the A.C.-derived subpopulation than in the H.b.-derived subpopulation, but in the region showing the greatest distortion, the cause was more likely allelic variation at the ZEO1 plant height locus rather than to DH production method. The effects of segregation distortion and pleiotropy had greater impacts on estimates of quantitative trait locus effect than population size for reproductive fitness traits assayed under greenhouse conditions. The Oregon Wolfe Barley (OWB) population and data are community resources. Seed is available from three distribution centers located in North America, Europe, and Asia. Details on ordering seed sets, as well as complete genotype and phenotype data files, are available at http://wheat.pw.usda.gov/ggpages/maps/OWB/.     

Genetic characterisation of dough rheological properties in a wheat doubled haploid population: additive genetic effects and epistatic interactions

Doubled haploid lines (n=160) from a cross between wheat cultivars Cranbrook (high dough extensibility) and Halberd (low dough extensibility) were grown at three Australian locations. The parents differ at all high- and low-molecular-weight glutenin loci. Dough rheological parameters were measured using small-scale testing procedures, and quantitative trait locus (QTL) mapping procedures were carried out using an existing well-saturated genetic linkage map for this cross. Genetic parameters were estimated using three software packages: QTLCartographer, Epistat and Genstat. Results indicated that environmental factors are a major determinant of dough extensibility across the three trial sites, whereas genotypic factors are the major determinants of dough strength. Composite interval mapping analysis across the 21 linkage groups revealed that as expected, the main additive QTLs for dough rheological properties are located at the high- and low-molecular-weight glutenin loci. A new QTL on chromosome 5A for M-extensibility (a mixograph-estimated measure of extensibility) was detected. Analysis of epistatic interactions revealed that there were significant conditional epistatic interactions related with the additive effects of glutenin loci on dough rheological properties. Therefore, the additive genetic effects of glutenins on dough rheological properties are conditional upon the genetic background of the wheat line. The molecular basis of the interactions with the glutenin loci may be via proteins that modify or alter the gluten protein matrix or variations in the expression level of the glutenin genes. Reverse-phase high performance liquid chromatography analysis of the molar number of individual glutenin subunits across the population showed that certain conditional epistases resulted in increased expression of the affected glutenin. The epistatic interactions detected in this study provide a possible explanation of the variable genetic effects of some glutenins on quality attributes in different genetic backgrounds and provide essential information for the accurate prediction of glutenin related variance in marker-assisted wheat breeding.     

Mapping QTLs with epistatic effects and QTL×environment interactions for plant height using a doubled haploid population in cultivated wheat

Quantitative trait loci (QTLs) for plant height in wheat (Triticum aestivum L.) were studied using a set of 168 doubled haploid (DH) lines, which were derived from the cross Huapei 3/Yumai 57. A genetic linkage map was constructed using 283 SSR and 22 EST-SSR markers. The DH population and the parents were evaluated for wheat plant height in 2005 and 2006 in Tai’an and 2006 in Suzhou. QTL analyses were performed using the software of QTLNetwork version 2.0 based on the mixed linear model. Four additive QTLs and five pairs of epistatic effects were detected, which were distributed on chromosomes 3A, 4B, 4D, 5A, 6A, 7B, and 7D. Among them, three additive QTLs and three pairs of epistatic QTLs showed QTL×environment interactions (QEs). Two major QTLs, Qph4B and Qph4D, which accounted for 14.51% and 20.22% of the phenotypic variation, were located similar to the reported locations of the dwarfing genes Rht1 and Rht2, respectively. The Qph3A-2 with additive effect was not reported in previous linkage mapping studies. The total QTL effects detected for the plant height explained 85.04% of the phenotypic variation, with additive effects 46.07%, epistatic effects 19.89%, and QEs 19.09%. The results showed that both additive effects and epistatic effects were important genetic bases of wheat plant height, which were subjected to environmental modifications, and caused dramatic changes in phenotypic effects. The information obtained in this study will be useful for manipulating the QTLs for wheat plant height by molecular marker-assisted selection (MAS).     

Tapping the genetic diversity of landraces in allogamous crops with doubled haploid lines: a case study from European flint maize

Key message

Using landraces for broadening the genetic base of elite maize germplasm is hampered by heterogeneity and high genetic load. Production of DH line libraries can help to overcome these problems.

Abstract

Landraces of maize (Zea mays L.) represent a huge reservoir of genetic diversity largely untapped by breeders. Genetic heterogeneity and a high genetic load hamper their use in hybrid breeding. Production of doubled haploid line libraries (DHL) by the in vivo haploid induction method promises to overcome these problems. To test this hypothesis, we compared the line per se performance of 389 doubled haploid (DH) lines across six DHL produced from European flint landraces with that of four flint founder lines (FFL) and 53 elite flint lines (EFL) for 16 agronomic traits evaluated in four locations. The genotypic variance (\(\sigma _{G}^{2}\)) within DHL was generally much larger than that among DHL and exceeded also \(\sigma _{G}^{2}\) of the EFL. For most traits, the means and \(\sigma _{G}^{2}\) differed considerably among the DHL, resulting in different expected selection gains. Mean grain yield of the EFL was 25 and 62% higher than for the FFL and DHL, respectively, indicating considerable breeding progress in the EFL and a remnant genetic load in the DHL. Usefulness of the best 20% lines was for individual DHL comparable to the EFL and grain yield (GY) in the top lines from both groups was similar. Our results corroborate the tremendous potential of landraces for broadening the narrow genetic base of elite germplasm. To make best use of these “gold reserves”, we propose a multi-stage selection approach with optimal allocation of resources to (1) choose the most promising landraces for DHL production and (2) identify the top DH lines for further breeding.

     

The production of haploid wheat plants from wheat x maize crosses

Summary Hybrid embryos from hexaploid wheat x maize crosses rapidly lose the maize chromosomes to produce haploid wheat embryos. Such embryos almost always aborted when left to develop on the plant, and only 1 was recovered from 2440 florets (0.17% of the expected number). Embryos had greater viability in spikelet culture, 47 (26.5% of the expected number) being recovered from 706 ovaries. Thirty-two of these embryos germinated to give green plants, 31 of which were haploid (21 wheat chromosomes) and 1 of which was euploid (42 wheat chromosomes). Spikelet culture enabled 17.1% of the expected number of embryos to be recovered as haploid plants, a 100-fold improvement on allowing embryos to develop in vivo. Ten haploid plants of Chinese Spring (kr1, kr2), 13 plants of Chinese Spring (Hope 5A) (kr1, Kr2), and 8 of Hope (Kr1, Kr2) were recovered. The potential of wheat x maize crosses for wheat haploid production and for gene transfer from maize to wheat is discussed.     

A green fluorescent protein-engineered haploid inducer line facilitates haploid mutant screens and doubled haploid breeding in maize

Northern corn leaf blight (NCLB) is a prevalent foliar disease in maize. Deployment of resistant cultivars is an effective way to control NCLB. In this study, 207 recombinant inbred lines derived from a K22 × By815 cross were planted in Yangling, China, in 2012 and 2013. NCLB score and lesion size were investigated after artificial inoculation. Significant phenotypic variation in NCLB resistance was observed in both years. Using a genetic map containing high-density single-nucleotide polymorphisms with average genetic distance of 0.74 cM, quantitative trait loci (QTL) for NCLB score and lesion size were analyzed. For NCLB score, four and three QTL were identified in 2012 and 2013, respectively. Two stable QTL were identified in both years. Of these, qNCLB5.04, located on chromosome 5 (bin 5.04), had the largest resistance effect, accounting for 19 and 20 % of the phenotypic variation in 2012 and 2013, respectively. For lesion size, six QTL were identified. Of these, one consensus QTL was associated with both lesion length and width, and the other five were associated only with lesion width. Among all QTL identified, only qNCLB5.04 was associated with both NCLB score and lesion size. Thus, our mapping results suggest that qNCLB5.04 could be a desirable target for marker-assisted selection for NCLB resistance in maize breeding programs.

     

Mapping QTL for grain yield, yield components, and spike features in a doubled haploid population of bread wheat

A doubled haploid (DH) population derived from a cross between the Japanese cultivar 'Fukuho-kumogi' and the Israeli wheat line 'Oligoculm' was used to map genome regions involved in the expression of grain yield, yield components, and spike features in wheat (Triticum aestivum L). A total of 371 markers (RAPD, SSR, RFLP, AFLP, and two morphological traits) were used to construct the linkage map that covered 4190 cM of wheat genome including 28 linkage groups. The results of composite interval mapping for all studied traits showed that some of the quantitative trait loci (QTL) were stable over experiments conducted in 2004 and 2005. The major QTL located in the Hair-Xpsp2999 interval on chromosome 1A controlled the expression of grains/spike (R(2) = 12.9% in 2004 and 22.4% in 2005), grain weight/spike (R(2) = 21.4% in 2004 and 15.8% in 2005), and spike number (R(2) = 15.6% in 2004 and 5.4% in 2005). The QTL for grain yield located on chromosomes 6A, 6B, and 6D totally accounted for 27.2% and 31.7% of total variation in this trait in 2004 and 2005, respectively. Alleles inherited from 'Oligoculm' increased the length of spikes and had decreasing effects on spike number. According to the data obtained in 2005, locus Xgwm261 was associated with a highly significant spike length QTL (R(2) = 42.33%) and also the major QTL for spikelet compactness (R(2) = 26.1%).     

In vitro production of haploid and doubled haploid plants from pollinated ovaries of maize (Zea mays)

Haploid induction has potential application for maize breeding. This paper reports that maize haploid plants have been induced by in vitro culture of pollinated ovaries. From a total of 26,400 cultured ovaries, 24 haploid plants were obtained and two of them were doubled after colchicine treatment. The maximum frequency of gynogenesis was 0.17% at 19.5 h post-pollination (HPP). The results showed that HPP was an important factor affecting plant induction from ovaries. Regenerated diploid R₀ plants were then subjected to genetic analysis using SSR molecular markers. One R₀ plant, whose progeny revealed a high level of homogeneity for several agro-morphological traits, was homozygous at 20 loci tested, with 11 showing paternal and 9 maternal banding pattern. This demonstrates that it is feasible to induce maize haploid plants by in vitro culture of pollinated ovaries.     

QTL mapping of yield-associated traits in Brassica juncea: meta-analysis and epistatic interactions using two different crosses between east European and Indian gene pool lines

Genetic analysis of 12 yield-associated traits was undertaken by dissection of quantitative trait loci (QTL) through meta-analysis and epistatic interaction studies in Brassica juncea. A consensus (integrated) map in B. juncea was constructed using two maps. These were VH map, developed earlier in the laboratory by using a DH population from the cross between Varuna and Heera (Pradhan et al. in Theor Appl Genet 106:607–614, 2003; Ramchiary et al. in Theor Appl Genet. 115:807–817, 2007; Panjabi et al. in BMC Genomics 9:113, 2008), and the TD map, developed in the present study using a DH population of 100 lines from the cross between TM-4 and Donskaja-IV. The TD map was constructed with 911 markers consisting of 585 AFLP, 8 SSR and 318 IP markers covering a total genome length of 1,629.9?cM. The consensus map constructed by using the common markers between the two maps contained a total of 2,662 markers and covered a total genome length of 1,927.1?cM. Firstly, QTL analysis of 12 yield-associated traits was undertaken for the TD population based on three-environment phenotypic data. Secondly, the three-environment phenotypic data for the same 12 quantitative traits generated by Ramchiary et al. (2007) were re-analyzed for the QTL detection in the VH map. Comparative analysis identified both common and population-specific QTL. The study revealed the presence of QTL clusters on LG A7, A8 and A10 in both TD and VH maps. Meta-analyses resolved 187 QTL distributed over nine linkage groups of TD and VH maps into 20 meta-QTL. Maximum resolution was recorded for the LG A10 wherein all the 54 QTL were mapped to a single meta-QTL within a confidence interval of 3.0?cM. Digenic epistatic interactions of QTL in both TD and VH maps revealed substantial additive?×?additive interactions showing a higher frequency of Type 1 and Type 2 interactions than Type 3 interactions. Some of the loci interacted with more than one locus indicating the presence of higher order epistatic interactions. These findings provided some detailed insight into the genetic architecture of the yield-associated traits in B. juncea.     

A genetic linkage map of Brassica carinata constructed with a doubled haploid population

Brassica carinata is an important oilseed crop with unique favourable traits that are desirable for other Brassica crops. However, given the limited research into genetic resources in B. carinata, knowledge of the genetic structure of this species is relatively poor. Nine homozygous, genetically distinct accessions of B. carinata were obtained via microspore culture, from which two divergent doubled haploid (DH) lines were used to develop a DH mapping population that consisted of 183 lines. The mapping population showed segregation of multiple traits of interest. A genetic map was constructed with PCR-based markers, and a total of 212 loci, which covered 1,703?cM, were assigned to eight linkage groups in the B genome and nine linkage groups in the C genome, which allowed comparison with genetic maps of other important Brassica species that contain the B/C genome(s). Loci for two Mendelian-inherited traits related to pigmentation (petal and anther tip colour) and one quantitative trait (seed coat colour) were identified using the linkage map. The significance of the mapping population in the context of genetic improvement of Brassica crops is discussed.     

Linkage Arrangement of RFLP loci in progenies from crosses between doubled haploid Asparagus officinalis L. clones

A preliminary genetic map of the dioecious species Asparagus officinalis L. (2n = 20) has been constructed on the basis of restriction fragment length polymorphism (RFLP) and isozyme marker data. With DNA samples digested with either EcoRI or HindIII 61 out of 148 probes (41%) identified RFLPs in six families of doubled haploid lines obtained through anther culture. A higher level of polymorphism (65%) was observed when a single family was screened for RFLPs using six distinct restriction enzymes. Segregation analysis of the BC progenies (40–80 individuals) resulted in a 418-cM extended map comprising 43 markers: 39 RFLPs, three isozymes and one morphological (sex). These markers are clustered in 12 linkage groups and four of them exhibited significant deviations from the expected 11 ratio. One isozyme and three RFLP markers were assigned to the sex chromosome.     

The mapping of epistatic effects onto a genealogical tree in haploid populations

In this paper we present a model that maps epistatic effects onto a genealogical tree for a haploid population. Prior work has demonstrated that genealogical structure causes the genotypic values of individuals to covary. Our results indicate that epistasis can reduce genotypic covariance that is caused by genealogical structure. Genotypic effects (both additive and epistatic) occur along the branches of a genealogical tree, from the base of the tree to its tips. Epistasis reduces genotypic covariance because there is a reweighting of the contribution of branches to the states of genotypes compared to the additive case. Branches near the tips of a genealogical tree contribute proportionally more genetic effects with epistasis than without epistasis. Epistatic effects are most numerous at basal positions in a genealogical tree when a population is constant in size and experiencing no selection, optimizing selection, diversifying selection or directional selection, indicating that epistatic effects are typically old. For a population that is growing in size, epistatic effects are most numerous at midpoints in a genealogical tree, indicating epistatic effects are of moderate age. Our results are important in that they suggest epistatic effects may typically explain deep (old) divergences and broad patterns of divergence that exist in populations, except in growing populations. In a growing population, epistatic effects may cause more within group divergence higher up in a tree and less between group divergence that is deep in a tree. The distribution of the number of epistatic effects and the expected variance and covariance in the number of epistatic effects is also provided assuming neutrality.     

Production of doubled haploid lines in frequencies sufficient for barley breeding programs

Winter barley (Hordeum vulgare L.) anthers were cultured on different liquid and on starch-solidified media. The optimal embryo and callus formation with different F₁-lines and the cv. Igri was obtained on a liquid medium with 20% Ficoll, 20 g/l maltose and barley starch. But the influence of the growth conditions of the donor plants and the genotypical differences are still enormous. The procedure has been optimized to such an extent that it can be used economically.     

QTL mapping with near-isogenic lines in maize

A set of 89 near-isogenic lines (NILs) of maize was created using marker-assisted selection. Nineteen genomic regions, identified by restriction fragment length polymorphism loci and chosen to represent portions of all ten maize chromosomes, were introgressed by backcrossing three generations from donor line Tx303 into the B73 genetic background. NILs were genotyped at an additional 128 simple sequence repeat loci to estimate the size of introgressions and the amount of background introgression. Tx303 introgressions ranged in size from 10 to 150 cM, with an average of 60 cM. Across all NILs, 89% of the Tx303 genome is represented in targeted and background introgressions. The average proportion of background introgression was 2.5% (range 0–15%), significantly lower than the expected value of 9.4% for third backcross generation lines developed without marker-assisted selection. The NILs were grown in replicated field evaluations in two years to map QTLs for flowering time traits. A parallel experiment of testcrosses of each NIL to the unrelated inbred, Mo17, was conducted in the same environments to map QTLs in NIL testcross hybrids. QTLs affecting days to anthesis, days to silking, and anthesis-silk interval were detected in both inbreds and hybrids in both environments. The testing environments differed dramatically for drought stress, and different sets of QTLs were detected across environments. Furthermore, QTLs detected in inbreds were typically different from QTLs detected in hybrids, demonstrating the genetic complexity of flowering time. NILs can serve as a valuable genetic mapping resource for maize breeders and geneticists. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Comparative mapping of the barley genome with male and female recombination-derived, doubled haploid populations

Male (anther culture) and female (Hordeum bulbosum) derived, doubled haploid populations were used to map the barley genome and thus determine the different recombination rates occurring during meiosis in the F1 hybrid donor plants. The anther culture-derived (male recombination) population showed an 18% overall increase in recombination rate. This increased recombination rate was observed for every chromosome and most of the chromosome arms. Examination of linkage distances between individual markers revealed eight segments with significantly higher recombination in the anther culture-derived population, and one in the Hordeum bulbosum-derived population. Very strong distortions of single locus segregations were observed in the anther culture-derived population, but map distances were not affected significantly by these distortions. There were 1.047 and 0.912 recombinations per chromosome in the anther culture and Hordeum bulbosum-derived doubled haploid populations, respectively.