Proteomic analysis of anthers from wild-type and photosensitive genetic male sterile mutant cotton (Gossypium hirsutum L.)

Background

Iron (Fe) deficiency symptoms in maize (Zea mays subsp. mays) express as leaf chlorosis, growth retardation, as well as yield reduction and are typically observed when plants grow in calcareous soils at alkaline pH. To improve our understanding of genotypical variability in the tolerance to Fe deficiency-induced chlorosis, the objectives of this study were to (i) determine the natural genetic variation of traits related to Fe homeostasis in the maize intermated B73 × Mo17 (IBM) population, (ii) to identify quantitative trait loci (QTLs) for these traits, and (iii) to analyze expression levels of genes known to be involved in Fe homeostasis as well as of candidate genes obtained from the QTL analysis.

Results

In hydroponically-grown maize, a total of 47 and 39 QTLs were detected for the traits recorded under limited and adequate supply of Fe, respectively.

Conclusions

From the QTL results, we were able to identify new putative candidate genes involved in Fe homeostasis under a deficient or adequate Fe nutritional status, like Ferredoxin class gene, putative ferredoxin PETF, metal tolerance protein MTP4, and MTP8. Furthermore, our expression analysis of candidate genes suggested the importance of trans-acting regulation for 2’-deoxymugineic acid synthase 1 (DMAS1), nicotianamine synthase (NAS3, NAS1), formate dehydrogenase 1 (FDH1), methylthioribose-1-phosphate isomerase (IDI2), aspartate/tyrosine/aromatic aminotransferase (IDI4), and methylthioribose kinase (MTK).     

Genome-Wide Association Implicates Candidate Genes Conferring Resistance to Maize Rough Dwarf Disease in Maize

Maize rough dwarf disease (MRDD) is a destructive viral disease in China, which results in 20–30% of the maize yield losses in affected areas and even as high as 100% in severely infected fields. Understanding the genetic basis of resistance will provide important insights for maize breeding program. In this study, a diverse maize population comprising of 527 inbred lines was evaluated in four environments and a genome-wide association study (GWAS) was undertaken with over 556000 SNP markers. Fifteen candidate genes associated with MRDD resistance were identified, including ten genes with annotated protein encoding functions. The homologous of nine candidate genes were predicted to relate to plant defense in different species based on published results. Significant correlation (R² = 0.79) between the MRDD severity and the number of resistance alleles was observed. Consequently, we have broadened the resistant germplasm to MRDD and identified a number of resistance alleles by GWAS. The results in present study also imply the candidate genes in defense pathway play an important role in resistance to MRDD in maize.     

Molecular evidence for phytosiderophore‐induced improvement of iron nutrition of peanut intercropped with maize in calcareous soil

Peanut/maize intercropping is a sustainable and effective agroecosystem that evidently enhances the Fe nutrition of peanuts in calcareous soils. So far, the mechanism involved in this process has not been elucidated. In this study, we unravel the effects of phytosiderophores in improving Fe nutrition of intercropped peanuts in peanut/maize intercropping. The maize ys3 mutant, which cannot release phytosiderophores, did not improve Fe nutrition of peanut, whereas the maize ys1 mutant, which can release phytosiderophores, prevented Fe deficiency, indicating an important role of phytosiderophores in improving the Fe nutrition of intercropped peanut. Hydroponic experiments were performed to simplify the intercropping system, which revealed that phytosiderophores released by Fe‐deficient wheat promoted Fe acquisition in nearby peanuts and thus improved their Fe nutrition. Moreover, the phytosiderophore deoxymugineic acid (DMA) was detected in the roots of intercropped peanuts. The yellow stripe1‐like (YSL) family of genes, which are homologous to maize yellow stripe 1 (ZmYS1), were identified in peanut roots. Further characterization indicated that among five AhYSL genes, AhYSL1, which was localized in the epidermis of peanut roots, transported Fe(III)–DMA. These results imply that in alkaline soil, Fe(III)–DMA dissolved by maize might be absorbed directly by neighbouring peanuts in the peanut/maize intercropping system.     

玉米叶形相关性状的Meta-QTL及候选基因分析

叶长、叶宽、叶面积及叶夹角不仅影响玉米(Zea mays)光合效率, 也是株型的重要构成因素。通过对620个叶形QTL进行整合, 构建不同遗传背景下的叶形QTL整合图谱, 利用元分析发掘出22个叶长、22个叶宽、12个叶面积以及17个叶夹角mQTL; 进一步运用生物信息学手段, 确定44个与叶片发育密切相关的候选基因。分析发现, 仅有NAL7-like、YABBY6- like和GRF2等13个基因位于mQTL区间内, 而玉米中已克隆的KNOTTED1、AN3/GIF1、rgd1/lbl1、mwp1及SRL2-like、HYL1-like和CYCB2;4-like等水稻(Oryza sativa)和拟南芥(Arabidopsis thaliana)叶形同源基因位于未被整合的QTL内; 对44个候选基因在叶片长、宽、厚发育过程中基部-末端、中央-边缘、远轴-近轴的调控机理进行归纳分析, 发现玉米中仅有少数几个候选基因被报道, 揭示了叶形发育的部分分子机理。因此, 对玉米叶形相关mQTL/QTL及基因进行全面深入的分析, 不仅有助于增加对其遗传结构的了解, 发掘更多候选基因, 阐明叶形发育和形成的分子机制, 还可为耐密理想株型的分子标记辅助选择提供依据。     

Pleiotropic effects of the duplicate maize FLORICAULA/LEAFY genes zfl1 and zfl2 on traits under selection during maize domestication

Phenotypic variation on which selection can act during evolution may be caused by variation in activity level of developmental regulatory genes. In many cases, however, such genes affect multiple traits. This situation can lead to co-evolution of traits, or evolutionary constraint if some pleiotropic effects are detrimental. Here, we present an analysis of quantitative traits associated with gene copy number of two important maize regulatory genes, the duplicate FLORICAULA/LEAFY orthologs zfl1 and zfl2. We found statistically significant associations between several quantitative traits and copy number of both zfl genes in several maize genetic backgrounds. Despite overlap in traits associated with these duplicate genes, zfl1 showed stronger associations with flowering time, while zfl2 associated more strongly with branching and inflorescence structure traits, suggesting some divergence of function. Since zfl2 associates with quantitative variation for ear rank and also maps near a quantitative trait locus (QTL) on chromosome 2 controlling ear rank differences between maize and teosinte, we tested whether zfl2 might have been involved in the evolution of this trait using a QTL complementation test. The results suggest that zfl2 activity is important for the QTL effect, supporting zfl2 as a candidate gene for a role in morphological evolution of maize.     

Maize grain concentrations and above-ground shoot acquisition of micronutrients as affected by intercropping with turnip, faba bean, chickpea, and soybean

Most research on micronutrients in maize has focused on maize grown as a monocrop. The aim of this study was to determine the effects of intercropping on the concentrations of micronutrients in maize grain and their acquisition via the shoot. We conducted field experiments to investigate the effects of intercropping with turnip (Brassica campestris L.), faba bean (Vicia faba L.), chickpea (Cicer arietinum L.), and soybean (Glycine max L.) on the iron (Fe), manganese (Mn), copper (Cu) and zinc (Zn) concentrations in the grain and their acquisition via the above-ground shoots of maize (Zea mays L.). Compared with monocropped maize grain, the grain of maize intercropped with legumes showed lower concentrations of Fe, Mn, Cu, and Zn and lower values of their corresponding harvest indexes. The micronutrient concentrations and harvest indexes in grain of maize intercropped with turnip were the same as those in monocropped maize grain. Intercropping stimulated the above-ground maize shoot acquisition of Fe, Mn, Cu and Zn, when averaged over different phosphorus (P) application rates. To our knowledge, this is the first report on the effects of intercropping on micronutrient concentrations in maize grain and on micronutrients acquisition via maize shoots (straw+grain). The maize grain Fe and Cu concentrations, but not Mn and Zn concentrations, were negatively correlated with maize grain yields. The concentrations of Fe, Mn, Cu, and Zn in maize grain were positively correlated with their corresponding harvest indexes. The decreased Fe, Mn, Cu, and Zn concentrations in grain of maize intercropped with legumes were attributed to reduced translocation of Fe, Mn, Cu, and Zn from vegetative tissues to grains. This may also be related to the delayed senescence of maize plants intercropped with legumes. We conclude that turnip/maize intercropping is beneficial to obtain high maize grain yield without decreased concentrations of Fe, Mn, Cu, and Zn in the grain. Further research is required to clarify the mechanisms underlying the changes in micronutrient concentrations in grain of intercropped maize.     

The role of regulatory genes during maize domestication: evidence from nucleotide polymorphism and gene expression

We investigated DNA sequence variation in 72 candidate genes in maize landraces and the wild ancestor of maize, teosinte. The candidate genes were chosen because they exhibit very low sequence diversity among maize inbreds and have sequence homology to known regulatory genes. We observed signatures of selection in 17 candidate genes, indicating that they were potential targets of artificial selection during domestication. In addition, 21 candidate genes were identified as potential targets of natural selection in teosinte. A comparison of the proportion of selected genes between our regulatory genes and genes unfiltered for their potential function (but also with very low sequence diversity among maize inbreds) provided some weak evidence that regulatory genes are overrepresented among selected genes. We detected no significant association between the positions of genes identified as potential targets of selection during domestication and quantitative trait loci (QTL) responsible for maize domestication traits. However, a subset of these genes, those identified by sequence homology as kinase/phosphatase genes, significantly cluster with the domestication QTL. We also analyzed expression profiles of genes in distinct maize tissues and observed that domestication genes are expressed on average at a significantly higher level than neutral genes in reproductive organs, including kernels.     

Fine mapping of Rscmv2, a major gene for resistance to sugarcane mosaic virus in maize

Sugarcane mosaic virus (SCMV) is one of the most devastating virus diseases in maize. In our previous study, two dominant complementary genes, Rscmv1 and Rscmv2, were identified in the resistant inbred line Siyi to confer resistance against SCMV. In this study, we report on the fine mapping of Rscmv2 and prediction of candidate genes. We developed a near-isogenic mapping population comprising 9,856 individuals segregating at Rscmv2, but not the Rscmv1 region. Combining the screening of the recombinants in the Rscmv2 region with 16 newly developed molecular markers, we fine-mapped Rscmv2 to an interval of 196.5?kb. Two overlapping bacterial artificial chromosomes covering this region were analyzed to identify putative candidate genes. Two predicted genes, encoding an auxin-binding protein and a Rho GTPase-activating protein, respectively, can be suggested as candidate genes for Rscmv2. The information from fine-mapping Rscmv2 can contribute to map-based cloning of this gene and molecular-assisted breeding in maize resistance breeding programs.     

A large-scale screen for artificial selection in maize identifies candidate agronomic loci for domestication and crop improvement

Maize (Zea mays subsp mays) was domesticated from teosinte (Z. mays subsp parviglumis) through a single domestication event in southern Mexico between 6000 and 9000 years ago. This domestication event resulted in the original maize landrace varieties, which were spread throughout the Americas by Native Americans and adapted to a wide range of environmental conditions. Starting with landraces, 20th century plant breeders selected inbred lines of maize for use in hybrid maize production. Both domestication and crop improvement involved selection of specific alleles at genes controlling key morphological and agronomic traits, resulting in reduced genetic diversity relative to unselected genes. Here, we sequenced 1095 maize genes from a sample of 14 inbred lines and chose 35 genes with zero sequence diversity as potential targets of selection. These 35 genes were then sequenced in a sample of diverse maize landraces and teosintes and tested for selection. Using two statistical tests, we identified eight candidate genes. Extended gene sequencing of these eight candidate loci confirmed that six were selected throughout the gene, and the remaining two exhibited evidence of selection in the 3' portion of each gene. The selected genes have functions consistent with agronomic selection for nutritional quality, maturity, and productivity. Our large-scale screen for artificial selection allows identification of genes of potential agronomic importance even when gene function and the phenotype of interest are unknown.     

Dissection of maize kernel composition and starch production by candidate gene association

Cereal starch production forms the basis of subsistence for much of the world's human and domesticated animal populations. Starch concentration and composition in the maize (Zea mays ssp mays) kernel are complex traits controlled by many genes. In this study, an association approach was used to evaluate six maize candidate genes involved in kernel starch biosynthesis: amylose extender1 (ae1), brittle endosperm2 (bt2), shrunken1 (sh1), sh2, sugary1, and waxy1. Major kernel composition traits, such as protein, oil, and starch concentration, were assessed as well as important starch composition quality traits, including pasting properties and amylose levels. Overall, bt2, sh1, and sh2 showed significant associations for kernel composition traits, whereas ae1 and sh2 showed significant associations for starch pasting properties. ae1 and sh1 both associated with amylose levels. Additionally, haplotype analysis of sh2 suggested this gene is involved in starch viscosity properties and amylose content. Despite starch concentration being only moderately heritable for this particular panel of diverse maize inbreds, high resolution was achieved when evaluating these starch candidate genes, and diverse alleles for breeding and further molecular analysis were identified.     

Identification of genetic variants associated with maize flowering time using an extremely large multi‐genetic background population

Flowering time is one of the major adaptive traits in domestication of maize and an important selection criterion in breeding. To detect more maize flowering time variants we evaluated flowering time traits using an extremely large multi‐ genetic background population that contained more than 8000 lines under multiple Sino‐United States environments. The population included two nested association mapping (NAM) panels and a natural association panel. Nearly 1 million single‐nucleotide polymorphisms (SNPs) were used in the analyses. Through the parallel linkage analysis of the two NAM panels, both common and unique flowering time regions were detected. Genome wide, a total of 90 flowering time regions were identified. One‐third of these regions were connected to traits associated with the environmental sensitivity of maize flowering time. The genome‐wide association study of the three panels identified nearly 1000 flowering time‐associated SNPs, mainly distributed around 220 candidate genes (within a distance of 1 Mb). Interestingly, two types of regions were significantly enriched for these associated SNPs – one was the candidate gene regions and the other was the approximately 5 kb regions away from the candidate genes. Moreover, the associated SNPs exhibited high accuracy for predicting flowering time.     

Identification and validation of genomic regions influencing kernel zinc and iron in maize

Key message

Genome-wide association study (GWAS) on 923 maize lines and validation in bi-parental populations identified significant genomic regions for kernel-Zinc and-Iron in maize.

Abstract

Bio-fortification of maize with elevated Zinc (Zn) and Iron (Fe) holds considerable promise for alleviating under-nutrition among the world’s poor. Bio-fortification through molecular breeding could be an economical strategy for developing nutritious maize, and hence in this study, we adopted GWAS to identify markers associated with high kernel-Zn and Fe in maize and subsequently validated marker-trait associations in independent bi-parental populations. For GWAS, we evaluated a diverse maize association mapping panel of 923 inbred lines across three environments and detected trait associations using high-density Single nucleotide polymorphism (SNPs) obtained through genotyping-by-sequencing. Phenotyping trials of the GWAS panel showed high heritability and moderate correlation between kernel-Zn and Fe concentrations. GWAS revealed a total of 46 SNPs (Zn-20 and Fe-26) significantly associated (P?≤?5.03?×?10^?05) with kernel-Zn and Fe concentrations with some of these associated SNPs located within previously reported QTL intervals for these traits. Three double-haploid (DH) populations were developed using lines identified from the panel that were contrasting for these micronutrients. The DH populations were phenotyped at two environments and were used for validating significant SNPs (P?≤?1?×?10^?03) based on single marker QTL analysis. Based on this analysis, 11 (Zn) and 11 (Fe) SNPs were found to have significant effect on the trait variance (P?≤?0.01, R²?≥?0.05) in at least one bi-parental population. These findings are being pursued in the kernel-Zn and Fe breeding program, and could hold great value in functional analysis and possible cloning of high-value genes for these traits in maize.