RESUMO
KEY MESSAGE: We found metabolites, enzyme activities and enzyme transcript abundances vary significantly across the maize lifecycle, but weak correlation exists between the three groups. We identified putative genes regulating nitrate assimilation. Progress in improving nitrogen (N) use efficiency (NUE) of crop plants has been hampered by the complexity of the N uptake and utilisation systems. To understand this complexity we measured the activities of seven enzymes and ten metabolites related to N metabolism in the leaf and root tissues of Gaspe Flint maize plants grown in 0.5 or 2.5 mM NO3 (-) throughout the lifecycle. The amino acids had remarkably similar profiles across the lifecycle except for transient responses, which only appeared in the leaves for aspartate or in the roots for asparagine, serine and glycine. The activities of the enzymes for N assimilation were also coordinated to a certain degree, most noticeably with a peak in root activity late in the lifecycle, but with wide variation in the activity levels over the course of development. We analysed the transcriptional data for gene sets encoding the measured enzymes and found that, unlike the enzyme activities, transcript levels of the corresponding genes did not exhibit the same coordination across the lifecycle and were only weakly correlated with the levels of various amino acids or individual enzyme activities. We identified gene sets which were correlated with the enzyme activity profiles, including seven genes located within previously known quantitative trait loci for enzyme activities and hypothesise that these genes are important for the regulation of enzyme activities. This work provides insights into the complexity of the N assimilation system throughout development and identifies candidate regulatory genes, which warrant further investigation in efforts to improve NUE in crop plants.
Assuntos
Regulação da Expressão Gênica de Plantas , Nitrogênio/metabolismo , Zea mays/genética , Zea mays/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Locos de Características Quantitativas/genética , Zea mays/enzimologia , Zea mays/crescimento & desenvolvimentoRESUMO
The maize genome, with its large complement of transposons and repeats, is a paradigm for the study of epigenetic mechanisms such as paramutation and imprinting. Here, we present the genome-wide map of cytosine methylation for two maize inbred lines, B73 and Mo17. CG (65%) and CHG (50%) methylation (where H = A, C, or T) is highest in transposons, while CHH (5%) methylation is likely guided by 24-nt, but not 21-nt, small interfering RNAs (siRNAs). Correlations with methylation patterns suggest that CG methylation in exons (8%) may deter insertion of Mutator transposon insertion, while CHG methylation at splice acceptor sites may inhibit RNA splicing. Using the methylation map as a guide, we used low-coverage sequencing to show that parental methylation differences are inherited by recombinant inbred lines. However, frequent methylation switches, guided by siRNA, persist for up to eight generations, suggesting that epigenetic inheritance resembling paramutation is much more common than previously supposed. The methylation map will provide an invaluable resource for epigenetic studies in maize.
Assuntos
Metilação de DNA , DNA de Plantas/genética , Genoma de Planta , Sítios de Splice de RNA , RNA de Plantas/metabolismo , Zea mays/genética , Zea mays/metabolismo , Processamento Alternativo , Elementos de DNA Transponíveis , DNA de Plantas/metabolismo , Epigênese Genética , Éxons , Regulação da Expressão Gênica de Plantas , Impressão Genômica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Análise de SequênciaRESUMO
Elucidation of the gene networks underlying the response to N supply and demand will facilitate the improvement of the N uptake efficiency of plants. We undertook a transcriptomic analysis of maize to identify genes responding to both a non-growth-limiting decrease in NO3- provision and to development-based N demand changes at seven representative points across the life cycle. Gene co-expression networks were derived by cluster analysis of the transcript profiles. The majority of NO3--responsive transcription occurred at 11 (D11), 18 (D18) and 29 (D29) days after emergence, with differential expression predominating in the root at D11 and D29 and in the leaf at D18. A cluster of 98 probe sets was identified, the expression pattern of which is similar to that of the high-affinity NO3- transporter (NRT2) genes across the life cycle. The cluster is enriched with genes encoding enzymes and proteins of lipid metabolism and transport, respectively. These are candidate genes for the response of maize to N supply and demand. Only a few patterns of differential gene expression were observed over the entire life cycle; however, the composition of the classes of the genes differentially regulated at individual time points was unique, suggesting tightly controlled regulation of NO3--responsive gene expression.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Nitratos/farmacologia , Transcrição Gênica/efeitos dos fármacos , Zea mays/crescimento & desenvolvimento , Zea mays/genética , Análise por Conglomerados , Perfilação da Expressão Gênica , Ontologia Genética , Redes Reguladoras de Genes/efeitos dos fármacos , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Tempo , Zea mays/efeitos dos fármacosRESUMO
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.
Assuntos
Elementos de DNA Transponíveis/genética , Éxons/genética , Duplicação Gênica , Variação Genética , Zea mays/genética , Sequência de Bases , DNA de Plantas/química , Genes de Plantas/genética , Genoma de Planta , Dados de Sequência Molecular , Especificidade da EspécieRESUMO
An understanding of nitrate (NO3-) uptake throughout the lifecycle of plants, and how this process responds to nitrogen (N) availability, is an important step towards the development of plants with improved nitrogen use efficiency (NUE). NO3- uptake capacity and transcript levels of putative high- and low-affinity NO3- transporters (NRTs) were profiled across the lifecycle of dwarf maize (Zea mays) plants grown at reduced and adequate NO3-. Plants showed major changes in high-affinity NO3- uptake capacity across the lifecycle, which varied with changing relative growth rates of roots and shoots. Transcript abundances of putative high-affinity NRTs (predominantly ZmNRT2.1 and ZmNRT2.2) were correlated with two distinct peaks in high-affinity root NO3- uptake capacity and also N availability. The reduction in NO3- supply during the lifecycle led to a dramatic increase in NO3- uptake capacity, which preceded changes in transcript levels of NRTs, suggesting a model with short-term post-translational regulation and longer term transcriptional regulation of NO3- uptake capacity. These observations offer new insight into the control of NO3- uptake by both plant developmental processes and N availability, and identify key control points that may be targeted by future plant improvement programmes to enhance N uptake relative to availability and/or demand.
Assuntos
Nitratos/metabolismo , Nitrogênio/farmacologia , Zea mays/crescimento & desenvolvimento , Zea mays/metabolismo , Aminoácidos/metabolismo , Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Transporte Biológico/efeitos dos fármacos , Transporte Biológico/genética , Biomassa , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Transportadores de Nitrato , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Zea mays/efeitos dos fármacos , Zea mays/genéticaRESUMO
BACKGROUND: Endo-(1,4)-ß-glucanase (cellulase) glycosyl hydrolase GH9 enzymes have been implicated in several aspects of cell wall metabolism in higher plants, including cellulose biosynthesis and degradation, modification of other wall polysaccharides that contain contiguous (1,4)-ß-glucosyl residues, and wall loosening during cell elongation. RESULTS: The endo-(1,4)-ß-glucanase gene families from barley (Hordeum vulgare), maize (Zea mays), sorghum (Sorghum bicolor), rice (Oryza sativa) and Brachypodium (Brachypodium distachyon) range in size from 23 to 29 members. Phylogenetic analyses show variations in clade structure between the grasses and Arabidopsis, and indicate differential gene loss and gain during evolution. Map positions and comparative studies of gene structures allow orthologous genes in the five species to be identified and synteny between the grasses is found to be high. It is also possible to differentiate between homoeologues resulting from ancient polyploidizations of the maize genome. Transcript analyses using microarray, massively parallel signature sequencing and quantitative PCR data for barley, rice and maize indicate that certain members of the endo-(1,4)-ß-glucanase gene family are transcribed across a wide range of tissues, while others are specifically transcribed in particular tissues. There are strong correlations between transcript levels of several members of the endo-(1,4)-ß-glucanase family and the data suggest that evolutionary conservation of transcription exists between orthologues across the grass family. There are also strong correlations between certain members of the endo-(1,4)-ß-glucanase family and other genes known to be involved in cell wall loosening and cell expansion, such as expansins and xyloglucan endotransglycosylases. CONCLUSIONS: The identification of these groups of genes will now allow us to test hypotheses regarding their functions and joint participation in wall synthesis, re-modelling and degradation, together with their potential role in lignocellulose conversion during biofuel production from grasses and cereal crop residues.
Assuntos
Celulase/genética , Família Multigênica , Filogenia , Poaceae/genética , Parede Celular/enzimologia , Parede Celular/genética , Bases de Dados Genéticas , Evolução Molecular , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Funções Verossimilhança , Análise de Sequência com Séries de Oligonucleotídeos , Poaceae/enzimologia , Transcrição GênicaRESUMO
DNA polymorphisms such as insertion/deletions and duplications affecting genome segments larger than 1 kb are known as copy-number variations (CNVs) or structural variations (SVs). They have been recently studied in animals and humans by using array-comparative genome hybridization (aCGH), and have been associated with several human diseases. Their presence and phenotypic effects in plants have not been investigated on a genomic scale, although individual structural variations affecting traits have been described. We used aCGH to investigate the presence of CNVs in maize by comparing the genome of 13 maize inbred lines to B73. Analysis of hybridization signal ratios of 60,472 60-mer oligonucleotide probes between inbreds in relation to their location in the reference genome (B73) allowed us to identify clusters of probes that deviated from the ratio expected for equal copy-numbers. We found CNVs distributed along the maize genome in all chromosome arms. They occur with appreciable frequency in different germplasm subgroups, suggesting ancient origin. Validation of several CNV regions showed both insertion/deletions and copy-number differences. The nature of CNVs detected suggests CNVs might have a considerable impact on plant phenotypes, including disease response and heterosis.
Assuntos
Genoma de Planta , Zea mays/genética , Alelos , Hibridização Genômica Comparativa , Dosagem de Genes , Endogamia , Análise de Sequência com Séries de OligonucleotídeosRESUMO
Maize is an important crop with a high level of genome diversity and heterosis. The genome sequence of a typical female line, B73, was previously released. Here, we report a de novo genome assembly of a corresponding male representative line, Mo17. More than 96.4% of the 2,183 Mb assembled genome can be accounted for by 362 scaffolds in ten pseudochromosomes with 38,620 annotated protein-coding genes. Comparative analysis revealed large gene-order and gene structural variations: approximately 10% of the annotated genes were mutually nonsyntenic, and more than 20% of the predicted genes had either large-effect mutations or large structural variations, which might cause considerable protein divergence between the two inbred lines. Our study provides a high-quality reference-genome sequence of an important maize germplasm, and the intraspecific gene order and gene structural variations identified should have implications for heterosis and genome evolution.
Assuntos
Ordem dos Genes/genética , Variação Genética/genética , Genoma de Planta/genética , Zea mays/genética , Cromossomos de Plantas/genética , Regulação da Expressão Gênica de Plantas/genética , Vigor Híbrido/genéticaRESUMO
Two species with genomes of almost identical size, maize and human, have different evolutionary histories, and as a result their genomes differ greatly in their content of retroelements, average size of the genes and amount of genetic diversity. However, there are also significant similarities: they both have undergone bottlenecks during their recent history and seem to have non-uniform distribution of recombination events. The human genome has been shown to contain large linkage blocks characterized by a limited number of haplotypes. A similar linkage block structure is likely to exist in maize. Although highly diverse maize populations show rapid decline of linkage disequilibrium, as in humans, it is possible to define populations with strong linkage disequilibrium, suitable for whole-genome scan association mapping. The genetic diversity and lack of sequence homology found in maize influences recombinational properties and local linkage disequilibrium levels but also challenges our understanding of the relationship between the genome sequence and species definition.
Assuntos
Frequência do Gene/genética , Genoma , Desequilíbrio de Ligação/genética , Zea mays/genética , Alelos , Variação Genética/genética , Genética Populacional , Humanos , Recombinação Genética/genéticaRESUMO
The discovery of single nucleotide polymorphisms (SNPs) and insertions/deletions, which are the basis of most differences between alleles, has been simplified by recent developments in sequencing technology. SNP discovery in many crop species, such as corn and soybean, is relatively straightforward because of their high level of intraspecific nucleotide diversity, and the availability of many gene and expressed sequence tag (EST) sequences. For these species, direct readout of SNP haplotypes is possible. Haplotype-based analysis is more informative than analysis based on individual SNPs, and has more power in analyzing association with phenotypes. The elite germplasm of some crops may have been subjected to bottlenecks relatively recently, increasing the amount of linkage disequilibrium (LD) present and facilitating the association of SNP haplotypes at candidate gene loci with phenotypes. Whole-genome scans may help identify genome regions that are associated with interesting phenotypes if sufficient LD is present. Technological improvements make the use of SNP and indel markers attractive for high-throughput use in marker-assisted breeding, EST mapping and the integration of genetic and physical maps.
Assuntos
Produtos Agrícolas/genética , Polimorfismo de Nucleotídeo Único/genética , Sequência de Bases , Etiquetas de Sequências Expressas , Marcadores Genéticos , Haplótipos/genética , Desequilíbrio de Ligação/genética , Mapeamento Físico do Cromossomo/métodos , Alinhamento de Sequência , Homologia de Sequência do Ácido NucleicoRESUMO
Cellulose is a fundamentally important component of cell walls of higher plants. It provides a scaffold that allows the development and growth of the plant to occur in an ordered fashion. Cellulose also provides mechanical strength, which is crucial for both normal development and to enable the plant to withstand both abiotic and biotic stresses. We quantified the cellulose concentration in the culm of 288 two--rowed and 288 six--rowed spring type barley accessions that were part of the USDA funded barley Coordinated Agricultural Project (CAP) program in the USA. When the population structure of these accessions was analysed we identified six distinct populations, four of which we considered to be comprised of a sufficient number of accessions to be suitable for genome-wide association studies (GWAS). These lines had been genotyped with 3072 SNPs so we combined the trait and genetic data to carry out GWAS. The analysis allowed us to identify regions of the genome containing significant associations between molecular markers and cellulose concentration data, including one region cross-validated in multiple populations. To identify candidate genes we assembled the gene content of these regions and used these to query a comprehensive RNA-seq based gene expression atlas. This provided us with gene annotations and associated expression data across multiple tissues, which allowed us to formulate a supported list of candidate genes that regulate cellulose biosynthesis. Several regions identified by our analysis contain genes that are co-expressed with cellulose synthase A (HvCesA) across a range of tissues and developmental stages. These genes are involved in both primary and secondary cell wall development. In addition, genes that have been previously linked with cellulose synthesis by biochemical methods, such as HvCOBRA, a gene of unknown function, were also associated with cellulose levels in the association panel. Our analyses provide new insights into the genes that contribute to cellulose content in cereal culms and to a greater understanding of the interactions between them.
Assuntos
Celulose/genética , Estudo de Associação Genômica Ampla , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Hordeum/genética , Família Multigênica , Agricultura , Parede Celular/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Marcadores Genéticos , Genética Populacional , Genótipo , Glicosídeo Hidrolases/metabolismo , Desequilíbrio de Ligação , Fenótipo , Locos de Características Quantitativas , Análise de Sequência de RNA , Estados UnidosRESUMO
BACKGROUND: Recent studies of ancestral maize populations indicate that linkage disequilibrium tends to dissipate rapidly, sometimes within 100 bp. We set out to examine the linkage disequilibrium and diversity in maize elite inbred lines, which have been subject to population bottlenecks and intense selection by breeders. Such population events are expected to increase the amount of linkage disequilibrium, but reduce diversity. The results of this study will inform the design of genetic association studies. RESULTS: We examined the frequency and distribution of DNA polymorphisms at 18 maize genes in 36 maize inbreds, chosen to represent most of the genetic diversity in U.S. elite maize breeding pool. The frequency of nucleotide changes is high, on average one polymorphism per 31 bp in non-coding regions and 1 polymorphism per 124 bp in coding regions. Insertions and deletions are frequent in non-coding regions (1 per 85 bp), but rare in coding regions. A small number (2-8) of distinct and highly diverse haplotypes can be distinguished at all loci examined. Within genes, SNP loci comprising the haplotypes are in linkage disequilibrium with each other. CONCLUSIONS: No decline of linkage disequilibrium within a few hundred base pairs was found in the elite maize germplasm. This finding, as well as the small number of haplotypes, relative to neutral expectation, is consistent with the effects of breeding-induced bottlenecks and selection on the elite germplasm pool. The genetic distance between haplotypes is large, indicative of an ancient gene pool and of possible interspecific hybridization events in maize ancestry.
Assuntos
Frequência do Gene/genética , Haplótipos/genética , Desequilíbrio de Ligação/genética , Polimorfismo de Nucleotídeo Único/genética , Zea mays/genética , Alelos , Animais , DNA de Plantas/genética , Genes de Plantas/genética , Marcadores Genéticos/genética , Variação Genética/genética , Genótipo , Camundongos , Dados de Sequência Molecular , Recombinação Genética/genética , Análise de Sequência de DNARESUMO
BACKGROUND: Plant height is an important agronomic trait that affects yield and tolerance to certain abiotic stresses. Understanding the genetic control of plant height is important for elucidating the regulation of maize development and has practical implications for trait improvement in plant breeding. METHODOLOGY/PRINCIPAL FINDINGS: In this study, two independent, semi-dwarf maize EMS mutants, referred to as dwarf & irregular leaf (dil1), were isolated and confirmed to be allelic. In comparison to wild type plants, the mutant plants have shorter internodes, shorter, wider and wrinkled leaves, as well as smaller leaf angles. Cytological analysis indicated that the leaf epidermal cells and internode parenchyma cells are irregular in shape and are arranged in a more random fashion, and the mutants have disrupted leaf epidermal patterning. In addition, parenchyma cells in the dil1 mutants are significantly smaller than those in wild-type plants. The dil1 mutation was mapped on the long arm of chromosome 6 and a candidate gene, annotated as an AP2 transcription factor-like, was identified through positional cloning. Point mutations near exon-intron junctions were identified in both dil1 alleles, resulting in mis-spliced variants. CONCLUSION: An AP2 transcription factor-like gene involved in stalk and leaf development in maize has been identified. Mutations near exon-intron junctions of the AP2 gene give mis-spliced transcript variants, which result in shorter internodes and wrinkled leaves.
Assuntos
Genes de Plantas/genética , Fenótipo , Epiderme Vegetal/metabolismo , Folhas de Planta/crescimento & desenvolvimento , Fator de Transcrição AP-2/genética , Zea mays/genética , Mapeamento Cromossômico , Clonagem Molecular , Primers do DNA/genética , Metanossulfonato de Etila , Hibridização In Situ , Mutagênese , Mutação/genética , Epiderme Vegetal/citologia , Folhas de Planta/anatomia & histologia , Folhas de Planta/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Zea mays/crescimento & desenvolvimentoRESUMO
We used whole genome scan association mapping to identify loci with major effect on oleic acid content in maize kernels. Single nucleotide polymorphism haplotypes at 8,590 loci were tested for association with oleic acid content in 553 maize inbreds. A single locus with major effect on oleic acid was mapped between 380 and 384 cM in the IBM2 neighbors genetic map on chromosome 4 and confirmed in a biparental population. A fatty acid desaturase, fad2, identified approximately 2 kb from the associated genetic marker, is the most likely candidate gene responsible for the differences in the phenotype. The fad2 alleles with high- and low-oleic acid content were sequenced and allelic differences in fad2 RNA level in developing embryos was investigated. We propose that a non-conservative amino acid polymorphism near the active site of fad2 contributes to the effect on oleic acid content. This is the first report of the use of a high resolution whole genome scan association mapping where a putative gene responsible for a quantitative trait was identified in plants.
Assuntos
Ácidos Graxos Dessaturases/genética , Ácidos Graxos Dessaturases/metabolismo , Ácido Oleico/metabolismo , Zea mays/genética , Zea mays/metabolismo , Alelos , Mapeamento Cromossômico , DNA de Plantas/genética , Expressão Gênica , Variação Genética , Genoma de Planta , Dados de Sequência Molecular , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismoRESUMO
A spontaneous maize mutant, brittle stalk-2 (bk2-ref), exhibits dramatically reduced tissue mechanical strength. Reduction in mechanical strength in the stalk tissue was highly correlated with a reduction in the amount of cellulose and an uneven deposition of secondary cell wall material in the subepidermal and perivascular sclerenchyma fibers. Cell wall accounted for two-thirds of the observed reduction in dry matter content per unit length of the mutant stalk in comparison to the wildtype stalk. Although the cell wall composition was significantly altered in the mutant in comparison to the wildtype stalks, no compensation by lignin and cell wall matrix for reduced cellulose amount was observed. We demonstrate that Bk2 encodes a Cobra-like protein that is homologous to the rice Bc1 protein. In the bk2-ref gene, a 1 kb transposon-like element is inserted in the beginning of the second exon, disrupting the open reading frame. The Bk2 gene was expressed in the stalk, husk, root, and leaf tissues, but not in the embryo, endosperm, pollen, silk, or other tissues with comparatively few or no secondary cell wall containing cells. The highest expression was in the isolated vascular bundles. In agreement with its role in secondary wall formation, the expression pattern of the Bk2 gene was very similar to that of the ZmCesA10, ZmCesA11, and ZmCesA12 genes, which are known to be involved in secondary wall formation. We have isolated an independent Mutator-tagged allele of bk2, referred to as bk2-Mu7, the phenotype of which is similar to that of the spontaneous mutant. Our results demonstrate that mutations in the Bk2 gene affect stalk strength in maize by interfering with the deposition of cellulose in the secondary cell wall in fiber cells.
Assuntos
Parede Celular/metabolismo , Proteínas de Plantas/fisiologia , Zea mays/fisiologia , Fenômenos Biomecânicos , Parede Celular/fisiologia , Elementos de DNA Transponíveis , Éxons , Expressão Gênica , Genes de Plantas , Glicosilfosfatidilinositóis/metabolismo , Mutação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Zea mays/genética , Zea mays/metabolismoRESUMO
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.
Assuntos
Cromossomos de Plantas , Elementos de DNA Transponíveis , Família Multigênica , Proteínas de Plantas/genética , Transcrição Gênica , Zea mays/genética , Sequência de Bases , Mapeamento Cromossômico , Evolução Molecular , Regulação da Expressão Gênica de Plantas , Dados de Sequência Molecular , Homologia de Sequência do Ácido NucleicoRESUMO
Allelic chromosomal regions totaling more than 2.8 Mb and located on maize (Zea mays) chromosomes 1L, 2S, 7L, and 9S have been sequenced and compared over distances of 100 to 350 kb between the two maize inbred lines Mo17 and B73. The alleles contain extended regions of nonhomology. On average, more than 50% of the compared sequence is noncolinear, mainly because of the insertion of large numbers of long terminal repeat (LTR)-retrotransposons. Only 27 LTR-retroelements are shared between alleles, whereas 62 are allele specific. The insertion of LTR-retrotransposons into the maize genome is statistically more recent for nonshared than shared ones. Most surprisingly, more than one-third of the genes (27/72) are absent in one of the inbreds at the loci examined. Such nonshared genes usually appear to be truncated and form clusters in which they are oriented in the same direction. However, the nonshared genome segments are gene-poor, relative to regions shared by both inbreds, with up to 12-fold difference in gene density. By contrast, miniature inverted terminal repeats (MITEs) occur at a similar frequency in the shared and nonshared fractions. Many times, MITES are present in an identical position in both LTRs of a retroelement, indicating that their insertion occurred before the replication of the retroelement in question. Maize ESTs and/or maize massively parallel signature sequencing tags were identified for the majority of the nonshared genes or homologs of them. In contrast with shared genes, which are usually conserved in gene order and location relative to rice (Oryza sativa), nonshared genes violate the maize colinearity with rice. Based on this, insertion by a yet unknown mechanism, rather than deletion events, seems to be the origin of the nonshared genes. The intergenic space between conserved genes is enlarged up to sixfold in maize compared with rice. Frequently, retroelement insertions create a different sequence environment adjacent to conserved genes.
Assuntos
DNA de Plantas/genética , Evolução Molecular , Genoma de Planta , Zea mays/genética , Alelos , Cruzamento , Mapeamento Cromossômico , Cromossomos Artificiais Bacterianos , Cromossomos de Plantas/genética , DNA de Plantas/química , Dados de Sequência Molecular , Família Multigênica , Oryza/genética , Proteínas de Plantas/genética , Retroelementos/genética , Análise de Sequência de DNA , Sequências Repetidas Terminais/genéticaRESUMO
We describe an effective systematic approach to genetic mapping of cDNA clones, including those obtained from EST sequencing. The EST of interest is first partially sequenced from the 3'-end. PCR primers which bracket the 3'-UTR segment of the cDNA are designed. The corresponding gene segment is amplified from the parents of the mapping population, using primers equipped with 3'- and 5'-extensions to facilitate direct sequencing of PCR products. Comparison of the sequences obtained from the mapping parents frequently reveals single nucleotide polymorphisms or insertion / deletion polymorphisms, which can then be genotyped in a mapping population. The genotyping of SNPs is performed by pyrosequencing, a sequencing-by-synthesis method that has been used successfully in SNP diagnostics. SNP analysis of up to 96 samples, a number required to produce meaningful genetic segregation data, can be rapidly accomplished in parallel. The parental genotype of three loci, stearoyl-ACP desaturase, nucleoside-diphosphate kinase and sucrose synthetase-1 were determined by conventional sequencing, and the polymorphism so identified were scored by the pyrosequencing of 94 individuals of a maize recombinant-inbred population. These loci were successfully placed onto chromosomes 3, 7 and 9 respectively. This method is generally applicable to most plant species, which show sufficient sequence diversity in the 3'-UTR region of genes.
Assuntos
Mapeamento Cromossômico/métodos , Plantas/genética , Regiões 3' não Traduzidas , Sequência de Bases , Primers do DNA/genética , DNA Complementar/genética , DNA de Plantas/genética , Etiquetas de Sequências Expressas , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA/métodos , Zea mays/genéticaRESUMO
Both yellow and white corn occurs among ancestral open pollinated varieties. More recently, breeders have selected yellow endosperm variants of maize over ancestral white phenotypes for their increased nutritional value resulting from the up-regulation of the Y1 phytoene synthase gene product in endosperm tissue. As a result, diversity within yellow maize lines at the Y1 gene is dramatically decreased as compared to white corn. We analyzed patterns of sequence diversity and linkage disequilibrium in nine low copy regions located at varying distances from the Y1 gene, including a homolog of the barley Mlo gene. Patterns consistent with a selective sweep, such as significant associations of informative single-nucleotide polymorphisms with endosperm color phenotype, linkage disequilibrium, and significantly reduced diversity within the yellow endosperm haplotypes, were observed up to 600 kb downstream of Y1, whereas the upstream region showed a more rapid recovery. The starch branching enzyme 1 (sbe1) gene is the first region downstream of Y1 that does not have a highly conserved haplotype in the yellow endosperm germplasm.