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1.
Nucleic Acids Res ; 47(14): 7247-7261, 2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31265077

RESUMO

Scaffold/matrix attachment regions (S/MARs) are DNA elements that serve to compartmentalize the chromatin into structural and functional domains. These elements are involved in control of gene expression which governs the phenotype and also plays role in disease biology. Therefore, genome-wide understanding of these elements holds great therapeutic promise. Several attempts have been made toward identification of S/MARs in genomes of various organisms including human. However, a comprehensive genome-wide map of human S/MARs is yet not available. Toward this objective, ChIP-Seq data of 14 S/MAR binding proteins were analyzed and the binding site coordinates of these proteins were used to prepare a non-redundant S/MAR dataset of human genome. Along with co-ordinate (location) details of S/MARs, the dataset also revealed details of S/MAR features, namely, length, inter-SMAR length (the chromatin loop size), nucleotide repeats, motif abundance, chromosomal distribution and genomic context. S/MARs identified in present study and their subsequent analysis also suggests that these elements act as hotspots for integration of retroviruses. Therefore, these data will help toward better understanding of genome functioning and designing effective anti-viral therapeutics. In order to facilitate user friendly browsing and retrieval of the data obtained in present study, a web interface, MARome (http://bioinfo.net.in/MARome), has been developed.


Assuntos
Cromatina/genética , DNA/genética , Genoma Humano/genética , Proteínas de Ligação à Região de Interação com a Matriz/genética , Regiões de Interação com a Matriz/genética , Sítios de Ligação/genética , Cromatina/metabolismo , Mapeamento Cromossômico/métodos , Biologia Computacional/métodos , DNA/metabolismo , Mineração de Dados/métodos , Genômica/métodos , Humanos , Internet , Proteínas de Ligação à Região de Interação com a Matriz/metabolismo , Ligação Proteica , Reprodutibilidade dos Testes
2.
Nat Commun ; 10(1): 2982, 2019 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-31278256

RESUMO

Hybrid rice breeding for exploiting hybrid vigor, heterosis, has greatly increased grain yield. However, the heterosis-related genes associated with rice grain production remain largely unknown, partly because comprehensive mapping of heterosis-related traits is still labor-intensive and time-consuming. Here, we present a quantitative trait locus (QTL) mapping method, GradedPool-Seq, for rapidly mapping QTLs by whole-genome sequencing of graded-pool samples from F2 progeny via bulked-segregant analysis. We implement this method and map-based cloning to dissect the heterotic QTL GW3p6 from the female line. We then generate the near isogenic line NIL-FH676::GW3p6 by introgressing the GW3p6 allele from the female line Guangzhan63-4S into the male inbred line Fuhui676. The NIL-FH676::GW3p6 exhibits grain yield highly increased compared to Fuhui676. This study demonstrates that it may be possible to achieve a high level of grain production in inbred rice lines without the need to construct hybrids.


Assuntos
Mapeamento Cromossômico/métodos , Grão Comestível/genética , Vigor Híbrido/genética , Oryza/genética , Melhoramento Vegetal/métodos , Cromossomos de Plantas/genética , Locos de Características Quantitativas/genética
3.
Mol Genet Genomics ; 294(5): 1327-1341, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31187273

RESUMO

A survey of genome-wide polymorphisms between closely related species is required to understand the molecular basis of the evolutionary differentiation of their genomes. Two wild diploid wheat species, namely Triticum monococcum ssp. aegilopoides and T. urartu, are closely related and harbour the Am and A genomes, respectively. The A-genome donor of tetraploid and common wheat is T. urartu, and T. monococcum ssp. monococcum is the cultivated form derived from the wild einkorn wheat subspecies aegilopoides. Although subspecies aegilopoides has been a useful genetic resource in wheat breeding, genome-wide molecular markers for this subspecies have not been sufficiently developed. Here, we describe the detection of genome-wide polymorphisms such as single-nucleotide polymorphisms (SNPs) and insertions/deletions (indels) from RNA sequencing (RNA-seq) data of leaf transcripts in 15 accessions of the two diploid wheat species. The SNPs and indels, detected using the A genome of common wheat as the reference genome, covered the entire chromosomes of these species. The polymorphism information facilitated a comparison of the genetic diversity of einkorn wheat with that of two related diploid Aegilops species, namely, Ae. tauschii and Ae. umbellulata. Cleaved amplified polymorphic sequence (CAPS) markers converted from the SNP data were efficiently developed to confirm the addition of aegilopoides subspecies chromosomes to tetraploid wheat in nascent allohexaploid lines with AABBAmAm genomes. In addition, the CAPS markers permitted linkage map construction in mapping populations of aegilopoides subspecies accessions. Therefore, these RNA-seq data provide information for further breeding of closely related species with no reference genome sequence data.


Assuntos
Marcadores Genéticos/genética , Folhas de Planta/genética , Polimorfismo de Nucleotídeo Único/genética , Triticum/genética , Mapeamento Cromossômico/métodos , Cromossomos de Plantas/genética , Diploide , Ligação Genética/genética , Genoma de Planta/genética , Estudo de Associação Genômica Ampla/métodos , Filogenia , Análise de Sequência de RNA/métodos , Tetraploidia
4.
Nat Commun ; 10(1): 2680, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31213597

RESUMO

Genetic studies of complex traits in animals have been hindered by the need to generate, maintain, and phenotype large panels of recombinant lines. We developed a new method, C. elegans eXtreme Quantitative Trait Locus (ceX-QTL) mapping, that overcomes this obstacle via bulk selection on millions of unique recombinant individuals. We use ceX-QTL to map a drug resistance locus with high resolution. We also map differences in gene expression in live worms and discovered that mutations in the co-chaperone sti-1 upregulate the transcription of HSP-90. Lastly, we use ceX-QTL to map loci that influence fitness genome-wide confirming previously reported causal variants and uncovering new fitness loci. ceX-QTL is fast, powerful and cost-effective, and will accelerate the study of complex traits in animals.


Assuntos
Caenorhabditis elegans/genética , Mapeamento Cromossômico/métodos , Aptidão Genética/genética , Locos de Características Quantitativas/genética , Característica Quantitativa Herdável , Animais , Mapeamento Cromossômico/economia , Resistência a Medicamentos/genética , Feminino , Regulação da Expressão Gênica/genética , Masculino , Fatores de Tempo
5.
Gene ; 710: 291-306, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31185283

RESUMO

The WD40 transcription factor family is a superfamily found in all eukaryotes that plays important roles in regulating growth and development. To our knowledge, to date, WD40 superfamily genes have been identified and characterized in several plant species, but little information is available on the WD40 superfamily genes in peach. In this study, we identified 220 members of the WD40 superfamily in the peach genome, and these members were further classified into five subfamilies based on phylogenetic comparison with those in Arabidopsis. The members within each subfamily had conserved motifs and gene structures. The WD40 genes were unevenly distributed on chromosomes 1 to 8 of the peach genome. Additionally, 58 pairs of paralog WD40 members were found on eight chromosomes in peach, and 242 pairs of orthologous WD40 genes in peach and Arabidopsis were matched. The 54 selected putative WD40 genes in peach had diverse expression patterns in red-fleshed and white-fleshed peach fruits at five developmental stages. Prupe.6G211800.1 was located only on the cytomembrane, while Prupe.1G428200.1 and Prupe.I003200.1 were located on both the cytomembrane and in the nucleus; Prupe.1G558700.1 was densely localized around the nuclear rim but relatively faintly localized in the nucleoplasm; Prupe.5G116300.1 was located in the nucleus and cytomembrane with strong signals but showed weak signals in the cytoplasm; and Prupe.8G212400.1 and Prupe.1G053600.1 were located mainly in the nuclear envelope and cytomembrane but relatively faintly in the nucleoplasm. This study provides a foundation for the further functional verification of WD40 genes in peach.


Assuntos
Mapeamento Cromossômico/métodos , Família Multigênica , Proteínas de Plantas/genética , Prunus persica/genética , Núcleo Celular/genética , Núcleo Celular/metabolismo , Cromossomos de Plantas/genética , Sequência Conservada , Citoplasma/genética , Citoplasma/metabolismo , Regulação da Expressão Gênica de Plantas , Filogenia , Proteínas de Plantas/metabolismo , Prunus persica/metabolismo , Repetições WD40
6.
PLoS Genet ; 15(5): e1008191, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31150378

RESUMO

Increasing agricultural productivity is one of the most important goals of plant science research and imperative to meet the needs of a rapidly growing population. Rice (Oryza sativa L.) is one of the most important staple crops worldwide. Grain size is both a major determinant of grain yield in rice and a target trait for domestication and artificial breeding. Here, a genome-wide association study of grain length and grain width was performed using 996,722 SNP markers in 270 rice accessions. Five and four quantitative trait loci were identified for grain length and grain width, respectively. In particular, the novel grain size gene OsSNB was identified from qGW7, and further results showed that OsSNB negatively regulated grain size. Most notably, knockout mutant plants by CRISPR/Cas9 technology showed increased grain length, width, and weight, while overexpression of OsSNB yielded the opposite. Sequencing of this gene from the promoter to the 3'-untranslated region in 168 rice accessions from a wide geographic range identified eight haplotypes. Furthermore, Hap 3 has the highest grain width discovered in japonica subspecies. Compared to other haplotypes, Hap 3 has a 225 bp insertion in the promoter. Based on the difference between Hap 3 and other haplotypes, OsSNB_Indel2 was designed as a functional marker for the improvement of rice grain width. This could be directly used to assist selection toward an improvement of grain width. These findings suggest OsSNB as useful for further improvements in yield characteristics in most cultivars.


Assuntos
Grão Comestível/genética , Oryza/genética , Alelos , Cruzamento , Mapeamento Cromossômico/métodos , Cromossomos de Plantas/genética , Genes de Plantas/genética , Variação Genética , Genética Populacional/métodos , Estudo de Associação Genômica Ampla/métodos , Haplótipos , Desequilíbrio de Ligação , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , Sementes/genética
7.
J Biosci ; 44(2)2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31180051

RESUMO

Restriction enzymes have been identified in the early 1950s of the past century and have quickly become key players in the molecular biology of DNA. Forty years ago, the scientists whose pioneering work had explored the activity and sequence specificity of these enzymes, contributing to the definition of their enormous potential as tools for DNA characterization, mapping and manipulation, were awarded the Nobel Prize. In this short review, we celebrate the history of these enzymes in the light of their many different uses, as these proteins have accompanied the history of DNA for over 50 years representing active witnesses of major steps in the field.


Assuntos
Mapeamento Cromossômico/história , Clonagem Molecular/métodos , Enzimas de Restrição do DNA/história , DNA/história , Biologia Molecular/história , Mapeamento de Nucleotídeos/história , Sistemas CRISPR-Cas , Cromatina/química , Cromatina/metabolismo , Mapeamento Cromossômico/métodos , DNA/química , DNA/genética , DNA/metabolismo , Metilação de DNA , Enzimas de Restrição do DNA/genética , Enzimas de Restrição do DNA/metabolismo , História do Século XX , História do Século XXI , Humanos , Biologia Molecular/métodos , Prêmio Nobel , Mapeamento de Nucleotídeos/métodos , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição/genética , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição/história , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição/metabolismo
8.
PLoS Comput Biol ; 15(6): e1006994, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31166948

RESUMO

The computational reconstruction of genome sequences from shotgun sequencing data has been greatly simplified by the advent of sequencing technologies that generate long reads. In the case of relatively small genomes (e.g., bacterial or viral), complete genome sequences can frequently be reconstructed computationally without the need for further experiments. However, large and complex genomes, such as those of most animals and plants, continue to pose significant challenges. In such genomes, assembly software produces incomplete and fragmented reconstructions that require additional experimentally derived information and manual intervention in order to reconstruct individual chromosome arms. Recent technologies originally designed to capture chromatin structure have been shown to effectively complement sequencing data, leading to much more contiguous reconstructions of genomes than previously possible. Here, we survey these technologies and the algorithms used to assemble and analyze large eukaryotic genomes, placed within the historical context of genome scaffolding technologies that have been in existence since the dawn of the genomic era.


Assuntos
Algoritmos , Mapeamento Cromossômico/métodos , Genoma Humano/genética , Genômica/métodos , Alinhamento de Sequência/métodos , Humanos , Análise de Sequência de DNA
9.
Nucleic Acids Res ; 47(15): e89, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31165870

RESUMO

Optical DNA mapping (ODM) allows visualization of long-range sequence information along single DNA molecules. The data can for example be used for detecting long range structural variations, for aiding DNA sequence assembly of complex genomes and for mapping epigenetic marks and DNA damage across the genome. ODM traditionally utilizes sequence specific marks based on nicking enzymes, combined with a DNA stain, YOYO-1, for detection of the DNA contour. Here we use a competitive binding approach, based on YOYO-1 and netropsin, which highlights the contour of the DNA molecules, while simultaneously creating a continuous sequence specific pattern, based on the AT/GC variation along the detected molecule. We demonstrate and validate competitive-binding-based ODM using bacterial artificial chromosomes (BACs) derived from the human genome and then turn to DNA extracted from white blood cells. We generalize our findings with in-silico simulations that show that we can map a vast majority of the human genome. Finally, we demonstrate the possibility of combining competitive binding with enzymatic labeling by mapping DNA damage sites induced by the cytotoxic drug etoposide to the human genome. Overall, we demonstrate that competitive-binding-based ODM has the potential to be used both as a standalone assay for studies of the human genome, as well as in combination with enzymatic approaches, some of which are already commercialized.


Assuntos
Benzoxazóis/química , Mapeamento Cromossômico/métodos , DNA/química , Genoma Humano , Netropsina/química , Compostos de Quinolínio/química , Análise de Sequência de DNA/métodos , Antineoplásicos Fitogênicos/farmacologia , Sítios de Ligação , Ligação Competitiva , Cromossomos Artificiais Bacterianos/química , DNA/genética , Etoposídeo/farmacologia , Corantes Fluorescentes/química , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Imagem Individual de Molécula/métodos
10.
BMC Plant Biol ; 19(1): 201, 2019 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-31096901

RESUMO

BACKGROUND: The kernel row number (KRN) of an ear is an important trait related to yield and domestication in maize. Exploring the underlying genetic mechanisms of KRN has great research significance and application potential. RESULTS: In the present study, N531 with a KRN of 18-22 and SLN with a KRN of 4-6 were used as the recurrent parent and the donor parent, respectively, to develop two introgression lines (ILs), IL_A and IL_B, both of which have common negative-effect alleles from SLN on chromosomes 1, 5 and 10 and significantly reduced inflorescence meristem (IM) diameter and KRN compared with those of N531. We used RNA-Seq to investigate the transcriptome profiles of 5-mm immature ears of N531, IL_A and IL_B. We identified a total of 2872 differentially expressed genes (DEGs) between N531 and IL_A, 2428 DEGs between N531 and IL_B and 1811 DEGs between IL_A and IL_B. A total of 1252 DEGs were detected as overlapping DEGs, and 89 DEGs were located on the common introgression fragments. Furthermore, three DEGs (Zm00001d013277, Zm00001d015310 and Zm00001d015377) containing three SNPs associated with KRN were identified using regional association mapping. CONCLUSIONS: These results will facilitate our understanding of ear development and provide important candidate genes for further study on KRN.


Assuntos
Grão Comestível/crescimento & desenvolvimento , Zea mays/genética , Mapeamento Cromossômico/métodos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/genética , Genes de Plantas/genética , Genes de Plantas/fisiologia , Estudos de Associação Genética , Locos de Características Quantitativas/genética , Característica Quantitativa Herdável , Reação em Cadeia da Polimerase em Tempo Real
11.
BMC Genomics ; 20(1): 355, 2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31072298

RESUMO

BACKGROUND: Anaerobic germination tolerance is an important trait for direct-seeded rice varieties. Understanding the genetic basis of anaerobic germination is a key for breeding direct-seeded rice varieties. RESULTS: In this study, a recombinant inbred line (RIL) population derived from a cross between YZX and 02428 exhibited obvious coleoptile phenotypic differences. Mapping analysis using a high-density bin map indicated that a total of 25 loci were detected across two cropping seasons, including 10 previously detected loci and a total of 13 stable loci. Analysis of the 13 stable loci demonstrated that the more elite alleles that were pyramided in an individual, the higher the values of these traits were in the two cropping seasons. Furthermore, some anaerobic germination-tolerant recombinant inbred lines, namely G9, G10, G16, and G151, were identified. A total of 84 differentially expressed genes were obtained from the 13 stable loci via genome-wide expression analysis of the two parents at three key periods. Among them, Os06g0110200, Os07g0638300, Os07g0638400, Os09g0532900, Os09g0531701 and Os12g0539751 constitute the best candidates associated with anaerobic germination. CONCLUSIONS: Both the anaerobic germination-tolerant recombinant inbred lines and the loci identified in this study will provide new genetic resources for improving the anaerobic germination tolerance of rice using molecular breeding strategies, as well as will broaden our understanding of the genetic control of germination tolerance under anaerobic conditions.


Assuntos
Mapeamento Cromossômico/métodos , Cromossomos de Plantas , Germinação , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Oryza/genética , Proteínas de Plantas/genética , Locos de Características Quantitativas , Anaerobiose , Oryza/crescimento & desenvolvimento , Oryza/fisiologia
12.
Mol Genet Genomics ; 294(5): 1231-1239, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31098741

RESUMO

The great majority of terrestrial plants produce epicuticular wax that is used to protect plants from a variety of biotic and abiotic stresses. Cabbage epicuticular wax is a white crystalline compound of various lipids. Wax-less cabbage has the characteristics of lustrous green leaves and beautiful exterior, which facilitates the brilliant green cabbage breeding. CGL-3 is a spontaneous wax-less mutant identified from cabbage. Genetic analysis indicated that the waxy deficiency of the mutant was controlled by a single dominant gene. To clarify the mechanism of the waxy deficiency, fine-mapping and transcriptome analysis of the wax-less gene, BoGL-3, were carried out in this study. The result of fine mapping showed that the wax-less gene, BoGL-3, was delimited in a 33.5-kb interval which is between the flanking marker C08-98 and the end of chromosome 8. Two cDNA libraries, constructed with wax-less cabbage CGL-3 and the wild-type cabbage WT, were sequenced for screening of the target gene BoGL-3. A total of 8340 genes were identified with significant differential expression between CGL-3 and WT. Among these genes, 3187 were up-regulated and 5153 were down-regulated in CGL-3. With homologous analysis, four differential expressed genes related to wax metabolism were obtained. Among these four genes, only Bol018504 is located within the region of fine-mapping. Bol08504 is homologous to CER1, which encodes fatty acid hydroxylase and plays an important role in wax synthesis in Arabidopsis. However, there was no difference of Bol08504 sequence between CGL-3 and WT. We suggested that Bol018504 was regulated by BoGL-3. The suppression of Bol018504 leads to the reduction of wax. These findings will be helpful to reveal the mechanism of the wax metabolism in cabbage and develop lustrous green cabbage germplasm material.


Assuntos
Brassica/genética , Genes de Plantas/genética , Transcriptoma/genética , Arabidopsis/genética , Mapeamento Cromossômico/métodos , Regulação para Baixo/genética , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas/genética , Genes Dominantes/genética , Marcadores Genéticos/genética , Genoma de Planta/genética , Folhas de Planta/genética , Estresse Fisiológico/genética , Regulação para Cima/genética
13.
Int J Mol Sci ; 20(10)2019 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-31108903

RESUMO

Leaf rust caused by Puccinia triticina Eriks is one of the most problematic diseases of wheat throughout the world. The gene Lr42 confers effective resistance against leaf rust at both seedling and adult plant stages. Previous studies had reported Lr42 to be both recessive and dominant in hexaploid wheat; however, in diploid Aegilops tauschii (TA2450), we found Lr42 to be dominant by studying segregation in two independent F2 and their F2:3 populations. We further fine-mapped Lr42 in hexaploid wheat using a KS93U50/Morocco F5 recombinant inbred line (RIL) population to a 3.7 cM genetic interval flanked by markers TC387992 and WMC432. The 3.7 cM Lr42 region physically corresponds to a 3.16 Mb genomic region on chromosome 1DS based on the Chinese Spring reference genome (RefSeq v.1.1) and a 3.5 Mb genomic interval on chromosome 1 in the Ae. tauschii reference genome. This region includes nine nucleotide-binding domain leucine-rich repeat (NLR) genes in wheat and seven in Ae. tauschii, respectively, and these are the likely candidates for Lr42. Furthermore, we developed two kompetitive allele-specific polymorphism (KASP) markers (SNP113325 and TC387992) flanking Lr42 to facilitate marker-assisted selection for rust resistance in wheat breeding programs.


Assuntos
Mapeamento Cromossômico/métodos , Resistência à Doença , Proteínas de Plantas/genética , Triticum/microbiologia , Basidiomycota/patogenicidade , Sítios de Ligação , Doenças das Plantas/microbiologia , Folhas de Planta/genética , Folhas de Planta/microbiologia , Proteínas de Plantas/química , Poliploidia , Triticum/genética
14.
Mol Genet Genomics ; 294(5): 1277-1288, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31139941

RESUMO

Stalk lodging severely limits the grain yield of maize (Zea mays L.). Mechanical stalk strength can be reflected by the traits of stalk diameter (SD), stalk bending strength (SBS), and lodging rind penetrometer resistance (RPR). To determine the genetic basis of maize stalk lodging, quantitative trait loci (QTLs) were mapped for these three traits using the IBM Syn10 DH population in three environments. The results indicated that there were strong genetic correlations among the three traits, and the analyses of phenotypic variations for SD, SBS, and RPR across the three environments showed high broad-sense heritability (0.6843, 0.5175, and 0.7379, respectively). In total, 44 significant QTLs were identified control the above traits across the 3 environments. A total of 14, 14, and 16 QTLs were identified for SD, SBS, and RPR across single-environment mapping, respectively. Notably, ten QTLs were stably expressed across multiple-environments, including two QTLs for SD, three for SBS, and five for RPR. Three major QTLs each accounting for over 10% of the phenotypic variation were qSD6-2 (10.03%), qSD8-2 (13.73%), and qSBS1-2 (11.89%). Comprehensive analysis of all QTLs in this study revealed that 5 QTL clusters including 12 QTLs were located on chromosomes 1, 3, 7, and 8, respectively. Among these 44 QTLs, 9 harbored 13 stalk lodging-associated SNPs that were detected by our recently published work, with 1 SNP successfully validated in the IBM Syn10 DH population. These chromosomal regions will be useful for marker-assisted selection and fine mapping of stalk lodging-related traits in maize.


Assuntos
Genes de Plantas/genética , Zea mays/genética , Mapeamento Cromossômico/métodos , Cruzamentos Genéticos , Grão Comestível/genética , Ligação Genética/genética , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética
15.
BMC Genomics ; 20(1): 371, 2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31088355

RESUMO

BACKGROUND: Barbarea vulgaris is a wild cruciferous plant and include two distinct types: the G- and P-types named after their glabrous and pubescent leaves, respectively. The types differ significantly in resistance to a range of insects and diseases as well as glucosinolates and other chemical defenses. A high-density linkage map was needed for further progress to be made in the molecular research of this plant. RESULTS: We performed restriction site-associated DNA sequencing (RAD-Seq) on an F2 population generated from G- and P-type B. vulgaris. A total of 1545 SNP markers were mapped and ordered in eight linkage groups, which represents the highest density linkage map to date for the crucifer tribe Cardamineae. A total of 722 previously published genome contigs (50.2 Mb, 30% of the total length) can be anchored to this high density genetic map, an improvement compared to a previously published map (431 anchored contigs, 38.7 Mb, 23% of the assembly genome). Most of these (572 contigs, 31.2 Mb) were newly anchored to the map, representing a significant improvement. On the basis of the present high-density genetic map, 37 QTL were detected for eleven traits, each QTL explaining 2.9-71.3% of the phenotype variation. QTL of glucosinolates, leaf size and color traits were in most cases overlapping, possibly implying a functional connection. CONCLUSIONS: This high-density linkage map and the QTL obtained in this study will be useful for further understanding of the genetic of the B. vulgaris and molecular basis of these traits, many of which are shared in the related crop watercress.


Assuntos
Barbarea/genética , Mapeamento Cromossômico/métodos , Locos de Características Quantitativas , Análise de Sequência de DNA/métodos , Barbarea/fisiologia , DNA de Plantas/genética , Ligação Genética , Fenótipo , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Polimorfismo de Nucleotídeo Único
16.
BMC Plant Biol ; 19(1): 167, 2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31035914

RESUMO

BACKGROUND: First flower node (FFN) is an important trait for evaluating fruit earliness in pepper (Capsicum annuum L.). The trait is controlled by quantitative trait loci (QTL); however, studies have been limited on QTL mapping and genes contributing to the trait. RESULTS: In this study, we developed a high density genetic map using specific-locus amplified fragment sequencing (SLAF-seq), a high-throughput strategy for de novo single nucleotide polymorphism discovery, based on 146 recombinant inbred lines (RILs) derived from an intraspecific cross between PM702 and FS871. The map contained 9328 SLAF markers on 12 linkage groups (LGs), and spanned a total genetic distance of 2009.69 centimorgan (cM) with an average distance of 0.22 cM. The sequencing depth for the map was 72.39-fold in the male parent, 57.04-fold in the female parent, and 15.65-fold in offspring. Using the genetic map, two major QTLs, named Ffn2.1 and Ffn2.2, identified on LG02 were strongly associated with FFN, with a phenotypic variance explanation of 28.62 and 19.56%, respectively. On the basis of the current annotation of C. annuum cv. Criollo de Morelos (CM334), 59 candidate genes were found within the Ffn2.1 and Ffn2.2 region, but only 3 of 59 genes were differentially expressed according to the RNA-seq results. Eventually we identified one gene associated with the FFN based on the function through GO, KEGG, and Swiss-prot analysis. CONCLUSIONS: Our research showed that the construction of high-density genetic map using SLAF-seq is a valuable tool for fine QTL mapping. The map we constructed is by far the most saturated complete genetic map of pepper, and using it we conducted fine QTL mapping for the important trait, FFN. QTLs and candidate genes obtained in this study lay a good foundation for the further research on FFN-related genes and other genetic applications in pepper.


Assuntos
Capsicum/genética , Mapeamento Cromossômico/métodos , Cromossomos de Plantas , Flores/genética , Locos de Características Quantitativas , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Ligação Genética , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Polimorfismo de Nucleotídeo Único
17.
Nucleic Acids Res ; 47(15): e85, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31114914

RESUMO

Whole-genome bisulfite sequencing (WGBS) is the current gold standard of methylome analysis. Post-bisulfite adaptor tagging (PBAT) is an increasingly popular WGBS protocol because of high sensitivity and low bias. PBAT originally relied on two rounds of random priming for adaptor-tagging of single-stranded DNA (ssDNA) to attain high efficiency but at a cost of library insert length. To overcome this limitation, we developed terminal deoxyribonucleotidyl transferase (TdT)-assisted adenylate connector-mediated ssDNA (TACS) ligation as an alternative to random priming. In this method, TdT attaches adenylates to the 3'-end of input ssDNA, which are then utilized by RNA ligase as an efficient connector to the ssDNA adaptor. A protocol that uses TACS ligation instead of the second random priming step substantially increased the lengths of PBAT library fragments. Moreover, we devised a dual-library strategy that splits the input DNA to prepare two libraries with reciprocal adaptor polarity, combining them prior to sequencing. This strategy ensured an ideal base-color balance to eliminate the need for DNA spike-in for color compensation, further improving the throughput and quality of WGBS. Adopting the above strategies to the HiSeq X Ten and NovaSeq 6000 platforms, we established a cost-effective, high-quality WGBS, which should accelerate various methylome analyses.


Assuntos
Mapeamento Cromossômico/métodos , DNA de Cadeia Simples/genética , Genoma Humano , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sulfitos/química , Monofosfato de Adenosina/genética , Monofosfato de Adenosina/metabolismo , Bacteriófago lambda/genética , Bacteriófago lambda/metabolismo , Metilação de DNA , DNA Nucleotidilexotransferase/genética , DNA Nucleotidilexotransferase/metabolismo , DNA de Cadeia Simples/metabolismo , Biblioteca Genômica , Humanos , RNA Ligase (ATP)/genética , RNA Ligase (ATP)/metabolismo
18.
PLoS Genet ; 15(5): e1008149, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31067226

RESUMO

Tomato (Solanum lycopersicum) is one of the highest-value vegetable crops worldwide. Understanding the genetic regulation of primary metabolite levels can inform efforts aimed toward improving the nutrition of commercial tomato cultivars, while maintaining key traits such as yield and stress tolerance. We identified 388 suggestive association loci (including 126 significant loci) for 92 metabolic traits including nutrition and flavor-related loci by genome-wide association study from 302 accessions in two different environments. Among them, an ascorbate quantitative trait locus TFA9 (TOMATO FRUIT ASCORBATEON CHROMOSOME 9) co-localized with SlbHLH59, which promotes high ascorbate accumulation by directly binding to the promoter of structural genes involved in the D-mannose/L-galactose pathway. The causal mutation of TFA9 is an 8-bp InDel, named InDel_8, located in the promoter region of SlbHLH59 and spanned a 5'UTR Py-rich stretch motif affecting its expression. Phylogenetic analysis revealed that differentially expressed SlbHLH59 alleles were selected during tomato domestication. Our results provide a dramatic illustration of how ascorbate biosynthesis can be regulated and was selected during the domestication of tomato. Furthermore, the findings provide novel genetic insights into natural variation of metabolites in tomato fruit, and will promote efficient utilization of metabolite traits in tomato improvement.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Lycopersicon esculentum/genética , Lycopersicon esculentum/metabolismo , Alelos , Ácido Ascórbico/genética , Ácido Ascórbico/metabolismo , Mapeamento Cromossômico/métodos , Frutas/genética , Galactose/biossíntese , Galactose/metabolismo , Regulação da Expressão Gênica de Plantas/genética , Variação Genética/genética , Genoma de Planta/genética , Estudo de Associação Genômica Ampla , Manose/biossíntese , Manose/metabolismo , Filogenia , Regiões Promotoras Genéticas/genética , Locos de Características Quantitativas/genética
19.
BMC Genomics ; 20(1): 331, 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31046664

RESUMO

BACKGROUND: Both a source of diversity and the development of genomic tools, such as reference genomes and molecular markers, are equally important to enable faster progress in plant breeding. Pear (Pyrus spp.) lags far behind other fruit and nut crops in terms of employment of available genetic resources for new cultivar development. To address this gap, we designed a high-density, high-efficiency and robust single nucleotide polymorphism (SNP) array for pear, with the main objectives of conducting genetic diversity and genome-wide association studies. RESULTS: By applying a two-step design process, which consisted of the construction of a first 'draft' array for the screening of a small subset of samples, we were able to identify the most robust and informative SNPs to include in the Applied Biosystems™ Axiom™ Pear 70 K Genotyping Array, currently the densest SNP array for pear. Preliminary evaluation of this 70 K array in 1416 diverse pear accessions from the USDA National Clonal Germplasm Repository (NCGR) in Corvallis, OR identified 66,616 SNPs (93% of all the tiled SNPs) as high quality and polymorphic (PolyHighResolution). We further used the Axiom Pear 70 K Genotyping Array to construct high-density linkage maps in a bi-parental population, and to make a direct comparison with available genotyping-by-sequencing (GBS) data, which suggested that the SNP array is a more robust method of screening for SNPs than restriction enzyme reduced representation sequence-based genotyping. CONCLUSIONS: The Axiom Pear 70 K Genotyping Array, with its high efficiency in a widely diverse panel of Pyrus species and cultivars, represents a valuable resource for a multitude of molecular studies in pear. The characterization of the USDA-NCGR collection with this array will provide important information for pear geneticists and breeders, as well as for the optimization of conservation strategies for Pyrus.


Assuntos
Mapeamento Cromossômico/métodos , Ligação Genética , Marcadores Genéticos , Genoma de Planta , Polimorfismo de Nucleotídeo Único , Pyrus/genética , Sementes/genética , Cromossomos de Plantas , Estudo de Associação Genômica Ampla , Técnicas de Genotipagem
20.
Nat Protoc ; 14(5): 1661-1685, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30996261

RESUMO

Nascent RNA may form a three-stranded structure with DNA, called an R-loop, which has been linked to fundamental biological processes such as transcription, replication and genome instability. Here, we provide a detailed protocol for a newly developed strategy, named R-ChIP, for robust capture of R-loops genome-wide. Distinct from R-loop-mapping methods based on the monoclonal antibody S9.6, which recognizes RNA-DNA hybrid structures, R-ChIP involves expression of an exogenous catalytically inactive RNASEH1 in cells to bind RNA-DNA hybrids but not resolve them. This is followed by chromatin immunoprecipitation (ChIP) of the tagged RNASEH1 and construction of a strand-specific library for deep sequencing. It takes ~3 weeks to establish a stable cell line expressing the mutant enzyme and 5 more days to proceed with the R-ChIP protocol. In principle, R-ChIP is applicable to both cell lines and animals, as long as the catalytically inactive RNASEH1 can be expressed to study the dynamics of R-loop formation and resolution, as well as its impact on the functionality of the genome. In our recent studies with R-ChIP, we showed an intimate spatiotemporal relationship between R-loops and RNA polymerase II pausing/pause release, as well as linking augmented R-loop formation to DNA damage response induced by driver mutations of key splicing factors associated with myelodysplastic syndrome (MDS).


Assuntos
Imunoprecipitação da Cromatina/métodos , Mapeamento Cromossômico/métodos , RNA/genética , Ribonuclease H/genética , Animais , Linhagem Celular , DNA/química , DNA/genética , RNA/química , Ribonuclease H/química , Ribonuclease H/metabolismo
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