fine mapping
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KEY MESSAGE: Making use of wheat chromosomal resources, we developed 11 gene-associated markers for the region of interest, which allowed reducing gene interval and spanning it by four BAC clones. Positional gene cloning and targeted marker development in bread wheat are hampered by high complexity and polyploidy of its nuclear genome. Aiming to clone a Russian wheat aphid resistance gene Dn2401 located on wheat chromosome arm 7DS, we have developed a strategy overcoming problems due to polyploidy and enabling efficient development of gene-associated markers from the region of interest. We employed information gathered by GenomeZipper, a synteny-based tool combining sequence data of rice, Brachypodium, sorghum and barley, and took advantage of a high-density linkage map of Aegilops tauschii. To ensure genome- and locus-specificity of markers, we made use of survey sequence assemblies of isolated wheat chromosomes 7A, 7B and 7D. Despite the low level of polymorphism of the wheat D subgenome, our approach allowed us to add in an efficient and cost-effective manner 11 new gene-associated markers in the Dn2401 region and narrow down the target interval to 0.83 cM. Screening 7DS-specific BAC library with the flanking markers revealed a contig of four BAC clones that span the Dn2401 region in wheat cultivar 'Chinese Spring'. With the availability of sequence assemblies and GenomeZippers for each of the wheat chromosome arms, the proposed strategy can be applied for focused marker development in any region of the wheat genome.
- MeSH
- býložravci MeSH
- chromozomy rostlin MeSH
- DNA primery MeSH
- DNA rostlinná genetika MeSH
- genetická vazba MeSH
- genetické markery MeSH
- genomika MeSH
- jednonukleotidový polymorfismus MeSH
- mapování chromozomů * MeSH
- mikrosatelitní repetice MeSH
- mšice * MeSH
- pšenice genetika MeSH
- rostlinné geny * MeSH
- syntenie MeSH
- umělé bakteriální chromozomy MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Rusko MeSH
Powdery mildew, caused by Blumeria graminis f. sp. tritici, is one of the most important wheat diseases in the world. In this study, a single dominant powdery mildew resistance gene MlIW172 was identified in the IW172 wild emmer accession and mapped to the distal region of chromosome arm 7AL (bin7AL-16-0.86-0.90) via molecular marker analysis. MlIW172 was closely linked with the RFLP probe Xpsr680-derived STS marker Xmag2185 and the EST markers BE405531 and BE637476. This suggested that MlIW172 might be allelic to the Pm1 locus or a new locus closely linked to Pm1. By screening genomic BAC library of durum wheat cv. Langdon and 7AL-specific BAC library of hexaploid wheat cv. Chinese Spring, and after analyzing genome scaffolds of Triticum urartu containing the marker sequences, additional markers were developed to construct a fine genetic linkage map on the MlIW172 locus region and to delineate the resistance gene within a 0.48 cM interval. Comparative genetics analyses using ESTs and RFLP probe sequences flanking the MlIW172 region against other grass species revealed a general co-linearity in this region with the orthologous genomic regions of rice chromosome 6, Brachypodium chromosome 1, and sorghum chromosome 10. However, orthologous resistance gene-like RGA sequences were only present in wheat and Brachypodium. The BAC contigs and sequence scaffolds that we have developed provide a framework for the physical mapping and map-based cloning of MlIW172.
Lung cancer has several genetic associations identified within the major histocompatibility complex (MHC); although the basis for these associations remains elusive. Here, we analyze MHC genetic variation among 26,044 lung cancer patients and 20,836 controls densely genotyped across the MHC, using the Illumina Illumina OncoArray or Illumina 660W SNP microarray. We impute sequence variation in classical HLA genes, fine-map MHC associations for lung cancer risk with major histologies and compare results between ethnicities. Independent and novel associations within HLA genes are identified in Europeans including amino acids in the HLA-B*0801 peptide binding groove and an independent HLA-DQB1*06 loci group. In Asians, associations are driven by two independent HLA allele sets that both increase risk in HLA-DQB1*0401 and HLA-DRB1*0701; the latter better represented by the amino acid Ala-104. These results implicate several HLA-tumor peptide interactions as the major MHC factor modulating lung cancer susceptibility.
- MeSH
- Asijci genetika MeSH
- běloši genetika MeSH
- frekvence genu MeSH
- genetická predispozice k nemoci etnologie genetika MeSH
- genotyp MeSH
- HLA antigeny genetika MeSH
- hlavní histokompatibilní komplex genetika MeSH
- jednonukleotidový polymorfismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- mapování chromozomů * MeSH
- nádory plic etnologie genetika MeSH
- peptidy genetika MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Genome-wide association studies (GWAS) have identified more than 200 common genetic variants independently associated with colorectal cancer (CRC) risk, but the causal variants and target genes are mostly unknown. We sought to fine-map all known CRC risk loci using GWAS data from 100,204 cases and 154,587 controls of East Asian and European ancestry. Our stepwise conditional analyses revealed 238 independent association signals of CRC risk, each with a set of credible causal variants (CCVs), of which 28 signals had a single CCV. Our cis-eQTL/mQTL and colocalization analyses using colorectal tissue-specific transcriptome and methylome data separately from 1299 and 321 individuals, along with functional genomic investigation, uncovered 136 putative CRC susceptibility genes, including 56 genes not previously reported. Analyses of single-cell RNA-seq data from colorectal tissues revealed 17 putative CRC susceptibility genes with distinct expression patterns in specific cell types. Analyses of whole exome sequencing data provided additional support for several target genes identified in this study as CRC susceptibility genes. Enrichment analyses of the 136 genes uncover pathways not previously linked to CRC risk. Our study substantially expanded association signals for CRC and provided additional insight into the biological mechanisms underlying CRC development.
- MeSH
- Asijci * genetika MeSH
- běloši * genetika MeSH
- celogenomová asociační studie * MeSH
- genetická predispozice k nemoci * MeSH
- jednonukleotidový polymorfismus * MeSH
- kolorektální nádory * genetika MeSH
- lidé MeSH
- lokus kvantitativního znaku * MeSH
- mapování chromozomů MeSH
- sekvenování exomu MeSH
- studie případů a kontrol MeSH
- transkriptom MeSH
- východní Asiaté MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Genome-wide association studies have identified breast cancer risk variants in over 150 genomic regions, but the mechanisms underlying risk remain largely unknown. These regions were explored by combining association analysis with in silico genomic feature annotations. We defined 205 independent risk-associated signals with the set of credible causal variants in each one. In parallel, we used a Bayesian approach (PAINTOR) that combines genetic association, linkage disequilibrium and enriched genomic features to determine variants with high posterior probabilities of being causal. Potentially causal variants were significantly over-represented in active gene regulatory regions and transcription factor binding sites. We applied our INQUSIT pipeline for prioritizing genes as targets of those potentially causal variants, using gene expression (expression quantitative trait loci), chromatin interaction and functional annotations. Known cancer drivers, transcription factors and genes in the developmental, apoptosis, immune system and DNA integrity checkpoint gene ontology pathways were over-represented among the highest-confidence target genes.
- MeSH
- Bayesova věta MeSH
- celogenomová asociační studie * MeSH
- genetická predispozice k nemoci * MeSH
- jednonukleotidový polymorfismus * MeSH
- lidé MeSH
- lokus kvantitativního znaku * MeSH
- mapování chromozomů metody MeSH
- nádorové biomarkery genetika MeSH
- nádory prsu genetika MeSH
- regulační oblasti nukleových kyselin MeSH
- rizikové faktory MeSH
- vazebná nerovnováha MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
BACKGROUND: QTL cloning for the discovery of genes underlying polygenic traits has historically been cumbersome in long-lived perennial plants like Populus. Linkage disequilibrium-based association mapping has been proposed as a cloning tool, and recent advances in high-throughput genotyping and whole-genome resequencing enable marker saturation to levels sufficient for association mapping with no a priori candidate gene selection. Here, multiyear and multienvironment evaluation of cell wall phenotypes was conducted in an interspecific P. trichocarpa x P. deltoides pseudo-backcross mapping pedigree and two partially overlapping populations of unrelated P. trichocarpa genotypes using pyrolysis molecular beam mass spectrometry, saccharification, and/ or traditional wet chemistry. QTL mapping was conducted using a high-density genetic map with 3,568 SNP markers. As a fine-mapping approach, chromosome-wide association mapping targeting a QTL hot-spot on linkage group XIV was performed in the two P. trichocarpa populations. Both populations were genotyped using the 34 K Populus Infinium SNP array and whole-genome resequencing of one of the populations facilitated marker-saturation of candidate intervals for gene identification. RESULTS: Five QTLs ranging in size from 0.6 to 1.8 Mb were mapped on linkage group XIV for lignin content, syringyl to guaiacyl (S/G) ratio, 5- and 6-carbon sugars using the mapping pedigree. Six candidate loci exhibiting significant associations with phenotypes were identified within QTL intervals. These associations were reproducible across multiple environments, two independent genotyping platforms, and different plant growth stages. cDNA sequencing for allelic variants of three of the six loci identified polymorphisms leading to variable length poly glutamine (PolyQ) stretch in a transcription factor annotated as an ANGUSTIFOLIA C-terminus Binding Protein (CtBP) and premature stop codons in a KANADI transcription factor as well as a protein kinase. Results from protoplast transient expression assays suggested that each of the polymorphisms conferred allelic differences in the activation of cellulose, hemicelluloses, and lignin pathway marker genes. CONCLUSION: This study illustrates the utility of complementary QTL and association mapping as tools for gene discovery with no a priori candidate gene selection. This proof of concept in a perennial organism opens up opportunities for discovery of novel genetic determinants of economically important but complex traits in plants.
- MeSH
- alely MeSH
- buněčná stěna genetika MeSH
- celulosa metabolismus MeSH
- fenotyp MeSH
- genetická vazba MeSH
- genotyp MeSH
- jednonukleotidový polymorfismus MeSH
- lignin biosyntéza MeSH
- lod skóre MeSH
- lokus kvantitativního znaku MeSH
- mapování chromozomů MeSH
- Populus genetika MeSH
- rostlinné geny * MeSH
- rostlinné proteiny chemie genetika MeSH
- sekvence nukleotidů MeSH
- sekvenční seřazení MeSH
- transkripční faktory chemie genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
Repetitive DNAs comprise large portion of eukaryote genomes. In genome projects, the assembly of repetitive DNAs is challenging due to the similarity between repeats, which generate ambiguities for alignment. Fluorescence in situ hybridization (FISH) is a powerful technique for the physical mapping of various sequences on chromosomes. This technique is thus very helpful in chromosome-based genome assemblies, providing information on the fine architecture of genomes and their evolution. However, various protocols are currently used for FISH mapping, most of which are relatively laborious and expensive, or work properly only with a specific type of probes or sequences, and there is a need for a universal and affordable FISH protocol. Here we tested a FISH protocol for mapping of different DNA repeats, such as multigene families (rDNAs, U snDNAs, histone genes), satellite DNAs, microsatellites, transposable elements, DOP-PCR products, and telomeric motif (TTAGG)n, on the chromosomes of various insects and other arthropods. Different cell types and stages obtained from diverse tissues were used. The FISH procedure proved high quality and reliable results in all experiments performed. We obtained data on the chromosomal distribution of DNA repeats in representatives of insects and other arthropods. Thus, our results allow us to conclude that the protocol is universal and requires only time adjustment for chromosome/DNA denaturation. The use of this FISH protocol will facilitate studies focused on understanding the evolution and role of repetitive DNA in arthropod genomes.
- MeSH
- členovci genetika MeSH
- DNA genetika MeSH
- fluorescence MeSH
- hmyz genetika MeSH
- hybridizace in situ fluorescenční metody MeSH
- mapování chromozomů metody MeSH
- molekulární evoluce MeSH
- multigenová rodina genetika MeSH
- repetitivní sekvence nukleových kyselin genetika MeSH
- telomery genetika MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The function of p53 is modulated by binding to a number of cellular and viral proteins, such as MDM2 and SV40 large T antigen. An initial immunochemical characterization of the p53-MDM2 complex in a rat fibroblast cell line (Clone 6) suggested that the anti-p53 monoclonal antibody Bp53-19 failed to immunoprecipitate the complex, and only recognized a fraction of the available p53 protein. Following the recent identification of the Bp53-19 epitope at the N-terminal end of p53, in the vicinity of where MDM2 protein was known to bind, we investigated the possibility that Bp53-19 might identify a region of p53 that interacts with MDM2 protein. MDM2 was found to bind with great specificity to short synthetic peptides derived from the N-terminus of p53. Several p53 synthetic peptides libraries, and an alanine substitution series at the optimal binding site, were used to establish the MDM2 binding site, in fine detail, to the sequence TFSGLW (aa 18-23) in mouse and TFSDLW in man (aa 18-23). The key residues required for MDM2 binding are almost identical to those required for the monoclonal antibody Bp53-19 to bind and this region of p53 is recognised by many other anti-p53 antibodies.
- MeSH
- imunochemie MeSH
- jaderné proteiny MeSH
- kultivované buňky MeSH
- molekulární sekvence - údaje MeSH
- myši MeSH
- nádorové proteiny metabolismus MeSH
- nádorový supresorový protein p53 metabolismus MeSH
- peptidové fragmenty metabolismus MeSH
- protoonkogenní proteiny c-mdm2 MeSH
- protoonkogenní proteiny MeSH
- sekvence aminokyselin MeSH
- vazebná místa MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
Adaxial, abaxial phylloplane (leaf), and spermoplane (seed) are proximal yet contrasting habitats for a microbiota that needs to be adequately explored. Here, we proposed novel methods to decipher the adaxial/abaxial-phylloplane and spermoplane-microbiomes. Comparison of 22 meta barcoded-NGS datasets (size of total data set-1980.48 Mb) enabled us to fine-map the microbiome of the rice foliar niche, which encompasses the lower, middle, top leaf as well panicle. Here, the total- and the cultivable-microbiome profiling revealed 157 genera representing ten phyla and 87 genera from 4 bacterial phyla, respectively, with a predominance of Proteobacteria and Actinobacteria. Interestingly, more bacterial communities (124-genera) preferred the abaxial than the adaxial phylloplane (104-genera) and spermoplane (67-genera) for colonization. The microbiome profiles were nearly identical on the aromatic (125-genera) and non-aromatic rice (116-genera) with high representation of Pantoea, Methylobacterium, Curtobacterium, Sphingopyxis, and Microbacterium. The culturomics investigation confirmed the abundance of Pantoea, Chryseobacterium, Pseudomonas, Acinetobacter, Sphingobacterium, and Exiguobacterium. One hundred bacterial isolates characterized and identified by polyphasic-taxonomic tools revealed the dominance of Acinetobacter, Chryseobacterium, Enterobacter, Massilia, Pantoea, Pseudomonas, and Stenotrophomonas on adaxial/abaxial-phylloplane and spermoplane. The study culminated in identifying hitherto unexplored bacterial communities on the adaxial/abaxial phylloplane and spermoplane of rice that can be harnessed for microbiome-assisted rice cultivation in the future.
- MeSH
- genotyp MeSH
- listy rostlin mikrobiologie MeSH
- mikrobiota * MeSH
- rýže (rod) * MeSH
- Sphingomonadaceae * MeSH
- Publikační typ
- časopisecké články MeSH
The novel wheat powdery mildew and stripe rust resistance genes Pm5V/Yr5V are introgressed from Dasypyrum villosum and fine mapped to a narrowed region in 5VS, and their effects on yield-related traits were characterized. The powdery mildew and stripe rust seriously threaten wheat production worldwide. Dasypyrum villosum (2n = 2x = 14, VV), a relative of wheat, is a valuable resource of resistance genes for wheat improvement. Here, we describe a platform for rapid introgression of the resistance genes from D. villosum into the wheat D genome. A complete set of new wheat-D. villosum V (D) disomic substitution lines and 11 D/V Robertsonian translocation lines are developed and characterized by molecular cytogenetic method. A new T5DL·5V#5S line NAU1908 shows resistance to both powdery mildew and stripe rust, and the resistances associated with 5VS are confirmed to be conferred by seedling resistance gene Pm5V and adult-plant resistance gene Yr5V, respectively. We flow-sort chromosome arm 5VS and sequence it using the Illumina NovaSeq 6000 system that allows us to generate 5VS-specific markers for genetic mapping of Pm5V/Yr5V. Fine mapping shows that Pm5V and Yr5V are closely linked and the location is narrowed to an approximately 0.9 Mb region referencing the sequence of Chinese Spring 5DS. In this region, a NLR gene in scaffold 24,874 of 5VS orthologous to TraesCS5D02G044300 is the most likely candidate gene for Pm5V. Soft- and hard-grained T5DL·5V#5S introgressions confer resistance to both powdery mildew and stripe rust in diverse wheat genetic backgrounds without yield penalty. Meanwhile, significant decrease in plant height and increase in yield were observed in NIL-5DL·5V#5S compared with that in NIL-5DL·5DS. These results indicate that Pm5V/Yr5V lines might have the potential value to facilitate wheat breeding for disease resistance.
