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1.
PLoS One ; 15(4): e0231436, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32298321

RESUMO

Molecular-based taxonomy, specifically DNA barcoding, has streamlined organism identification. For land plants, the recommended 2-locus barcode of rbcL and matK is not suitable for all groups, thus the second subunit of the nuclear internal transcribed spacer (ITS2) has received attention as a possible alternative. To date, evaluations of ITS2 have mostly been limited in scope to specific plant orders/families and single source material. Prior to using ITS2 to routinely characterize land plants present in environmental samples (i.e., DNA metabarcoding), a wet lab protocol optimized for bulk sample types is needed. To address this gap, in this study we determined the broad recoverability across land plants when using published ITS2 primer pairs, and subsequently optimized the PCR reaction constituents and cycling conditions for the best two performing primer pairs (ITS2F/ITSp4 and ITSp3/ITSu4). Using these conditions, both primer pairs were used to characterize land plants present in 17 diverse soils collected from across the US. The resulting PCR amplicons were prepared into libraries and pooled for sequencing on an Illumina® MiniSeq. Our existing bioinformatics workflow was used to process raw sequencing data and taxonomically assign unique ITS2 plant sequences by comparison to GenBank. Given strict quality criteria were imposed on sequences for inclusion in data analysis, only 43.6% and 7.5% of sequences from ITS2F/ITSp4 and ITSp3/ITSu4 respectively remained for taxonomic comparisons; ~7-11% of sequences originated from fungal co-amplification. The number of orders and families recovered did differ between primer pairs, with ITS2F/ITSp4 consistently outperforming ITSp3/ITSu4 by >15%. Primer pair bias was observed in the recovery of certain taxonomic groups; ITS2F/ITSp4 preferentially recovered flowering plants and grasses, whereas ITSp3/ITSu4 recovered more moss taxa. To maximize data recovery and reduce potential bias, we advocate that studies using ITS2 to characterize land plants from environmental samples such as soil use a multiple primer pair approach.


Assuntos
Código de Barras de DNA Taxonômico/métodos , DNA Intergênico/genética , DNA de Plantas/genética , Metagenômica/métodos , Briófitas/classificação , Briófitas/genética , Código de Barras de DNA Taxonômico/normas , DNA Intergênico/química , DNA de Plantas/química , Gleiquênias/classificação , Gleiquênias/genética , Magnoliopsida/classificação , Magnoliopsida/genética , Metagenômica/normas , Reação em Cadeia da Polimerase/métodos , Reação em Cadeia da Polimerase/normas , Solo/química
2.
Nat Commun ; 11(1): 1417, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32184398

RESUMO

Holliday junctions (HJs) are key DNA intermediates in genetic recombination and are eliminated by nuclease, termed resolvase, to ensure genome stability. HJ resolvases have been identified across all kingdoms of life, members of which exhibit sequence-dependent HJ resolution. However, the molecular basis of sequence selectivity remains largely unknown. Here, we present the chloroplast resolvase MOC1, which cleaves HJ in a cytosine-dependent manner. We determine the crystal structure of MOC1 with and without HJs. MOC1 exhibits an RNase H fold, belonging to the retroviral integrase family. MOC1 functions as a dimer, and the HJ is embedded into the basic cleft of the dimeric enzyme. We characterize a base recognition loop (BR loop) that protrudes into and opens the junction. Residues from the BR loop intercalate into the bases, disrupt the C-G base pairing at the crossover and recognize the cytosine, providing the molecular basis for sequence-dependent HJ resolution by a resolvase.


Assuntos
Arabidopsis/enzimologia , Cloroplastos/enzimologia , DNA Cruciforme/metabolismo , Oryza/enzimologia , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Recombinases/química , Recombinases/metabolismo , Soja/enzimologia , Arabidopsis/química , Arabidopsis/genética , Arabidopsis/metabolismo , Sequência de Bases , Cloroplastos/química , Cloroplastos/genética , DNA Cruciforme/química , DNA Cruciforme/genética , DNA de Plantas/química , DNA de Plantas/genética , DNA de Plantas/metabolismo , Oryza/química , Oryza/genética , Oryza/metabolismo , Recombinases/genética , Soja/química , Soja/genética , Soja/metabolismo
3.
PLoS One ; 15(2): e0228776, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32032368

RESUMO

The Mediterranean Basin is a biodiversity hotspot, where islands play a key role because of their high biological diversity, degree of endemicity and human pressure. One of these islands, Sardinia, is a good evolutionary laboratory, especially for the study of complex genera, such as Centaurea. In particular, endemic species of Centaurea sect. Centaurea from Sardinia provides an interesting case study of plant evolution on continental islands. We attempted to clarify the processes leading to the diversification of Centaurea species on Sardinia using bi-parentally inherited nuclear markers and maternally inherited plastid markers. Our plastid results revealed the presence of five lineages of sect. Centaurea on the island. Three of them were defined as three species: C. ferulacea, C. filiformis and C. horrida. The other two lineages highlighted the complex evolutionary history of the two polyploids C. corensis and C. magistrorum. Multiple colonization events from the mainland involving the C. deusta and C. paniculata lineages among others, have led to the diversity of sect. Centaurea on Sardinia. One colonization event likely followed a southern path via the land connection between the mainland, the Calabrian Plate and Sardinia. A second pathway likely followed a northern connection, probably through the Tuscan Archipelago. Implications of these findings on conservation efforts for Centaurea endemics on Sardinia are also discussed.


Assuntos
Evolução Biológica , Centaurea/crescimento & desenvolvimento , Centaurea/classificação , Centaurea/genética , DNA de Plantas/química , DNA de Plantas/isolamento & purificação , DNA de Plantas/metabolismo , Ligação Genética , Ilhas , Itália , Filogenia , Plastídeos/genética , Poliploidia
4.
Mol Phylogenet Evol ; 145: 106727, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31899222

RESUMO

Trichophoreae is a nearly cosmopolitan Cyperaceae tribe that contains ~17 species displaying striking variation in size, inflorescence complexity, and perianth morphology. Although morphologically distinct, the status of its three genera (Cypringlea, Oreobolopsis and Trichophorum) is controversial because recent phylogenetic studies have suggested they might not be reciprocally monophyletic. However, previous analyses have shown conflicting topologies and consistently poor support due to an initial rapid diversification of the tribe. We analysed restriction-site associated DNA sequencing (RADseq) data from nearly all species of the clade, combined with five Sanger-based markers (matK, ndhF, rps16, ETS-1f, ITS) sampled extensively within species. This approach allowed us to resolve deep and shallow relationships within Trichophoreae for the first time, despite an anomaly zone spanning several successive short branches that produced considerable gene tree incongruence. Analyses reveal a primary phylogenetic split of the tribe into two clades roughly corresponding to an East Asian-North American disjunction that dates back to the mid-Miocene, with both clades comprised of a mixture of reduced unispicate and larger taxa with highly compound inflorescences. Morphological characters traditionally used in the circumscription of Trichophoreae genera are shown to be homoplasious. Several of these characters correlate best with climatic conditions, with the most reduced species occurring in open habitats at high latitudes and altitudes. Close relatives with highly compound inflorescences are found in temperate or subtropical forest understories. Cypringlea and Oreobolopsis are deeply nested within Trichophorum, and we merge all three genera into a more broadly circumscribed Trichophorum. We also show that Scirpus filipes is another previously unrecognized East Asian species of Trichophorum with highly compound inflorescences.


Assuntos
Cyperaceae/classificação , Teorema de Bayes , Biodiversidade , Cyperaceae/anatomia & histologia , Cyperaceae/genética , DNA de Plantas/química , DNA de Plantas/metabolismo , DNA Ribossômico/química , DNA Ribossômico/metabolismo , Funções Verossimilhança , Filogenia , Filogeografia , Plastídeos/genética , Análise de Sequência de DNA
5.
PLoS Biol ; 18(1): e3000582, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31995554

RESUMO

In most plants, centromeric DNA contains highly repetitive sequences, including tandem repeats and retrotransposons; however, the roles of these sequences in the structure and function of the centromere are unclear. Here, we found that multiple RNA sequences from centromeric retrotransposons (CRMs) were enriched in maize (Zea mays) centromeres, and back-spliced RNAs were generated from CRM1. We identified 3 types of CRM1-derived circular RNAs with the same back-splicing site based on the back-spliced sequences. These circular RNAs bound to the centromere through R-loops. Two R-loop sites inside a single circular RNA promoted the formation of chromatin loops in CRM1 regions. When RNA interference (RNAi) was used to target the back-splicing site of the circular CRM1 RNAs, the levels of R-loops and chromatin loops formed by these circular RNAs decreased, while the levels of R-loops produced by linear RNAs with similar binding sites increased. Linear RNAs with only one R-loop site could not promote chromatin loop formation. Higher levels of R-loops and lower levels of chromatin loops in the CRM1 regions of RNAi plants led to a reduced localization of the centromeric H3 variant (CENH3). Our work reveals centromeric chromatin organization by circular CRM1 RNAs via R-loops and chromatin loops, which suggested that CRM1 elements might help build a suitable chromatin environment during centromere evolution. These results highlight that R-loops are integral components of centromeric chromatin and proper centromere structure is essential for CENH3 localization.


Assuntos
Centrômero/metabolismo , Cromatina , Conformação de Ácido Nucleico , RNA de Plantas/metabolismo , Retroelementos/genética , Zea mays/genética , Sítios de Ligação/genética , Proteína Centromérica A/genética , Proteína Centromérica A/metabolismo , Cromatina/química , Cromatina/genética , Cromatina/metabolismo , DNA de Plantas/química , DNA de Plantas/genética , DNA de Plantas/metabolismo , Regulação da Expressão Gênica de Plantas , Plantas Geneticamente Modificadas , Processamento de RNA/fisiologia , RNA Circular/genética , RNA Circular/metabolismo , RNA de Plantas/genética , Zea mays/metabolismo
6.
Nucleic Acids Res ; 48(1): 460-471, 2020 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-31733060

RESUMO

As the largest group of MYB family transcription factors, R2R3-MYB proteins play essential roles during plant growth and development. However, the structural basis underlying how R2R3-MYBs recognize the target DNA remains elusive. Here, we report the crystal structure of Arabidopsis WEREWOLF (WER), an R2R3-MYB protein, in complex with its target DNA. Structural analysis showed that the third α-helices in both the R2 and R3 repeats of WER fit in the major groove of the DNA, specifically recognizing the DNA motif 5'-AACNGC-3'. In combination with mutagenesis, in vitro binding and in vivo luciferase assays, we showed that K55, N106, K109 and N110 are critical for the function of WER. Although L59 of WER is not involved in DNA binding in the structure, ITC analysis suggested that L59 plays an important role in sensing DNA methylation at the fifth position of cytosine (5mC). Like 5mC, methylation at the sixth position of adenine (6mA) in the AAC element also inhibits the interaction between WER and its target DNA. Our study not only unravels the molecular basis of how WER recognizes its target DNA, but also suggests that 5mC and 6mA modifications may block the interaction between R2R3-MYB transcription factors and their target genes.


Assuntos
Proteínas de Arabidopsis/química , Arabidopsis/genética , DNA de Plantas/química , Proteínas de Ligação a DNA/química , Regulação da Expressão Gênica de Plantas , Raízes de Plantas/genética , Sequência de Aminoácidos , Animais , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Sítios de Ligação , Galinhas/genética , Galinhas/metabolismo , Sequência Conservada , Cristalografia por Raios X , Metilação de DNA , DNA de Plantas/genética , DNA de Plantas/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Mamíferos , Modelos Moleculares , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Peixe-Zebra/genética , Peixe-Zebra/metabolismo
7.
Genome ; 63(1): 53-60, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31580739

RESUMO

Arctium lappa, commonly called burdock, has a long medicinal and edible history. It has recently gained increasing attention because of its economic value. In this study, we obtained the complete chloroplast genome of A. lappa by Illumina Hiseq. The complete chloroplast genome of A. lappa is a typical circular structure with 152 708 bp in length. The GC content in the whole chloroplast genome of A. lappa is 37.7%. A total of 37 tRNA genes, 8 rRNA genes, and 87 protein-coding genes were successfully annotated. And the chloroplast genome contains 113 unique genes, 19 of which are duplicated in the inverted repeat. The distribution of 39 simple sequence repeats was analysed, and most of them are in the large single-copy (LSC) sequence. An inversion comprising 16 genes was found in the LSC region, which is 26 283 bp long. We performed multiple sequence alignments using 72 common protein-coding genes of 29 species and constructed a Maximum Parsimony (MP) tree. The MP phylogenetic result shows that A. lappa grouped together with Carthamus tinctorius, Centaurea diffusa, and Saussurea involucrata. The chloroplast genome of A. lappa is a valuable resource for further studies in Asteraceae.


Assuntos
Arctium/genética , Genoma de Cloroplastos , Arctium/classificação , Uso do Códon , DNA de Plantas/química , Genes de Plantas , Sequências Repetidas Invertidas , Repetições de Microssatélites , Filogenia , Plantas Medicinais/genética , Sequências Repetitivas de Ácido Nucleico
8.
Talanta ; 206: 120220, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31514891

RESUMO

This work addresses a technological advance applied to the construction of a magnetogenoassay with electrochemical transduction for the maize taxon-specific (HMGA gene) detection using gold-coated magnetic nanoparticles as nanosized platform. Superparamagnetic core-shell Fe3O4@Au nanoparticles (10.4 ±â€¯1.7 nm) were used to assemble the genoassay through the covalent immobilization of HMGA DNA probes onto carboxylated self-assembled monolayers at the nanoparticles surface. A hybridization reaction using sandwich format was selected to prevent inefficient hybridization connected with stable secondary DNA structures using also fluorescein isothiocyanate as DNA signaling tag. The labelling of the hybridization reaction with enzymes allowed the chronoamperometric measurement of the peroxidase activity linked to the nanoplatform located on gold surface. Using this electrochemical magnetogenoassay a linear concentration range from 0.5 to 5 nM and a LOD of 90 pM with a RSD <1.2% was calculated. Certified maize was evaluated without further purification after PCR amplification. This work highlights the efficacy of the electrochemical magnetogenoassay for the HMGA detection, showing its potential as alternative procedure for the verification of the compliance of the legislation.


Assuntos
Técnicas Biossensoriais/métodos , Genes de Plantas , Ouro/química , Proteínas HMGA/genética , Nanopartículas de Magnetita/química , Zea mays/genética , Sequência de Bases , Sondas de DNA/química , Sondas de DNA/genética , DNA de Plantas/química , DNA de Plantas/genética , DNA de Cadeia Simples/química , DNA de Cadeia Simples/genética , Técnicas Eletroquímicas/métodos , Ácidos Nucleicos Imobilizados/química , Ácidos Nucleicos Imobilizados/genética , Limite de Detecção , Hibridização de Ácido Nucleico , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética
9.
Genome Biol Evol ; 12(1): 3663-3676, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31845987

RESUMO

In plants, parasitism triggers the reductive evolution of plastid genomes (plastomes). To disentangle the molecular evolutionary associations between feeding on other plants below- or aboveground and general transitions from facultative to obligate parasitism, we analyzed 34 complete plastomes of autotrophic, root- and stem-feeding hemiparasitic, and holoparasitic Santalales. We observed inexplicable losses of housekeeping genes and tRNAs in hemiparasites and dramatic genomic reconfiguration in holoparasitic Balanophoraceae, whose plastomes have exceptionally low GC contents. Genomic changes are related primarily to the evolution of hemi- or holoparasitism, whereas the transition from a root- to a stem-feeding mode plays no major role. In contrast, the rate of molecular evolution accelerates in a stepwise manner from autotrophs to root- and then stem-feeding parasites. Already the ancestral transition to root-parasitism coincides with a relaxation of selection in plastomes. Another significant selectional shift in plastid genes occurs as stem-feeders evolve, suggesting that this derived form coincides with trophic specialization despite the retention of photosynthetic capacity. Parasitic Santalales fill a gap in our understanding of parasitism-associated plastome degeneration. We reveal that lifestyle-genome associations unfold interdependently over trophic specialization and feeding mode transitions, where holoparasitic Balanophoraceae provide a system for exploring the functional realms of plastomes.


Assuntos
Evolução Molecular , Genomas de Plastídeos , Magnoliopsida/genética , Uso do Códon , DNA de Plantas/química , Raízes de Plantas/parasitologia , Caules de Planta/parasitologia , Sequências Repetitivas de Ácido Nucleico
10.
mBio ; 10(5)2019 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-31594815

RESUMO

The wild legume Lotus japonicus engages in mutualistic symbiotic relationships with arbuscular mycorrhiza (AM) fungi and nitrogen-fixing rhizobia. Using plants grown in natural soil and community profiling of bacterial 16S rRNA genes and fungal internal transcribed spacers (ITSs), we examined the role of the Lotus symbiosis genes RAM1, NFR5, SYMRK, and CCaMK in structuring bacterial and fungal root-associated communities. We found host genotype-dependent community shifts in the root and rhizosphere compartments that were mainly confined to bacteria in nfr5 or fungi in ram1 mutants, while symrk and ccamk plants displayed major changes across both microbial kingdoms. We observed in all AM mutant roots an almost complete depletion of a large number of Glomeromycota taxa that was accompanied by a concomitant enrichment of Helotiales and Nectriaceae fungi, suggesting compensatory niche replacement within the fungal community. A subset of Glomeromycota whose colonization is strictly dependent on the common symbiosis pathway was retained in ram1 mutants, indicating that RAM1 is dispensable for intraradical colonization by some Glomeromycota fungi. However, intraradical colonization by bacteria belonging to the Burkholderiaceae and Anaeroplasmataceae is dependent on AM root infection, revealing a microbial interkingdom interaction. Despite the overall robustness of the bacterial root microbiota against major changes in the composition of root-associated fungal assemblages, bacterial and fungal cooccurrence network analysis demonstrates that simultaneous disruption of AM and rhizobium symbiosis increases the connectivity among taxa of the bacterial root microbiota. Our findings imply a broad role for Lotus symbiosis genes in structuring the root microbiota and identify unexpected microbial interkingdom interactions between root symbionts and commensal communities.IMPORTANCE Studies on symbiosis genes in plants typically focus on binary interactions between roots and soilborne nitrogen-fixing rhizobia or mycorrhizal fungi in laboratory environments. We utilized wild type and symbiosis mutants of a model legume, grown in natural soil, in which bacterial, fungal, or both symbioses are impaired to examine potential interactions between the symbionts and commensal microorganisms of the root microbiota when grown in natural soil. This revealed microbial interkingdom interactions between the root symbionts and fungal as well as bacterial commensal communities. Nevertheless, the bacterial root microbiota remains largely robust when fungal symbiosis is impaired. Our work implies a broad role for host symbiosis genes in structuring the root microbiota of legumes.


Assuntos
Bactérias/crescimento & desenvolvimento , Fungos/crescimento & desenvolvimento , Interações entre Hospedeiro e Microrganismos , Lotus/microbiologia , Interações Microbianas , Proteínas de Plantas/genética , Simbiose , Bactérias/classificação , Bactérias/genética , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Fúngico/química , DNA Fúngico/genética , DNA de Plantas/química , DNA de Plantas/genética , DNA Ribossômico/química , DNA Ribossômico/genética , DNA Espaçador Ribossômico/química , DNA Espaçador Ribossômico/genética , Fungos/classificação , Fungos/genética , Genótipo , Lotus/genética , Filogenia , Raízes de Plantas/microbiologia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Microbiologia do Solo
11.
Plant Mol Biol ; 101(4-5): 415-437, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31542868

RESUMO

KEY MESSAGE: Naturally transgenic plant species occur on an unexpectedly large scale. Agrobacterium-mediated gene transfer leads to the formation of crown galls or hairy roots, due to expression of transferred T-DNA genes. Spontaneous regeneration of transformed cells can produce natural transformants carrying cellular T-DNA (cT-DNA) sequences of bacterial origin. This particular type of horizontal gene transfer (HGT) could play a role in plant evolution. However, the material available today is not enough for generalizations concerning the role of Agrobacterium in HGT from bacteria to plants. In this study, we searched for T-DNA-like genes in the sequenced genomes of dicots and monocots. We demonstrate the presence of cT-DNAs in 23 out of 275 dicot species, within genera Eutrema, Arachis, Nissolia, Quillaja, Euphorbia, Parasponia, Trema, Humulus, Psidium, Eugenia, Juglans, Azadirachta, Silene, Dianthus, Vaccinium, Camellia, and Cuscuta. Analysis of transcriptome data of 356 dicot species yielded 16 additional naturally transgenic species. Thus, HGT from Agrobacterium to dicots is remarkably widespread. Opine synthesis genes are most frequent, followed by plast genes. Species in the genera Parasponia, Trema, Camellia, Azadirachta, Quillaja, and Diospyros contain a combination of plast and opine genes. Some are intact and expressed, but the majority have internal stop codons. Among the sequenced monocot species, Dioscorea alata (greater yam) and Musa acuminata (banana) also contain T-DNA-like sequences. The identified examples are valuable material for future research on the role of Agrobacterium-derived genes in plant evolution, for investigations on Agrobacterium strain diversity, and for studies on the function and evolution of cT-DNA genes in natural transformants.


Assuntos
Agrobacterium/genética , Genoma de Planta , Transformação Genética , Evolução Biológica , DNA Bacteriano/química , DNA de Plantas/química , Transferência Genética Horizontal , Plantas Geneticamente Modificadas/genética , Análise de Sequência de DNA , Transcriptoma
12.
J Genet ; 982019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31544779

RESUMO

To detect the genetic variation and relationships among different Salvia ecotypes/species, the gene targeted CAAT box derived polymorphism (CBDP) markers were employed in terms of their efficiency. In this study, 25 CBDP primers amplified a total of 323 different polymorphic fragments that discriminate all 26 Salvia ecotypes/species and produced an informative and differentiated dendrogram and population structure. The CBDP markers were found to be effective in Salvia genetic diversity estimation with regard to the averages polymorphism (100%), polymorphism information content (PIC = 0.89), marker index (MI = 4.5) and the effective multiplex ratio (EMR = 5.01) which were higher than other reported markers on Salvia. The extent of heterozygosity (0.034≤H≤0.223) and Shannon index (0.042≤I≤0.278) indicated a high level of genetic variation among Salvia species. The species containing the highest basic chromosome number (X = 12) revealed the highest values for the number of different (Na) and effective (N e) alleles, Shannon index (I), and heterozygosity (H). Additionally, the tetraploid species showed high values of N a, Ne, I and H compared to the diploid species. Mean of gene differentiation (Gst) among Salvia species was 0.792, and the estimation of gene flow (Nm) was 0.13, indicating high genetic differentiation. Remarkably, similar results were obtained from the principal co-ordinate analysis (PCoA) as compared with the cluster analysis, in which all different Salvia species formed individual groups. In conclusion, because the CBDP markers are derived from the gene containing regions of the genome, consequently, the high genetic diversity among studied Salvia species would be more useful for crop improvement programmes, such as hybridization between species and QTL mapping. The potential of CBDPs for analysing the phylogeny and genetic diversity of Salvia species is another key result with practical implications.


Assuntos
Polimorfismo Genético , Salvia/genética , Alelos , Análise por Conglomerados , DNA de Plantas/química , Fluxo Gênico , Marcadores Genéticos , Genética Populacional , Motivos de Nucleotídeos , Filogenia
13.
Int J Mol Sci ; 20(19)2019 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-31546611

RESUMO

Methylation of cytosine (5-meC) is a critical epigenetic modification in many eukaryotes, and genomic DNA methylation landscapes are dynamically regulated by opposed methylation and demethylation processes. Plants are unique in possessing a mechanism for active DNA demethylation involving DNA glycosylases that excise 5-meC and initiate its replacement with unmodified C through a base excision repair (BER) pathway. Plant BER-mediated DNA demethylation is a complex process involving numerous proteins, as well as additional regulatory factors that avoid accumulation of potentially harmful intermediates and coordinate demethylation and methylation to maintain balanced yet flexible DNA methylation patterns. Active DNA demethylation counteracts excessive methylation at transposable elements (TEs), mainly in euchromatic regions, and one of its major functions is to avoid methylation spreading to nearby genes. It is also involved in transcriptional activation of TEs and TE-derived sequences in companion cells of male and female gametophytes, which reinforces transposon silencing in gametes and also contributes to gene imprinting in the endosperm. Plant 5-meC DNA glycosylases are additionally involved in many other physiological processes, including seed development and germination, fruit ripening, and plant responses to a variety of biotic and abiotic environmental stimuli.


Assuntos
5-Metilcitosina/metabolismo , Desmetilação do DNA , DNA Glicosilases/metabolismo , DNA de Plantas/genética , Plantas/enzimologia , DNA Glicosilases/química , Metilação de DNA , DNA de Plantas/química , Endosperma/metabolismo , Regulação da Expressão Gênica de Plantas , Instabilidade Genômica/genética , Óvulo Vegetal/metabolismo , Pólen/metabolismo , Estresse Fisiológico/genética
14.
Mol Phylogenet Evol ; 139: 106567, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31330266

RESUMO

The scaly tree ferns, Cyatheaceae, form a large natural group of ca. 640 species. They comprise an ideal model for studying the biogeography of plants due to their broad distribution across the tropical to south-temperate areas of the world. However, tracing the biogeographic history of this highly diversified group has been hampered by unresolved evolutionary relationships among the major clades. Here, we generated transcriptome sequences of five species in three genera of Cyatheaceae (Alsophila, Gymnosphaera, and Sphaeropteris) and used them to search for single-copy nuclear loci for phylogenetic reconstruction. We identified a total of 818 candidate single-copy loci across multiple Cyatheaceae species. To test their phylogenetic utility, we further obtained sequence data of 12 of these loci for 76 samples representing all 13 known species of scaly tree ferns in China and Vietnam. Phylogenetic analyses based on multispecies coalescent and, alternatively, concatenation models yielded congruent results with high resolution. Additionally, we used the 12 loci to identify genetic signals of hybridization. Overall, our results demonstrated that multiple, single-copy loci are informative and efficient tools for phylogenetic or evolutionary studies of scaly tree ferns.


Assuntos
Núcleo Celular/genética , Gleiquênias/genética , Transcriptoma , China , DNA de Plantas/química , DNA de Plantas/metabolismo , Gleiquênias/classificação , Fluxo Gênico , Filogenia , Vietnã
15.
Anal Chim Acta ; 1078: 24-31, 2019 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-31358225

RESUMO

A novel electrochemical DNA biosensor was developed and MON89788 of soybean transgenic gene sequence was detected based on a strategy of rolling circle amplification (RCA) and gold nanoparticle cube (AuNPC)-labeled multiple probes. First, the mercapto-modified capture DNA was immobilized on the surface of the Fe3O4@Au magnetic nanoparticles via an Au-S bond, and the capture DNA was opened and complementarily hybridized with the target DNA to form a double-stranded DNA. In the 10 × reaction buffer, Exonuclease III (ExoIII) specifically recognized and sheared the double-stranded DNA to release the target DNA, which led to the next round of reaction. Afterward, AuNP cube-loaded ssDNA (AuNPC/DNA) was added with the rolling circle reaction with the help of Phi29 DNA polymerase and T4 ligase. Finally, [Ru(NH3)6]3+ was attracted directly by the anionic phosphate of ssDNA via electrostatic interaction. The determination was carried out by using chronocoulometry (CC), and the CC signal was recorded. The mass amount of DNA strands extended infinitely on the AuNPs cube and numerous [Ru(NH3)6]3+ were absorbed, thus the detected signal was highly amplified. The corresponding CC signal showed a good linear relationship with the logarithm of the target DNA concentration in the range of 1 × 10-16 to 1 × 10-7 mol L-1, with a detection limit of 4.5 × 10-17 mol L-1. Specific gene sequence of MON89788 in soybean samples was determined, and the recoveries ranged from 97.3% to 102.0%. This sensor is one of the most sensitive sensors for genetic sequence assessment at present. Moreover, it demonstrates good selectivity, stability, and reproducibility.


Assuntos
Técnicas Biossensoriais/métodos , DNA de Plantas/análise , Técnicas Eletroquímicas/métodos , Plantas Geneticamente Modificadas/genética , Soja/genética , Sequência de Bases , Calibragem , Sondas de DNA/química , Sondas de DNA/genética , DNA de Plantas/química , DNA de Plantas/genética , Exodesoxirribonucleases/química , Ouro/química , Limite de Detecção , Nanopartículas de Magnetita/química , Técnicas de Amplificação de Ácido Nucleico/métodos , Hibridização de Ácido Nucleico , Oligodesoxirribonucleotídeos/química , Oligodesoxirribonucleotídeos/genética , Reprodutibilidade dos Testes , Compostos de Rutênio/química
16.
Mol Phylogenet Evol ; 139: 106572, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31351183

RESUMO

The Eurasian steppes occupy a significant portion of the worldwide land surface and their biota have been affected by specific past range dynamics driven by ice ages-related climatic fluctuations. The dynamic alterations in conditions during the Pleistocene often triggered reticulate evolution and whole genome duplication events. Employing genomic, genetic and cytogenetic tools as well as morphometry we investigate the intricate evolution of Astragalus onobrychis, a widespread Eurasian steppe plant with diploid, tetraploid and octoploid cytotypes. To analyse the heteroploid RADseq dataset we employ both genotype-based and genotype-free methods that result in highly consistent results, and complement our inference with information from the plastid ycf1 region. We uncover a complex and reticulate evolutionary history, including at least one auto-tetraploidization event and two allo-octoploidization events; one of them involved also genetic contributions from other species, most likely A. goktschaicus. The present genetic structure points to the existence of four main clades within A. onobrychis, which only partly correspond to different ploidies. Time-calibrated diffusion models suggest that diversification within A. onobrychis was associated with ice age-related climatic fluctuations during the last million years. We finally argue for the usefulness of uniparentally inherited plastid markers, even in the genomic era, especially when investigating heteroploid systems.


Assuntos
Astrágalo (Planta)/genética , Cromossomos de Plantas , Ásia , Astrágalo (Planta)/anatomia & histologia , Astrágalo (Planta)/classificação , DNA de Plantas/química , DNA de Plantas/metabolismo , Europa (Continente) , Filogenia , Plastídeos/genética , Poliploidia , Análise de Componente Principal
17.
Mol Biotechnol ; 61(9): 694-702, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31256331

RESUMO

Molecular characteristics including information of insertion site, flanking sequence, and copy numbers are the base for the safety assessment and subsequent monitoring of genetically modified organisms (GMOs), which has to be revealed thoroughly in a case-by-case manner. Although both polymerase chain reaction (PCR)-based and next-generation sequencing (NGS)-based approaches are proven to be effective in the molecular characterization of most of GM events, they often fail to work with GM maize events, mainly due to the genome complexity. In this study, by using NGS, we successfully identified the 3' end T-DNA insertion site and flanking sequence of a GM maize event IE09S034, which were confirmed by PCR amplification and Sanger sequencing. Notably, insertions of unintended exogenous elements were revealed in this event although the single copy of target exogenous genes was also confirmed by digital PCR. The output of this study provides novel and important genetic evidence for the safety assessment and monitoring of GM maize event IE09S034.


Assuntos
Mapeamento Cromossômico/métodos , DNA Bacteriano/genética , DNA de Plantas/química , Genoma de Planta , Mutagênese Insercional , Zea mays/genética , Sequência de Bases , DNA de Plantas/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase/métodos
18.
BMC Genomics ; 20(1): 508, 2019 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-31215402

RESUMO

BACKGROUND: DNA methylation is an important epigenetic modification. Recently the developed single-molecule real-time (SMRT) sequencing technology provided an efficient way to detect DNA N6-methyladenine (6mA) modification that played an important role in epigenetic and positively regulated gene expression. In addition, the gene expression was also regulated by genetic variation. However, the relationship between DNA 6mA modification and variation is still unknown. RESULTS: We collected the SMRT long-reads DNA, Illumina short reads DNA and RNA datasets from the young leaves of Herrania umbratica, and used them to detect 35,654 6mA modification sites, 829,894 DNA variations and 60,672 RNA variations respectively, among which, there are 303 DNA variations and 19 RNA variations with 6mA modification, and 57,468 transmitted genetic variations from DNA to RNA. The results illustrated that the genes with 6mA modification were significant disadvantage to mutate than those genes without modification (p-value< 4.9e-08). And result from the linear regression model showed the 6mA densities of genes were associated with the transmitted variations type 0/1 to 1/1 (p-value < 0.001). CONCLUSIONS: The variations of DNA and RNA in genes with 6mA modification were significant less than those in unmodified genes. Furthermore, the variations in 6mA modified genes were easily transmitted from DNA to RNA, especially the transmitted variation from DNA heterozygote to RNA homozygote.


Assuntos
Adenosina/análogos & derivados , DNA de Plantas/genética , DNA de Plantas/metabolismo , Variação Genética/genética , Genoma de Planta/genética , Magnoliopsida/genética , RNA de Plantas/genética , Adenosina/metabolismo , DNA Intergênico/genética , DNA Intergênico/metabolismo , DNA de Plantas/química , Heterozigoto , Homozigoto , Magnoliopsida/metabolismo
19.
Mol Phylogenet Evol ; 139: 106541, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31228555

RESUMO

Eremitis, Pariana, and Parianella are herbaceous bamboos (tribe Olyreae) included in the subtribe Parianinae, which is characterized by the presence of fimbriae at the apex of the leaf sheaths and exclusively spiciform synflorescences. We analyzed 43 samples of herbaceous and woody bamboos in order to infer relationships within the Parianinae, based on combined data from the nuclear ribosomal internal transcribed spacer (ITS) and plastid DNA (rpl32-trnL and trnD-trnT spacers). Bayesian inference, maximum likelihood, and maximum parsimony methods were applied, and macro- and micromorphological aspects were also analyzed, including the ectexine patterns of pollen grains. Parianinae is represented by three well-supported lineages in our analyses: (1) Parianella, endemic to southern Bahia, Brazil; (2) Pariana sensu stricto with a broad distribution in southern Central America and northern South America, especially in the Amazon region; and (3) Eremitis, endemic to the Brazilian Atlantic Forest, from the states of Pernambuco to Rio de Janeiro, including one species previously described as a member of Pariana. Our molecular phylogeny showed that Pariana, as historically circumscribed, is not monophyletic, by recovering Pariana sensu stricto as strongly supported and sister to Eremitis + Pariana multiflora, with Parianella sister to the Pariana-Eremitis clade. Morphological features of their synflorescences and differences in ectexine patterns characterize each lineage. Based on all these characters and the phylogenetic results, Pariana multiflora, endemic to the state of Espírito Santo, Brazil, is transferred to Eremitis.


Assuntos
Poaceae/classificação , Teorema de Bayes , Brasil , Núcleo Celular/genética , América Central , DNA de Plantas/química , Filogenia , Plastídeos/genética , Poaceae/anatomia & histologia , Poaceae/genética , Poaceae/ultraestrutura , Pólen/ultraestrutura , Análise de Sequência de DNA , América do Sul
20.
Plant J ; 99(2): 201-215, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31134682

RESUMO

Hexaploid common wheat is one of the most important food crops worldwide. Common wheat domestication began in the Fertile Crescent of the Near East approximately 10 000 years ago and then spread west into Europe and eastward into East Asia and China. However, the possible spreading route into and within China is still unclear. In this study, we successfully extracted DNA from single ancient wheat seeds and sequenced the whole genome of seven ancient samples from Xiaohe and Gumugou cemeteries in Xinjiang, China. Genomic inference and morphological observation confirmed their identity as hexaploid common wheat grown in prehistoric China at least 3200 years before present (BP). Phylogenetic and admixture analyses with RNA-seq data of modern hexaploid wheat cultivars from both China and Western countries demonstrated a close kinship of the ancient wheat to extant common wheat landraces in southwestern China. The highly similar allelic frequencies in modern landraces of the Qinghai-Tibetan plateau with the ancient wheat support the previously suggested southwestern spreading route into highland China. A subsequent dispersal route from the Qinghai-Tibetan plateau margins to the Yangtze valley was proposed in this study. Furthermore, the common wheat populations grown in the Middle and Lower Yangtze valley wheat zones were also proposed to be established by population admixture with the wheat grown in the Upper Yangtze valley. Our study reports ancient common wheat sequences at a genome-wide scale, providing important information on the origin, dispersal, and genetic improvement under cultivation of present-day wheat landraces grown in China.


Assuntos
Genoma de Planta , Triticum/genética , China , DNA de Plantas/química , Frequência do Gene , Filogenia , Dispersão de Sementes , Sementes/anatomia & histologia , Sementes/genética , Análise de Sequência de RNA , Triticum/anatomia & histologia
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