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1.
BMC Plant Biol ; 19(1): 246, 2019 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-31182023

RESUMO

BACKGROUND: Rapid-cycling Brassica rapa (RCBr), also known as Wisconsin Fast Plants, are small robust plants with a short lifecycle that are widely used in biology teaching. RCBr have been used for decades but there are no published reports of RCBr genetic transformation. Agrobacterium-mediated vacuum infiltration has been used to transform pakchoi (Brassica rapa ssp. chinensis) and may be suitable for RCBr transformation. The floral dip transformation method, an improved version of vacuum infiltration, could make the procedure easier. RESULTS: Based on previous findings from Arabidopsis and pakchoi, plants of three different ages were inoculated with Agrobacterium. Kanamycin selection was suboptimal with RCBr; a GFP screen was used to identify candidate transformants. RCBr floral bud dissection showed that only buds with a diameter less than 1 mm carried unsealed carpels, a key point of successful floral dip transformation. Plants across a wide range of inflorescence maturities but containing these immature buds were successfully transformed, at an overall rate of 0.1% (one per 1000 T1 seeds). Transformation was successful using either vacuum infiltration or the floral dip method, as confirmed by PCR and Southern blot. CONCLUSION: A genetic transformation system for RCBr was established in this study. This will promote development of new biology teaching tools as well as basic biology research on Brassica rapa.


Assuntos
Agrobacterium/fisiologia , Brassica rapa/genética , Brassica rapa/microbiologia , Engenharia Genética/métodos , Transformação Genética , Southern Blotting , Flores/genética , Reação em Cadeia da Polimerase
2.
BMC Genomics ; 20(1): 437, 2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-31146687

RESUMO

BACKGROUND: For spring-type Chinese cabbage production, premature bolting refers to the excessive elongation of dwarf stems before harvesting. Although quantitative trait loci (QTL) mapping for bolting-related traits have been studied extensively, the main flower stalk length (MFSL) have been rarely investigated. Two inbred lines, 06-247 and He102, have significant differences in the MFSL. In this study, these two materials were selected as parental lines for the construction of a recombinant inbred line (RIL) mapping population. High-density mapping of QTL for the MFSL was performed based on the deep resequencing of parental lines and specific locus-amplified fragment sequencing (SLAF-Seq) of individual recombination inbred lines. RESULTS: An F7 population consisting of 150 lines was developed. Deep resequencing of parental lines produced 21.08 gigabases, whereas SLAF-Seq produced an average of 428.35 million bases for each progeny. The total aligned data from the parental lines identified 1,082,885 high-quality single nucleotide polymorphisms (SNPs) between parental lines. Out of these, 5392 SNP markers with a segregation type of aa×bb and average integrity of > 99% were suitable for the genetic linkage map construction. The final map contained 10 linkage groups (LGs) was 1687.82 cM in length with an average distance of 0.32 cM between adjacent markers. Based on the high-density map, nine QTLs for MFSL were found to be distributed on seven chromosomes, and two major-effect QTLs were identified for the first time. The physical distance between adjacent markers of two major-effect QTLs was 44.37 kbp and 121.91 kbp, respectively. Approximately 2056 and 6769 SNP markers within confidence intervals were identified according to the results of parental line resequencing, which involved 24 and 199 mutant genes. CONCLUSIONS: The linkage map constructed in this study has the highest density in Chinese cabbage to date. Two major-effect QTLs for MFSL in Chinese cabbage were also identified. Among these, a novel QTL associated with bolting mapped on LG A04 was identified based on MFSL. The results of this study provide an important platform for gene/QTL mapping and marker-assisted selection (MAS) breeding for bolting-resistant Chinese cabbage.


Assuntos
Brassica rapa/genética , Locos de Características Quantitativas , Brassica rapa/anatomia & histologia , Mapeamento Cromossômico , Ligação Genética , Técnicas de Genotipagem , Sequenciamento de Nucleotídeos em Larga Escala , Fenótipo , Caules de Planta/anatomia & histologia , Polimorfismo de Nucleotídeo Único
3.
Biomed Res Int ; 2019: 2965035, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31073524

RESUMO

The type 2C protein which belongs to the major group of protein phosphatases (PP2C) plays a vital role in abscisic acid (ABA) signaling and signal transductions processes. In the present study, 131 PP2C genes were identified in total in Brassica rapa and categorized into thirteen subgroups based on their phylogenetic relationships. These B. rapa PP2C are structurally conserved based on amino acid sequence alignment, phylogenetic analysis, and conserved domains. Moreover, we utilized previously reported RNA-sequence data on various tissues (root, stem, leaf, flower, and silique), which suggests overlapping expression pattern in 29 paralogous gene pairs. The qRT-PCR validation of 15 paralogous gene pairs depicts distinct expression patterns in response to various abiotic stresses, such as heat, cold, ABA, and drought. Interestingly, stress-responsive BraPP2C candidate genes were also identified, suggesting their significance in stress-tolerance mechanism in B. rapa. The evolutionary analysis for 15 paralogous gene pairs suggested that only three pairs have the positive selection and remaining were purifying in nature. The presented results of this study hasten our understanding of the molecular evolution of the PP2C gene family in B. rapa. Thus, it will be ultimately helping in future research for facilitating the functional characterization of BraPP2C genes in developing the abiotic stress tolerant plants.


Assuntos
Brassica rapa/genética , Evolução Molecular , Filogenia , Proteína Fosfatase 2C/genética , Sequência de Aminoácidos/genética , Cromossomos de Plantas/genética , Flores/genética , Regulação da Expressão Gênica de Plantas , Genoma de Planta/genética , Família Multigênica/genética , Proteína Fosfatase 2C/classificação
4.
BMC Genomics ; 20(1): 343, 2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31064320

RESUMO

BACKGROUND: Caixin and Zicaitai (Brassica rapa) belong to Southern and Central China respectively. Zicaitai contains high amount of anthocyanin in leaf and stalk resulting to the purple color. Stalk is the major edible part and stalk color is an economically important trait for the two vegetables. The aim of this study is to construct a high density genetic map using the specific length amplified fragment sequencing (SLAF-seq) technique to explore genetic basis for anthocyanin pigmentation traits via quantitative trait loci (QTL) mapping. RESULTS: We constructed a high generation linkage map with a mapping panel of F2 populations derived from 150 individuals of parental lines "Xianghongtai 01" and "Yinong 50D" with purple and green stalk respectively. The map was constructed containing 4253 loci, representing 10,940 single nucleotide polymorphism (SNP) markers spanning 1030.04 centiMorgans (cM) over 10 linkage groups (LGs), with an average distance between markers of 0.27 cM. Quantitative trait loci (QTL) analysis revealed that a major locus on chromosome 7 and 4 minor QTLs explaining 2.69-61.21% of phenotypic variation (PVE) were strongly responsible for variation in stalk color trait. Bioinformatics analysis of the major locus identified 62 protein-coding genes. Among the major locus, there were no biosynthetic genes related to anthocyanin. However, there were several transcription factors like helix-loop-helix (bHLH) bHLH, MYB in the locus. Seven predicted candidate genes were selected for the transcription level analysis. Only bHLH49 transcription factor, was significantly higher expressed in both stalks and young leaves of Xianghongtai01 than Yinong50D. An insertion and deletion (InDel) marker developed from deletion/insertion in the promoter region of bHLH49 showed significant correlation with the stalk color trait in the F2 population. CONCLUSION: Using the constructed high-qualified linkage map, this study successfully identified QTLs for stalk color trait. The identified valuable markers and candidate genes for anthocyanin accumulation in stalk will provide useful information for molecular regulation of anthocyanin biosynthesis. Overall our findings will lay a foundation for functional gene cloning, marker-assisted selection (MAS) and molecular breeding of important economic traits in B. rapa.


Assuntos
Antocianinas/metabolismo , Brassica rapa/anatomia & histologia , Brassica rapa/genética , Cromossomos de Plantas , Locos de Características Quantitativas , Brassica rapa/crescimento & desenvolvimento , Mapeamento Cromossômico , Ligação Genética , Marcadores Genéticos , Técnicas de Genotipagem , Fenótipo , Pigmentação , Análise de Sequência de DNA
5.
Planta ; 250(2): 603-628, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31139927

RESUMO

MAIN CONCLUSION: Compared with its parents, Brassica hexaploid underwent significant AS changes, which may provide diversified gene expression regulation patterns and could enhance its adaptability during evolution Polyploidization is considered a significant evolution force that promotes species formation. Alternative splicing (AS) plays a crucial role in multiple biological processes during plant growth and development. To explore the effects of allopolyploidization on the AS patterns of genes, a genome-wide AS analysis was performed by RNA-seq in Brassica hexaploid and its parents. In total, we found 7913 (27540 AS events), 14447 (70179 AS events), and 13205 (60804 AS events) AS genes in Brassica rapa, Brassica carinata, and Brassica hexaploid, respectively. A total of 920 new AS genes were discovered in Brassica hexaploid. There were 56 differently spliced genes between Brassica hexaploid and its parents. In addition, most of the alternative 5' splice sites were located 4 bp upstream of the dominant 5' splice sites, and most of the alternative 3' splice sites were located 3 bp downstream of the dominant 3' splice sites in Brassica hexapliod, which was similar to B. carinata. Furthermore, we cloned and sequenced all amplicons from the RT-PCR products of GRP7/8, namely, Bol045859, Bol016025 and Bol02880. The three genes were found to produce AS transcripts in a new way. The AS patterns of genes were diverse between Brassica hexaploid and its parents, including the loss and gain of AS events. Allopolyploidization changed alternative splicing sites of pre-mRNAs in Brassica hexaploid, which brought about alterations in the sequences of transcripts. Our study provided novel insights into the AS patterns of genes in allopolyploid plants, which may provide a reference for the study of polyploidy adaptability.


Assuntos
Processamento Alternativo , Brassica/genética , Regulação da Expressão Gênica de Plantas/genética , Genoma de Planta/genética , Adaptação Fisiológica , Evolução Biológica , Brassica/fisiologia , Brassica rapa/genética , Brassica rapa/fisiologia , Poliploidia
6.
Ecotoxicol Environ Saf ; 180: 88-94, 2019 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-31078020

RESUMO

Cadmium (Cd) is a dangerous transition element that causes environmental and health problems due to its high mobility in the soil-plant system. In plants, Cd causes serious alterations in physiological processes, affecting different vital functions such as photosynthesis. Species such as Brassica juncea and Brassica rapa have been selected as suitable plants for phytoremediation purposes due to their ability to tolerate the toxic effect of heavy metals. In order to improve this strategy, techniques of plant mutagenesis such as TILLING (Targeting Induced Local Lessions in Genomes) have been employed. In the present work we studied the role of the HMA4 gene in the tolerance to Cd toxicity (100 µM CdCl2) using a TILLING mutant of B. rapa (BraA.hma4a-3). These mutant plants presented a lower biomass reduction and a higher Cd concentration in leaves. An increase in the GSH/GSSG ratio, in the content of photosynthetic pigments and a reduction of oxidative stress was observed, as well as a better photosynthetic index, confirming that BraA.hma4a-3 plants showed a higher tolerance to Cd. In conclusion, according to the results obtained in this work, BraA.hma4a-3 plants could be used for phytoremediation purposes of Cd contaminated soils.


Assuntos
Brassica rapa/efeitos dos fármacos , Cádmio/toxicidade , Genes de Plantas , Fotossíntese/efeitos dos fármacos , Poluentes do Solo/toxicidade , Biodegradação Ambiental , Brassica rapa/genética , Cádmio/metabolismo , Mutação , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Fotossíntese/genética , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/metabolismo , Solo/química , Poluentes do Solo/metabolismo
7.
BMC Genomics ; 20(1): 264, 2019 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-30943898

RESUMO

BACKGROUND: Genic male sterility (GMS) line is an important approach to utilize heterosis in Brassica rapa, one of the most widely cultivated vegetable crops in Northeast Asia. However, the molecular genetic mechanisms of GMS remain to be largely unknown. RESULTS: Detailed phenotypic observation of 'Bcajh97-01A/B', a B. rapa genic male sterile AB line in this study revealed that the aberrant meiotic cytokinesis and premature tapetal programmed cell death occurring in the sterile line ultimately resulted in microspore degeneration and pollen wall defect. Further gene expression profile of the sterile and fertile floral buds of 'Bcajh97-01A/B' at five typical developmental stages during pollen development supported the result of phenotypic observation and identified stage-specific genes associated with the main events associated with pollen wall development, including tapetum development or functioning, callose metabolism, pollen exine formation and cell wall modification. Additionally, by using ChIP-sequencing, the genomic and gene-level distribution of trimethylated histone H3 lysine 4 (H3K4) and H3K27 were mapped on the fertile floral buds, and a great deal of pollen development-associated genes that were covalently modified by H3K4me3 and H3K27me3 were identified. CONCLUSIONS: Our study provids a deeper understanding into the gene expression and regulation network during pollen development and pollen wall formation in B. rapa, and enabled the identification of a set of candidate genes for further functional annotation.


Assuntos
Brassica rapa/genética , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Pólen/fisiologia , Brassica rapa/crescimento & desenvolvimento , Brassica rapa/metabolismo , Imunoprecipitação da Cromatina , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Infertilidade das Plantas , Proteínas de Plantas/metabolismo , Pólen/genética , Transcriptoma
8.
Int J Mol Sci ; 20(7)2019 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-30987159

RESUMO

Glycoside hydrolase family 1 (GH1) ß-glucosidases (BGLUs) are encoded by a large number of genes, and are involved in many developmental processes and stress responses in plants. Due to their importance in plant growth and development, genome-wide analyses have been conducted in model plants (Arabidopsis and rice) and maize, but not in Brassica species, which are important vegetable crops. In this study, we systematically analyzed B. rapa BGLUs (BrBGLUs), and demonstrated the involvement of several genes in pollen development. Sixty-four BrBGLUs were identified in Brassica databases, which were anchored onto 10 chromosomes, with 10 tandem duplications. Phylogenetic analysis revealed that 64 genes were classified into 10 subgroups, and each subgroup had relatively conserved intron/exon structures. Clustering with Arabidopsis BGLUs (AtBGLUs) facilitated the identification of several important subgroups for flavonoid metabolism, the production of glucosinolates, the regulation of abscisic acid (ABA) levels, and other defense-related compounds. At least six BrBGLUs might be involved in pollen development. The expression of BrBGLU10/AtBGLU20, the analysis of co-expressed genes, and the examination of knocked down Arabidopsis plants strongly suggests that BrBGLU10/AtBGLU20 has an indispensable function in pollen development. The results that are obtained from this study may provide valuable information for the further understanding of ß-glucosidase function and Brassica breeding, for nutraceuticals-rich Brassica crops.


Assuntos
Brassica rapa/enzimologia , Brassica rapa/genética , Estudo de Associação Genômica Ampla , Família Multigênica , Pólen/crescimento & desenvolvimento , Pólen/genética , beta-Glucosidase/genética , Cromossomos de Plantas/genética , Éxons/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Ontologia Genética , Genes de Plantas , Íntrons/genética , Filogenia
9.
J Plant Physiol ; 237: 51-60, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31022665

RESUMO

Cation/H+ exchanger transporters (CAXs) are crucial in Ca2+ homeostasis and in the generation of Ca2+ profiles involved in signalling processes. Given the crucial role of CAX1 in Ca2+ homeostasis, CAX1 modifications could have effects on plant metabolism. Three Brassica rapa mutants for CAX1 were obtained through TILLING. The aim of this work is to assess the effect of the different mutations and different Ca2+ doses on plant metabolism. For this, the mutants and the parental line were grown under low, control and high Ca2+ doses and parameters related to nitrogen (N) and tricarboxylic acid (TCA) metabolisms, and amino acid (AAs) and phytohormone profiles were measured. The results show that BraA.cax1a mutations affect metabolism especially under high Ca2+ dose. Thus, BraA.cax1a-7 inhibited some N metabolism enzymes and activated photorespiration activity. On the opposite side, BraA.cax1a-12 mutation provides a better tolerance to high Ca2+ dose. This tolerance could be provided by an improved N and TCA metabolisms enzymes, and a higher glutamate, malate, indole-3-acetic acid and abscisic acid concentrations. Therefore, BraA.cax1a-12 mutation could be used for B. rapa improving; the metabolomics changes observed in this mutant could be responsible for a better tolerance to high Ca2+.


Assuntos
Brassica rapa/genética , Brassica rapa/metabolismo , Cálcio/metabolismo , Proteínas de Transporte de Cátions/genética , Proteínas de Plantas/genética , Proteínas de Transporte de Cátions/metabolismo , Relação Dose-Resposta a Droga , Mutação , Proteínas de Plantas/metabolismo
10.
Plant Genome ; 12(1)2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30951085

RESUMO

Epigenetic regulations in the form of changes in differential expression of noncoding RNAs (ncRNAs) are an essential mechanism of stress response in plants. Previously we showed that heat treatment in L. results in the differential processing and accumulation of ncRNA fragments (ncRFs) stemming from transfer RNAs (tRNAs), ribosomal RNAs (rRNAs), small nuclear RNAs (snRNAs), and small nucleolar RNAs (snoRNAs). In this work, we analyzed whether ncRFs are differentially expressed in the progeny of heat-stressed plants. We found significant changes in the size of tRF reads and a significant decrease in the percentage of tRFs mapping to tRNA-Ala, tRNA-Arg, and tRNA-Tyr and an increase in tRFs mapping to tRNA-Asp. The enrichment analysis showed significant differences in processing of tRFs from tRNA, tRNA, tRNA, tRNA, tRNA, and tRNA isoacceptors. Analysis of potential targets of tRFs showed that they regulate brassinosteroid metabolism, the proton pump ATPase activity, the antiporter activity, the mRNA decay activity as well as nucleosome positioning and the epigenetic regulation of transgenerational response. Gene ontology term analysis of potential targets demonstrated a significant enrichment in tRFs that potentially targeted a cellular component endoplasmic reticulum (ER) and in small nucleolar RNA fragments (snoRFs), the molecular function protein binding. To summarize, our work demonstrated that the progeny of heat-stressed plants exhibit changes in the expression of tRFs and snoRFs but not of small nuclear RNA fragments (snRFs) or ribosomal RNA fragments (rRFs) and these changes likely better prepare the progeny of stressed plants to future stress encounters.


Assuntos
Brassica rapa/genética , Resposta ao Choque Térmico/genética , RNA de Plantas/biossíntese , RNA não Traduzido/biossíntese , Brassica rapa/fisiologia , Ontologia Genética , Genes de Plantas , RNA Mensageiro/metabolismo , RNA de Plantas/genética , RNA não Traduzido/genética , Reprodução
11.
BMC Genomics ; 20(1): 227, 2019 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-30890148

RESUMO

BACKGROUND: Abiotic stresses like drought, heat, cold and salinity cause major productivity loss in the rapeseed-mustard crops (Brassica). Major efforts have been made in the past to identify genes that provide resistance against such stresses. Superoxide dismutase (SOD) proteins, member of the metallo-enzyme family play vital role in protecting plants against abiotic stresses. In the present study, genome-wide analysis of abiotic stress responsive SOD gene family has been done in B. juncea and B. rapa. RESULTS: A total of 29 and 18 SOD genes were identified in B. juncea and B. rapa respectively and chromosome location mapping indicated their wide distribution across genome. On the basis of domain composition, the SODs were phylogenetically classified into sub-groups which was also substantiated by the gene structure and sub-cellular locations of SOD proteins. Functional annotation of SODs was also done by Gene Ontology (GO) mapping and the result was corroborated by the identified cis-regulatory elements in the promoter region of SOD genes. Based on FPKM analysis of SRA data available for drought, heat and salt stress, we identified 14 and 10 abiotic stress responsive SOD genes in B. rapa and B. juncea respectively. The differential expression analysis under drought and heat stress of identified abiotic-stress responsive SOD genes was done through quantitative Real Time PCR. CONCLUSION: We identified abiotic-stress responsive genes that could help in improving the plant tolerance against abiotic stresses. This was the first study to describe the genome-wide analysis of SOD gene family in B. rapa and B. juncea, and the results will help in laying basic ground for future work of cloning and functional validation of SOD genes during abiotic stresses leading to Brassica crop improvement.


Assuntos
Brassica rapa/genética , Regulação Enzimológica da Expressão Gênica , Genoma de Planta , Mostardeira/genética , Proteínas de Plantas/genética , Superóxido Dismutase/genética , Brassica rapa/enzimologia , Brassica rapa/fisiologia , Mapeamento Cromossômico , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Família Multigênica , Mostardeira/enzimologia , Mostardeira/fisiologia , Filogenia , Estresse Fisiológico
12.
Nat Commun ; 10(1): 1154, 2019 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-30858362

RESUMO

Brassica napus (2n = 4x = 38, AACC) is an important allopolyploid crop derived from interspecific crosses between Brassica rapa (2n = 2x = 20, AA) and Brassica oleracea (2n = 2x = 18, CC). However, no truly wild B. napus populations are known; its origin and improvement processes remain unclear. Here, we resequence 588 B. napus accessions. We uncover that the A subgenome may evolve from the ancestor of European turnip and the C subgenome may evolve from the common ancestor of kohlrabi, cauliflower, broccoli, and Chinese kale. Additionally, winter oilseed may be the original form of B. napus. Subgenome-specific selection of defense-response genes has contributed to environmental adaptation after formation of the species, whereas asymmetrical subgenomic selection has led to ecotype change. By integrating genome-wide association studies, selection signals, and transcriptome analyses, we identify genes associated with improved stress tolerance, oil content, seed quality, and ecotype improvement. They are candidates for further functional characterization and genetic improvement of B. napus.


Assuntos
Aclimatação/genética , Brassica napus/genética , Loci Gênicos , Genoma de Planta/genética , Melhoramento Vegetal , Brassica rapa/genética , Cromossomos de Plantas , Ecótipo , Perfilação da Expressão Gênica , Especiação Genética , Sementes/genética , Sequenciamento Completo do Genoma
13.
Food Chem Toxicol ; 126: 313-321, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30831153

RESUMO

Examining tissue-specific expression and the measurement of protein abundance are important steps when assessing the performance of genetically engineered crops. Liquid chromatography-mass spectrometry offers many advantages over traditional methods for protein quantitation, especially when dealing with transmembrane proteins that are often difficult to express or generate antibodies against. In this study, discovery proteomics was used to detect the seven transgenic membrane-bound enzymes from the docosahexaenoic acid (DHA) biosynthetic pathway that had been engineered into canola. Subsequently, a targeted LC-MS/MS method for absolute quantitation was developed and applied to the simultaneous measurement of the seven DHA biosynthetic pathway enzymes in genetically modified canola grown across three sites. The results of this study demonstrated that the enzymatic proteins that drive the production of DHA using seed-specific promoters were detected only in mature and developing seed of DHA canola. None of the DHA biosynthesis pathway proteins were detected in wild-type canola planted in the same site or in the non-seed tissues of the transgenic canola, irrespective of the sampling time or the tissues tested. This study describes a streamlined approach to simultaneously measure multiple membrane-bound proteins in planta.


Assuntos
Brassica rapa/enzimologia , Ácidos Docosa-Hexaenoicos/biossíntese , Espectrometria de Massas/métodos , Proteínas de Membrana/química , Proteínas de Plantas/química , Plantas Geneticamente Modificadas/enzimologia , Brassica rapa/química , Brassica rapa/genética , Brassica rapa/metabolismo , Engenharia Genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/química , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Sementes/química , Sementes/enzimologia , Sementes/genética , Sementes/metabolismo
14.
Cell Mol Biol (Noisy-le-grand) ; 65(2): 37-49, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30860470

RESUMO

This experiment was conducted to assess the quantitative and qualitative changes in soluble proteins as well as some chlorophyll fluorescence parameters in the leaves of a winter canola (Brassica napus L., cv. Licord) under continuous low temperature. Over the experiment, seedlings were initially grown at 15/10 °C (d/n). At fourth fully expanded leafy stage (day 30), a part of the plants were transferred to 4/2°C for 4 weeks. Plants were sampled for protein extraction from leaves in which chlorophyll fluorescence parameters (Fo, Fv, Fm, Fv/Fo, Fm/Fo, Fv/Fm, Fo´, FV´, Fm´ and some other calculated) were also measured. The results showed a clear increase in soluble proteins quantity caused by cold treatment. The enhancements appeared abruptly following the cold exposure to 4°C and lasted. The electrophoretic protein patterns showed changes in the intensity of some polypeptides, besides, induction a new probable protein weighing 47-kW in response to cold treatment. Cold-triggered reduction in maximum quantum yield of PSII (Fv/Fm) was connected especially with drastic decreasing Fv and Fm. Interestingly, high quantitative amounts of soluble proteins along with induction of the new probable polypeptide induced at cold temperature, were attributed to low deduction of maximum quantum yield of PSII. Additionally, more imperative chlorophyll fluorescence parameters changed e.g. qP, NPQ, qL, Y(II) or фPSII etc at light. Nowadays, radar charts or spider plots are the most sophisticated multivariate statistical tools representing physiological responses of plants to abiotic stress conditions or even morphophysiological studies of plants. In rapeseed many researches performed by applying the radar charts for low temperature stresses and interpreted their effects more advancely than common statistical tools. We observed a good representation of the chl fluorescence parameters fluctuations using radar plots. Overall, cold-induced soluble proteins accumulated after longer cold-acclimation, can contribute in photosynthetic apparatus protection against low-temperature damages.


Assuntos
Aclimatação , Brassica rapa/metabolismo , Temperatura Baixa , Complexo de Proteína do Fotossistema II/metabolismo , Proteínas de Plantas/metabolismo , Brassica rapa/genética , Clorofila/metabolismo , Fluorescência , Genótipo , Solubilidade
15.
Int J Mol Sci ; 20(5)2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30832221

RESUMO

Low ambient air temperature limits the growth and selection of crops in cold regions, and cold tolerance is a survival strategy for overwintering plants in cold winters. Studies of differences in transcriptional levels of winter rapeseed (Brassica rapa L.) under cold stress can improve our understanding of transcript-mediated cold stress responses. In this study, two winter rapeseed varieties, Longyou-7 (cold-tolerant) and Lenox (cold-sensitive), were used to reveal morphological, physiological, and transcriptome levels after 24 h of cold stress, and 24 h at room temperature, to identify the mechanism of tolerance to cold stress. Compared to Lenox, Longyou-7 has a shorter growth period and greater belowground mass, and exhibits stronger physiological activity after cold stress. Subsequently, more complete genomic annotation was obtained by sequencing. A total of 10,251 and 10,972 differentially expressed genes (DEG) were identified in Longyou-7 and Lenox, respectively. Six terms closely related to cold stress were found by the Gene Ontology (GO) function annotation. Some of these terms had greater upregulated expression in Longyou-7, and the expression of these genes was verified by qRT-PCR. Most of these DEGs are involved in phenylpropanoid biosynthesis, plant hormone signal transduction, ribosome biogenesis, MAPK signaling pathway, basal transcription factors, and photosynthesis. Analysis of the genes involved in the peroxisome pathway revealed that Longyou-7 and Lenox may have different metabolic patterns. Some transcription factors may play an important role in winter rapeseed tolerance to cold stress, and Longyou-7 is slightly slower than Lenox. Our results provide a transcriptome database and candidate genes for further study of winter rapeseed cold stress.


Assuntos
Brassica rapa/genética , Resposta ao Choque Frio/genética , Genes de Plantas , Transcriptoma
16.
J Sci Food Agric ; 99(8): 4082-4093, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30761554

RESUMO

BACKGROUND: Selenium (Se)-induced phytotoxicity has been linked to oxidative injury triggered by the accumulation of reactive oxygen species (ROS) due to the disturbance of anti-oxidative systems. However, the way Se stress induces hydrogen peroxide (H2 O2 ) production in plants is a long-standing question. Here we identified the role of polyamine oxidase (PAO) in H2 O2 production in the root of Brassica rapa upon Se stress. RESULTS: Studying Se-induced growth inhibition, H2 O2 accumulation, and oxidative injury in the root of Brassica rapa, we found that excessive Se exposure resulted in a remarkable increase in PAO activity. Inhibition of PAO activity led to decreased H2 O2 content and alleviated oxidative injury in the Se-treated root. These results indicated that Se stress induced PAO-dependent H2 O2 production. A total of six BrPAO family members were discovered in the genome of B. rapa by in silico analysis. Se stress pronouncedly upregulated the expression of most BrPAOs and further transient expression analysis proved that it could lead to H2 O2 production. CONCLUSION: These results suggest that Se stress upregulates the expression of a set of BrPAOs which further enhances PAO activity, contributing to H2 O2 generation in roots. © 2019 Society of Chemical Industry.


Assuntos
Brassica rapa/genética , Peróxido de Hidrogênio/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/metabolismo , Proteínas de Plantas/metabolismo , Selênio/metabolismo , Brassica rapa/enzimologia , Brassica rapa/crescimento & desenvolvimento , Brassica rapa/metabolismo , Regulação da Expressão Gênica de Plantas , Estresse Oxidativo , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/genética , Proteínas de Plantas/genética , Espécies Reativas de Oxigênio/metabolismo
17.
Plant Mol Biol ; 99(6): 603-620, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30783953

RESUMO

KEY MESSAGE: Thirty-five IQD genes were identified and analysed in Chinese cabbage and BrIQD5 transgenic plants enhanced the drought resistance of plants. The IQD (IQ67-domain) family plays an important role in various abiotic stress responses in plant species. However, the roles of IQD genes in the Chinese cabbage response to abiotic stress remain unclear. Here, 35 IQD genes, from BrIQD1 to BrIQD35, were identified in Chinese cabbage (Brassica rapa ssp. pekinensis). Based on the phylogenetic analysis, these genes were clustered into three subfamilies (I-III), and members within the same subfamilies shared conserved exon-intron distribution and motif composition. The 35 BrIQD genes were unevenly distributed on 9 of the 10 chromosomes with 4 segmental duplication events. Ka/Ks ratios showed that the duplicated BrIQDs had mainly experienced strong purifying selection. Quantitative real-time polymerase chain reaction of 35 BrIQDs under PEG6000 indicated that BrIQD5 was significantly induced by PEG6000. To verify BrIQD5 function, BrIQD5 was heterologously overexpressed in tobacco and was silenced in Chinese cabbage. BrIQD5-overexpressed plants showed more tolerance to drought stress than wild-type plants, while BrIQD5-silenced plants in Chinese cabbage showed decreased drought tolerance. Additionally, six BrIQD5 potential interactive proteins were isolated by the yeast two-hybrid assay, including BrCaMa, BrCaMb and four other stress-related proteins. Motif IQ1 of BrIQD5 is important for the interaction with BrCaMa and BrCaMb, and the isoleucine in motif IQ1 is an essential amino acid for calmodulin binding to BrIQD5. The identification and cloning of the new Chinese cabbage drought tolerance genes will promote the drought-resistant breeding of Chinese cabbage and help to better understand the mechanism of IQD involved in the drought tolerance of plants.


Assuntos
Proteínas de Arabidopsis/classificação , Proteínas de Arabidopsis/genética , Brassica rapa/genética , Proteínas de Ligação a Calmodulina/classificação , Proteínas de Ligação a Calmodulina/genética , Secas , Regulação da Expressão Gênica de Plantas , Família Multigênica/genética , Filogenia , Mapeamento Cromossômico , Cromossomos de Plantas/genética , Perfilação da Expressão Gênica , Genoma de Planta , Duplicações Segmentares Genômicas , Alinhamento de Sequência , Estresse Fisiológico/genética , Tabaco/genética , Transcriptoma/genética
18.
Gene ; 696: 113-121, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30776462

RESUMO

Multiple-allele-inherited male sterility (MAMS) is important in Chinese cabbage (Brassica rapa L. ssp. pekinensis) breeding, but the molecular mechanisms leading to male sterility are poorly understood. In this study, we cloned a novel gene, BrSKS13, that is differentially expressed in fertile and sterile flower buds of Chinese cabbage. BrSKS13 is most similar to Arabidopsis thaliana AT3G13400 (SKS13) and encodes a predicted 61.87 kDa protein with three cupredoxin superfamily conserved domains in the multicopper oxidase family. Semi-quantitative reverse-transcription PCR (sqRT-PCR) showed that expression of BrSKS13 is higher in fertile buds than in sterile buds. Quantitative RT-PCR (qRT-PCR) and in situ hybridization showed that BrSKS13 is highly expressed in fertile anthers, peaking at pollen-maturation stage VI, but is weakly expressed in other tissues and floral organs. Expression patterns of BrSKS13 promoter::GUS reporter fusions in Arabidopsis showed that the BrSKS13 promoter drives expression of the GUS gene only in anthers. The relative expression of Brsks13 in fertile buds was higher than in sterile buds for all other MAMS lines of Chinese cabbage examined. These results suggest that BrSKS13 affects pollen development. In situ hybridization analysis of flower stigmas at different times after pollination showed that BrSKS13 expression was first observed in stigmas and immature seeds at 1 h after pollination, and the signal intensity in seeds increased with increasing maturity. Compared to Col-0, A. thaliana sks13 mutant plants have shorter and fewer siliques, shriveled pollen grains, pollen tube abnormalities, and reduced seed number. The phenotype of sks13 mutant was recovered by over-expressing BrSKS13. Our results suggest that BrSKS13 affects pollen development and the pollination/fertilization process, and will enable further study of the genetic mechanisms underlying MAMS in Chinese cabbage.


Assuntos
Brassica rapa/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica de Plantas/genética , Infertilidade das Plantas/genética , Proteínas de Plantas/genética , Polinização/genética , Alelos , Arabidopsis/genética , Brassica rapa/crescimento & desenvolvimento , Fertilização/genética , Perfilação da Expressão Gênica , Herança Multifatorial/genética , Plantas Geneticamente Modificadas , Pólen/crescimento & desenvolvimento
19.
BMC Plant Biol ; 19(1): 13, 2019 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-30621588

RESUMO

BACKGROUND: Plasmodiophora brassicae is a soil-borne plant pathogen that causes clubroot disease, which results in crop yield loss in cultivated Brassica species. Here, we investigated whether a quantitative trait locus (QTL) in B. rapa might confer resistance to a Korean P. brassicae pathotype isolate, Seosan. We crossed resistant and susceptible parental lines and analyzed the segregation pattern in a F2 population of 348 lines. We identified and mapped a novel clubroot resistance QTL using the same mapping population that included susceptible Chinese cabbage and resistant turnip lines. Forty-five resistant and 45 susceptible F2 lines along with their parental lines were used for double digest restriction site-associated DNA sequencing (ddRAD-seq). High resolution melting (HRM)-based validation of SNP positions was conducted to confirm the novel locus. RESULTS: A 3:1 ratio was observed for resistant: susceptible genotypes, which is in accordance with Mendelian segregation. ddRAD-seq identified a new locus, CRs, on chromosome A08 that was different from the clubroot resistance (CR) locus, Crr1. HRM analysis validated SNP positions and constricted CRs region. Four out of seventeen single nucleotide polymorphisms (SNPs) positions were within a 0.8-Mb region that included three NBS-LRR candidate genes but not Crr1. CONCLUSION: The newly identified CRs locus is a novel clubroot resistance locus, as the cultivar Akimeki bears the previously known Crr1 locus but remains susceptible to the Seosan isolate. These results could be exploited to develop molecular markers to detect Seosan-resistant genotypes and develop resistant Chinese cabbage cultivars.


Assuntos
Brassica rapa/genética , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , Brassica rapa/parasitologia , Plasmodioforídeos/patogenicidade
20.
Mol Biol Rep ; 46(1): 1227-1238, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30637624

RESUMO

Seed size and seed metabolites have been the targets of direct or indirect selection during domestication and subsequent crop breeding. Understanding these traits and associated genetics can prove very useful for plant translational research. Large germplasm assemblage (235) of Brassica juncea and its progenitor species (B. rapa and B. nigra) was evaluated to establish seed trait variations for seed size and seed metabolites. Seeds were smallest in B. nigra and largest in B. juncea. Australian B. juncea and Indian B. rapa var brown sarson types averaged more seed oil content. Seed size and oil content were generally higher in modern cultivars in comparison to the land races. Allelic diversity for known associated genes for seed-size and oil-content (AP2, ARF2, TTG2, GRF2, GL2, CYP78A5, CYP78A6, MINI3, IKU2, IKU1, BRI1, DGAT, GPDH, LPAAT, GPAT and DA1) was studied so as to infer the effect of domestication on seed traits. Three genes (IKU1, IKU2, AP2) in B. rapa, two (TTG2 and GL2) in B. nigra and two (IKU1 and GRF2) in natural B. juncea were identified as targets of selection on the basis of Fst outlier and/or sequence diversity tests. We report parallel divergence for seed traits between B. juncea and B. rapa. Directional selection appeared stronger for seed-size as compared to correlated seed metabolites. Positive selection on seed-size is likely to have played a significant role in structuring regional variation in the germplasm.


Assuntos
Mostardeira/genética , Sementes/genética , Alelos , Evolução Biológica , Brassica rapa/genética , Mapeamento Cromossômico/métodos , Diploide , Evolução Molecular , Ácidos Graxos/genética , Frequência do Gene/genética , Variação Genética/genética , Genoma de Planta/genética , Genótipo , Fenótipo , Seleção Genética/genética
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