RESUMO
BACKGROUND: 'Regal Splendour' (Hosta variety) is famous for its multi-color leaves, which are useful resources for exploring chloroplast development and color changes. The expressions of chlorophyll biosynthesis-related genes (HrHEMA, HrPOR and HrCAO) in Hosta have been demonstrated to be associated with leaf color. Herein, we isolated, sequenced, and analyzed HrHEMA, HrPOR and HrCAO genes. Subcellular localization was also performed to determine the location of the corresponding enzymes. After plasmid construction, virus-induced gene silencing (VIGS) was carried out to reduce the expressions of those genes. In addition, HrHEMA-, HrPOR- and HrCAO-overexpressing tobacco plants were made to verify the genes function. Changes of transgenic tobacco were recorded under 2000 lx, 6000 lx and 10,000 lx light intensity. Additionally, the contents of enzyme 5-aminolevulinic acid (5-ALA), porphobilinogen (PBG), chlorophyll a and b (Chla and Chlb), carotenoid (Cxc), superoxide dismutase (SOD), peroxidase (POD), malondialdehyde (MDA), proline (Pro) and catalase (CAT) under different light intensities were evaluated. RESULTS: The silencing of HrHEMA, HrPOR and HrCAO genes can induce leaf yellowing and chloroplast structure changes in Hosta. Specifically, leaves of Hosta with HrCAO silencing were the most affected, while those with HrPOR silencing were the least affected. Moreover, all three genes in tobacco were highly expressed, whereas no expression was detected in wild-type (WT). However, the sensitivities of the three genes to different light intensities were different. The highest expression level of HrHEMA and HrPOR was detected under 10,000 lx of illumination, while HrCAO showed the highest expression level under 6000 lx. Lastly, the 5-ALA, Chla, Cxc, SOD, POD, MDA, Pro and CAT contents in different transgenic tobaccos changed significantly under different light intensities. CONCLUSION: The overexpression of these three genes in tobacco enhanced photosynthesis by accumulating chlorophyll content, but the influential level varied under different light intensities. Furthermore, HrHEMA-, HrPOR- and HrCAO- overexpressing in tobacco can enhance the antioxidant capacity of plants to cope with stress under higher light intensity. However, under lower light intensity, the antioxidant capacity was declined in HrHEMA-, HrPOR- and HrCAO- overexpressing tobaccos.
Assuntos
Clorofila/biossíntese , Hosta/fisiologia , Nicotiana/fisiologia , Folhas de Planta/fisiologia , Proteínas de Plantas/genética , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Clorofila/genética , Clorofila/metabolismo , Regulação da Expressão Gênica de Plantas , Hosta/genética , Luz , Malondialdeído/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Oxigenases/genética , Oxigenases/metabolismo , Filogenia , Pigmentação/genética , Folhas de Planta/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Plântula/genética , Plântula/metabolismo , Estresse Fisiológico/genética , Estresse Fisiológico/fisiologia , Nicotiana/genéticaRESUMO
BACKGROUND: Hosta plantaginea (Lam.) Aschers (HPA), a species in the family Liliaceae, is an important landscaping plant and herbaceous ornamental flower. However, because the flower has only two colors, white and purple, color matching applications are extremely limited. To date, the mechanism underlying flower color regulation remains unclear. METHODS: In this study, the transcriptomes of three cultivars-H. plantaginea (HP, white flower), H. Cathayana (HC, purple flower), and H. plantaginea 'Summer Fragrance' (HS, purple flower)-at three flowering stages (bud stage, initial stage, and late flowering stage) were sequenced with the Illumina HiSeq 2000 (San Diego, CA, USA). The RNA-Seq results were validated by qRT-PCR of eight differentially expressed genes (DEGs). Then, we further analyzed the relationship between anthocyanidin synthase (ANS), chalcone synthase (CHS), and P450 and the flower color regulation by Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Eukaryotic Orthologous Groups (KOG) network and pathway enrichment analyses. The overexpression of CHS and ANS in transgenic tobacco petals was verified using qRT-PCR, and the petal colors associated with the overexpression lines were confirmed using absorbance values. RESULTS: Over 434,349 transcripts were isolated, and 302,832 unigenes were identified. Additionally, through transcriptome comparisons, 2098, 722, and 606 DEGs between the different stages were found for HP, HC, and HS, respectively. Furthermore, GO and KEGG pathway analyses showed that 84 color-related DEGs were enriched in 22 pathways. In particular, the flavonoid biosynthetic pathway, regulated by CHS, ANS, and the cytochrome P450-type monooxygenase gene, was upregulated in both purple flower varieties in the late flowering stage. In contrast, this gene was hardly expressed in the white flower variety, which was verified in the CHS and ANS overexpression transgenic tobacco petals. CONCLUSIONS: The results suggest that CHS, ANS, and the cytochrome P450s-regulated flavonoid biosynthetic pathway might play key roles in the regulation of flower color in HPA. These insights into the mechanism of flower color regulation could be used to guide artificial breeding of polychrome varieties of ornamental flowers.
Assuntos
Aciltransferases/genética , Perfilação da Expressão Gênica/métodos , Hosta/fisiologia , Nicotiana/genética , Oxigenases/genética , Vias Biossintéticas , Cor , Flores/genética , Flores/fisiologia , Regulação da Expressão Gênica de Plantas , Sequenciamento de Nucleotídeos em Larga Escala , Hosta/genética , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Locos de Características Quantitativas , Análise de Sequência de RNA , Nicotiana/crescimento & desenvolvimentoRESUMO
Chloroplast (cp) genome sequences have become a useful tool for phylogeneitc and evolutionary study in recent reports. Here the complete chloroplast genome of the Paphiopedilum tranlimianum has been reconstructed from the whole-genome Illumina sequencing data. The circular genome is 162,127 bp in size, and comprises of a pair of inverted repeat (IR) regions of 31,457 bp each, a large single-copy (LSC) region of 91,711 bp, and a small single-copy (SSC) region of 7,502 bp. The total GC content is 35.4%, while the corresponding values of the LSC, SSC, and IR regions are 32.7%, 26.9%, and 40.9%, respectively. The chloroplast genome contains 133 genes, including 87 protein-coding genes, eight ribosomal RNA genes, and 38 transfer RNA genes. Three genes (ndhA, ndhE, and ndhI) are pseudogenized or lost in its cpDNA. The Maximum-Likelihood phylogenetic analysis showed a close relationship with P. niveum in Orchidaceae. Our findings provide useful information for phylogenetic and evolutionary research of Paphiopedilum species.
RESUMO
Chloroplast (cp) genome sequences provide a valuable source for phylogenetic analysis, which becomes a popular tool for population and phylogeny in a recent report. Here, the complete chloroplast genome of the Paphiopedilum wardii has been reconstructed from the whole-genome Illumina sequencing data. The circular genome is 169,820 bp in size and comprises a pair of inverted repeat (IR) regions of 33,641 bp each, a large single-copy (LSC) region of 98,872 bp and a small single-copy (SSC) region of 3666 bp. The total GC content is 33.8%, while the corresponding values of the IR regions, LSC, and SSC and are 38.7%, 31.7% and 26.7%, respectively. The chloroplast genome contains 151 genes, including 91 protein-coding genes, eight ribosomal RNA genes and 52 transfer RNA genes. Two genes (ndhA and ndhB) are pseudogenized or lost in its cpDNA. The maximum-likelihood phylogenetic analysis showed a close relationship with P. armeniacum in Orchidaceae. Our findings provide useful information for phylogenetic and evolutionary research of Paphiopedilum species.
