RESUMO
In order to ascertain the regulatory mechanism of fruit development in Isatis indigotica Fortune, the complementary DNA (cDNA) sequence of the SHATTERPROOF 2 (SHP2) orthologous gene was identified by Rapid Amplification of cDNA Ends technology and the corresponding gene was named IiSHP2. The expression pattern of IiSHP2 was determined by quantitative reverse transcription-polymerase chain reaction and wild-type Col-0 Arabidopsis plants were transformed with the IiSHP2 gene using Agrobacterium tumefaciens and the floral-dip method. Expression analyses indicated that IiSHP2 was highly expressed in flowers, silicles and seeds. Compared to wild-type plants, IiSHP2 transgenic lines bolted earlier. Detailed phenotypic observations showed that the size of the rosette and cauline leaves in transgenic lines was reduced and the cauline leaves of the transgenic lines were incurved and displayed a funnel-like shape. During the reproductive growth stage, IiSHP2 transgenic plants produced shortened sepals and the flower buds were not encapsulated completely. Moreover, the petals of the transgenic lines were converted into stamineous tissues, accompanied by exposed stamens, short malformed siliques and wrinkled valves, indicating a severe decline in fertility. These experimental conclusions are valuable as a reference for the breeding of medicinal plants.
RESUMO
The coding sequence of IiFUL in Isatis indigotica was isolated and was used in transformation of Arabidopsis. IiFUL overexpressing Arabidopsis plants exhibited early flowering phenotype, accompanied with the reduction of flower number and the production of terminal flower on the top of the main stems. In development process, the flowers located on the top of the main stems generated a lot of variations in phenotype, including abnormal swelling of pistil, withering and numerical change of stamens and petals, appearance of stigmatoid tissues and naked ovules at the margin or inside of sepals. Besides, secondary flower could be formed within the flowers on the top of the main stems. These observations illustrated that IiFUL mainly affected the development of inflorescence meristems and pistils, but its ectopic expression could also disturb the normal growth of other floral organs. Moreover, the fertile siliques produced by the lateral inflorescences of IiFUL overexpressing Arabidopsis plants showed indehiscent phenotype, and the shape of the cauline leaves was changed significantly. The results of quantitative real-time PCR revealed that higher transcriptional levels of IiFUL could be detected in flowers and silicles of I. indigotica. In comprehensive consideration of the previous reports about the dehiscence phenotype of Arabidopsis siliques and the fact that the siliques of IiFUL overexpressing Arabidopsis plants were indehiscent in the present work, it can be speculated that high expression of IiFUL in pericarp is likely the reason why the silicles of I. indigotica possess an indehiscent phenotype.
Assuntos
Arabidopsis , Flores , Regulação da Expressão Gênica de Plantas , Isatis/genética , Proteínas de Domínio MADS , Proteínas de Plantas , Plantas Geneticamente Modificadas , Arabidopsis/genética , Arabidopsis/metabolismo , Flores/genética , Flores/metabolismo , Proteínas de Domínio MADS/biossíntese , Proteínas de Domínio MADS/genética , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Reprodução/genéticaRESUMO
The function of AZI1 in systemic acquired resistance of Arabidopsis was confirmed by investigation of the phenotypic features of wild-type Col-0, AZI1 T-DNA knockout and AZI1 overexpressing plants after infection with virulent and avirulent Pseudomonas syringae. Real-time quantitative PCR and Northern blotting analyses showed that the transcript abundances of PR genes increased significantly in local and systemic leaves of wild-type Col-0 and AZI1 overexpressing plants challenged with avirulent P. syringae, whereas the mRNA accumulation of PR genes was obviously attenuated in local and systemic leaves of AZI1 T-DNA knockout plants after localized infiltration with avirulent Psm avrRpm1. The changes of metabolomic profiles in distal leaves of three types of materials infected with avirulent P. syringae were determined by (1)H NMR spectrometry and data mining showed that the soluble carbonhydrates might function as signal substances in the systemic immunity of Arabidopsis. At the same time, the expression of the sugar signaling genes in local and distal leaves after infection of avirulent P. syringae was compared. As a result, it was found that the transcript abundances of sugar signaling genes, including SUS1, SUS2, SUS3, SUS6, SUT1, HXK1, HXK2, SNRK1.2, ERD6, TPS1, TOR, SNRK1.1, SNRK1.3 and bZIP11, were obviously changed in distal leaves of different materials with the modulated AZI1 activities, indicating sugar-related genes are involved in regulation of the systemic immunity mediated by AZI1. These results also illustrated that the immune system associated with sugar molecules probably was an important part of the systemic acquired resistance in Arabidopsis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/imunologia , Arabidopsis/metabolismo , Metabolismo dos Carboidratos , Imunidade Inata , Metabolômica/métodos , Transdução de Sinais , Arabidopsis/genética , Arabidopsis/microbiologia , Proteínas de Arabidopsis/genética , Proteínas de Bactérias/metabolismo , Metabolismo dos Carboidratos/genética , Análise Discriminante , Regulação da Expressão Gênica de Plantas , Técnicas de Inativação de Genes , Análise dos Mínimos Quadrados , Sulfato de Magnésio/farmacologia , Fenótipo , Doenças das Plantas/imunologia , Doenças das Plantas/microbiologia , Folhas de Planta/genética , Folhas de Planta/microbiologia , Espectroscopia de Prótons por Ressonância Magnética , Pseudomonas syringae/fisiologia , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/genética , Transcrição GênicaRESUMO
Fifteen SPL (SQUAMOSA PROMOTER BINDING PROTEIN-LIKE) genes were identified and characterized in Nicotiana tabacum L. cv. Qinyan95. The exon-intron structures of these genes were determined according to the coding sequences confirmed by RT-PCR and the genomic DNA sequences downloaded from the databases in Sol Genomics Network, and thirteen of them were found to carry the response element of miR156. To elucidate the origin of the validated NtabSPL genes, multiple alignments of the nucleotide sequences encompassing the open reading frames were conducted by using the orthologs in N. tabacum, Nicotiana sylvestris, Nicotiana tomentosiformis, and Nicotiana otophora. The results showed that six NtabSPL genes were derived from a progenitor of N. sylvestris, and nine NtabSPL genes were derived from a progenitor of N. tomentosiformis, further corroborating that N. tabacum came from the interspecific hybridization between the ancestors of N. sylvestris and N. tomentosiformis. In contrast to previous statements about highly repetitive sequences, the genome of N. tabacum mainly retained the paternal-derived SPL genes in diploidization process. Phylogenetic analyses based on the highly conserved SBP (SQUAMOSA PROMOTER BINDING PROTEIN) domains and the full-length amino acid sequences reveal that the SPL proteins of tobacco, tomato, and Arabidopsis can be categorized into eight groups. It is worth noting that N. tabacum contains seven NtabSPL6 genes originated from two parental genomes and NtabSPL6-2 possesses a GC-AG intron. In addition, transgenic tobacco plants harboring Arabidopsis Pri-miR156A were generated by Agrobacterium-mediated transformation method, and the constitutive expression of miR156 could obviously inhibit the activity of the NtabSPL genes containing its target site, suggesting the function of miR156 is conservative in tobacco and Arabidopsis.
