| Tomato (Solanum lycopersicum) is typical climacteric fruit, which is not only amodel plant to study the mechanism of fruit development and maturation, but also anideal material to study the stress responses. Abscisic acid (ABA) is an importantphytohormone involving in the various stages of plant growth and development, such asseed dormancy, fruit ripening, leaf senescence, stomatal closure. On the other hand,ABA also regulates stress responses (drought, salt, disease, high or low temperature)and is called as stress hormones.This experiment we separated nine ABA receptor (SlPYL), six negative regulator ofABA signal transduction (SlPP2C) and seven positive regulator (SlSnRK2) in tomato,screening out some genes that are sensitive to ABA and osmosis stress. We clonedSlSnRK2.1, SlSnRK2.2and SlSnRK2.3which belonged SlSnRK2Ⅲ subclasses, studiedits expression mode in tomato tissue and subcellular localization. Overexpression ofSlSnRK2.1, SlSnRK2.2, SlSnRK2.3and get transgenic plants of SlSnRK2.1andSlSnRK2.2. Treating plants with drought and salt to analyze the stress response ofSlSnRK2s. RNAi interference vector of SlSnRK2.1/2.2/2.3were constructed and theresults suggested that maybe SlSnRK2.1, SlSnRK2.2and SlSnRK2.3related to the signalintersection of ABA and ethylene. The main results are as follows:①Nine SlPYLs genes, six SlPP2Cs genes, seven SlSnRK2s genes in tomato wereidentified, the deduced amino acid sequence lengths of SlPYLs were123-218, theSlPP2Cs were397-544and the SlSnRK2s were140-362. The SlPYLs and SlSnRK2swere clustered into three subfamilies/subclasses. Within the SlPYL family, SlPYL4,SlPYL7, SlPYL8and SlPYL9belonged to subfamilyⅠ, SlPYL5, SlPYL6and SlPYL10belonged to subfamilyⅡ,the others belonged to subfamilyⅢ. SlSnRK2subclassⅠincluded SlSnRK2.5, SlSnRK2.7, SlSnRK2subclassⅡincluded SlSnRK2.4, SlSnRK2subclassⅢincluded SlSnRK2.1, SlSnRK2.2, SlSnRK2.3and SlSnRK2.6.②Within SlPYL gene family, the general trend was down-regulated by ABA,specially the SlPYL10. SlABI2-1, SlABI2-2and SlPP2CA2was induced by ABA, whileSlPP2CA1was suppressed slowly. Within SlSnRK2family, in addition to SlSnRK2.1and SlSnRK2.7, the rest of the genes had a response to ABA, suggested that theyinvolved in ABA signal transduction.③All the SlPYL genes except for SlPYL1and SlPYL7had responses to salt stress. Almost all the SlPP2C genes were inhibited by salt stress, specially the SlHAB andSlPP2CA1. While SlPP2CA2had a small amplitude increase expression at72hour. Allthe SlSnRK2s genes had significant changes under salt stress.④Within SlPYL gene family, all the genes in addition to SlPYL2and SlPYL10hadsignificant changes under drought stress. All the SlPP2C genes were up-regulated underdrought stress, particularly the SlABA2-2. Within SlSnRK2family, the expression levelsof SlSnRK2.1SlSnRK2.3, SlSnRK2.5and SlSnRK2.6were increased, involved in theresponse to drought stress.⑤The full-length cDNA of SlSnRK2.1, SlSnRK2.2and SlSnRK2.3were isolated.Construction features were analyzed through bioinformatics: SlSnRK2.1contained acomplete ORF encoding362amino acids, SlSnRK2.2encoding339amino acids,SlSnRK2.3encoding352amino acids; contained phosphorylation sites belonging to theSlSnRK2III subclass; predicted promoter included ABA response element andstress-related cis-acting elements.⑥The expression pattern showed that SlSnRK2.1, SlSnRK2.2and SlSnRK2.3hadexpressed in all organizations involved in entire process of plant growth anddevelopment.The subcellular localizations of the SlSnRK2.1/2.2/2.3proteins wereperformed by fusion to the green fluorescent protein (GFP).Results showed thatSlSnRK2.1, SlSnRK2.2and SlSnRK2.3existed in the cytoplasm and the nucleus,interacted with other transcription factor in the cell.⑦Overexpression of SlSnRK2.1, SlSnRK2.2, SlSnRK2.3and get transgenic plantsof SlSnRK2.1and SlSnRK2.2. The transgenic plants had reduced seed germination andincreased sensitivity to osmotic stress decreasing stress tolerance.⑧RNAi interference vector of SlSnRK2.1/2.2/2.3were constructed and somephenotype in transgenic plants were observed like premature aging (leaf senescence,flower withering) and branch increased response. Quantitative PCR analyses ofethylene-synthesis-related genes expression were changed in transgenic plants.,speculated that maybe SlSnRK2.1, SlSnRK2.2and SlSnRK2.3related to the signalintersection of ABA and ethylene. |
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