| 周 荣 1，刘嘉超 1,2，杨凤玺 2.墨兰 CsAP1-A 基因克隆及表达分析[J].广东农业科学,2023,50(9):99-107 |
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墨兰 CsAP1-A 基因克隆及表达分析 |
| Analysis on Cloning and Expression of CsAP1-A Gene in Cymbidium sinense |
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| DOI：10.16768/j.issn.1004-874X.2023.09.010 |
| 中文关键词: 墨兰 CsAP1-A 基因 生物信息学 表达分析 花器官发育 |
| 英文关键词: Cymbidium sinense CsAP1-A gene bioinformatics expression analysis flower development |
| 基金项目:广州市科技计划项目（2023B03J1322）；国家重点研发计划项目（2019YFD1001003） |
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| 摘要点击次数: 283 |
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| 中文摘要: |
| 【目的】AP1 基因在植物的花器官发育和开花调控中发挥重要作用，对墨兰 AP1 基因进行克隆与表达分析可为研究其在墨兰花发育和开花调控中的作用提供前期基础。【方法】以墨兰品种‘小香’为材料，克隆到 1 个 AP1 基因，命名为 CsAP1-A。通过生物信息学分析其基因结构、蛋白结构域和进化关系，利用 RT-qPCR 方法分别检测 CsAP1-A 在墨兰不同器官、不同花发育阶段和不同花组织部位的表达情况；通过转录组测序分析 CsAP1-A 在 5 个不同花型墨兰品种花组织部位的表达情况；通过蛋白互作预测软件分析 CsAP1-A 与其他蛋白的互作关系。【结果】CsAP1-A 基因编码区为 744 bp，编码 248 个氨基酸，含有高度保守的 MADS-box 和K-box 结构域，符合 MADS-box 转录因子家族特征。CsAP1-A 与其他兰科植物 AP1 蛋白相似性较高，其中与春兰 AP1/FUL3（APY18447.1）和蕙兰 MADS1（AGE15496.1）的同源性最高。RT-qPCR 分析结果表明，CsAP1-A在墨兰不同器官中均有表达，在花中表达量最高，且集中在花芽分化初期（S1）高表达。在不同花型的墨兰品种中，CsAP1-A 在 WT（普通型）、MPV（重瓣花型）、LaPV（花瓣唇瓣化花型）和 NLV（唇瓣萼片化花型）4 种花型的合蕊柱中表达量均最高，而在萼片中表达量最低；在合蕊柱异常发育的 MPV 中，CsAP1-A 在合蕊柱的表达量显著提高，而在花瓣蕊柱化的梅瓣花型（GPV）中整体表达量最高，且表达区域从合蕊柱扩展到花瓣。蛋白互作预测 CsAP1-A 蛋白可与 MADS1、MADS6、MADS47、MADS8 等 10 个蛋白存在互作关系。【结论】墨兰 CsAP1-A 的基因结构、进化关系、时空表达情况和蛋白互作预测可为墨兰花发育的研究提供理论依据，对进一步揭示 CsAP1-A 基因在墨兰成花过程中的作用奠定基础。 |
| 英文摘要: |
| 【Objective】AP1 gene plays an important role in the development of floral organs and regulation of flowering in plants. The analysis on cloning and expression of the AP1 gene of Cymbidium sinense can provide a preliminary foundation for the study of its role in the development of C. sinense flowers and the regulation of flowering.【Method】An AP1 gene, named CsAP1-A, was cloned from the C. sinense variety ‘Xiao Xiang’, and its gene structure, protein structural domains and evolutionary relationship were analyzed by bioinformatics. RT-qPCR was used to detect the expression of CsAP1-A in different organs, flower development stages and flower tissue parts, respectively, and the expression of CsAP1-A in flower tissues of five different flower types of C. sinense was analyzed by RNA-seq. The interactions between CsAP1-A and other proteins were analyzed by the Protein Interaction Prediction Software.【Result】The coding region of CsAP1-A gene was 744 bp, encoding 248 amino acids, with highly conserved MADS-box and K-box structural domains, which is in line with the characteristics of the MADS-box family of transcription factors. The protein sequence comparison revealed that the similarity between CsAP1-A and the AP1 proteins of other orchid plants was relatively high, among which the highest homology was found with the AP1/FUL3（APY18447.1） of C. goeringii and MADS1 (AGE15496.1) of C. faberi. RT-qPCR showed CsAP1-A was expressed in different organs of C. sinense, the highest expression was in flowers, and concentrated in stage of early flower development (S1). In the five varieties with different floral patterns, the expression of CsAP1-A in common type variety (WT), multi-perianth variety (MPV), labellum-like petal variety (LaPV) and null-labellum variety (NLV) were the highest in gynostegiums and the lowest in sepals. CsAP1-A was significantly up-regulated expression in MPV with abnormal development of gynostegium. The overall expression was the highest in gynostegium-like petal variety (GPV), and the expression area expanded from gynostegium to petal. Protein interactions predicted that CsAP1-A could interact with 10 proteins, including MADS1, MADS6, MADS47 and MADS8 etc. 【Conclusion】The gene structure, evolutionary relationship, spatio-temporal expression and protein interaction prediction of CsAP1-A can provide theoretical basis for the study of flower development in C. sinense, and lay a foundation for further revealing the role of CsAP1-A gene in flower formation of C. sinense. |
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