Xinjiang Key Laboratory of Crop Biotechnology, The State Key Laboratory of Genetic Improvement and Germplasm Innovation of Crop Resistance in Arid Desert Regions (Preparation), Institute of Nuclear and Biological Technology, Xinjiang Academy of Agricultural Sciences, Urumqi 830091, China.
College of Agronomy, Xinjiang Agricultural University, Urumqi 830052, China.
Int J Mol Sci. 2024 Feb 5;25(3):1917. doi: 10.3390/ijms25031917.
Multiple cis-acting elements are present in promoter sequences that play critical regulatory roles in gene transcription and expression. In this study, we isolated the cotton (Fiddlehead) gene promoter (pGhFDH) using a real-time reverse transcription-PCR (qRT-PCR) expression analysis and performed a cis-acting elements prediction analysis. The plant expression vector pGhFDH::GUS was constructed using the Gateway approach and was used for the genetic transformation of and upland cotton plants to obtain transgenic lines. Histochemical staining and a β-glucuronidase (GUS) activity assay showed that the GUS protein was detected in the roots, stems, leaves, inflorescences, and pods of transgenic lines. Notably, high GUS activity was observed in different tissues. In the transgenic lines, high GUS activity was detected in different tissues such as leaves, stalks, buds, petals, androecium, endosperm, and fibers, where the pGhFDH-driven GUS expression levels were 3-10-fold higher compared to those under the CaMV 35S promoter at 10-30 days post-anthesis (DPA) during fiber development. The results indicate that pGhFDH can be used as an endogenous constitutive promoter to drive the expression of target genes in various cotton tissues to facilitate functional genomic studies and accelerate cotton molecular breeding.
多个顺式作用元件存在于启动子序列中，这些元件在基因转录和表达中起着关键的调控作用。在这项研究中，我们使用实时逆转录-聚合酶链反应（qRT-PCR）表达分析分离了棉花（Fiddlehead）基因启动子（pGhFDH），并进行了顺式作用元件预测分析。使用 Gateway 方法构建了植物表达载体 pGhFDH::GUS，并用于 和陆地棉植物的遗传转化，以获得转基因系。组织化学染色和β-葡萄糖醛酸酶（GUS）活性测定表明，在转基因 系的根、茎、叶、花序和荚果中检测到 GUS 蛋白。值得注意的是，不同组织中检测到高 GUS 活性。在转基因系中，不同组织如叶片、茎秆、芽、花瓣、雄蕊、胚乳和纤维中检测到高 GUS 活性，在纤维发育过程中，与 CaMV 35S 启动子相比，pGhFDH 驱动的 GUS 表达水平在 10-30 天开花后天（DPA）时高出 3-10 倍。结果表明，pGhFDH 可作为内源性组成型启动子，在各种棉花组织中驱动靶基因的表达，从而促进功能基因组学研究和加速棉花分子育种。
