Cloning and expression analysis of tonoplast dicarboxylate transporter gene ScTDT in sugarcane

【Objective】To clone the sugarcane tonoplast dicarboxylate transporter gene(ScTDT) and analyze its expression pattern in different tissues of sugarcane and under aluminum stress, so as to provide a theoretical basis for indepth study of the gene function and the molecular mechanism of sugarcane resistance to aluminum stress.【Method】Homologous cloning technology was used to clone ScTDT from sugarcane variety ROC22.Bioinformatics software was used to analyze the gene sequence.Real-time fluorescence quantitative PCR was used to detect the gene expression levels in different sugarcane tissues(root, stem and leaf) and under aluminum stress(0, 10 and 20μmol/L Al³⁺).【Result】The cloned ScTDT gene had an open reading frame(ORF) of 1623 bp, encoding 540 amino acid residues with a relative molecular weight of 57.34 kD and a theoretical isoelectric point(pI) of 5.77.ScTDT protein was an unstable hydrophobin and may be located in plasma membrane, vacuole and/or Golgi.The amino acid sequence of ScTDT was highly similar to the amino acid sequence of TDT in sorghum(XP_002460443.1), rice(XP_015612609.1) and maize(PWZ04635.1).Among them, the TDT amino acid sequence of sorghum had the highest similarity, reaching 96.30%, indicating that ScTDT had the closest genetic relationship with TDT of sorghum.The ScTDT gene was expressed in sugarcane roots, stems and leaves, and the expression level in roots was significantly higher than that in stems and leaves(P< 0.05, the same below).Under aluminum stress, the expression of ScTDT gene in the roots of the three sugarcane varieties(ROC222, Liucheng 05-136 and Zhongtang 1202) was significantly higher than that of the control group(0μmol/L Al³⁺), especially in Liucheng 05-136 and Zhongtang 1202.With the increase of Al³⁺concentration in the nutrient solution, the expression of ScTDT gene in these two varieties showed a significant increase trend, indicating that high-concentration Al³⁺stress could induce the high-efficiency expression of ScTDT gene.Among the three sugarcane varieties, Zhongtang 1202 had the largest change in ScTDT gene expression, followed by Liucheng 05-136, and ROC22 had the smallest change.【Conclusion】The cloned ScTDT gene is tissue-specific, and its high expression in roots may be related to the resistance of sugarcane to aluminum stress.That is, plants can up-regulate the expression of ScTDT gene in roots to accelerate the release of malate from vacuoles, and then promote the secretion of malate from the roots into the soil to react with Al³⁺, thereby reducing the toxicity of aluminum, which indicates that the ScTDT gene may be involved in sugarcane resistance to aluminum stress.Different sugarcane varieties have different resistance abilities to aluminum stress.