ABSTRACT
Objective: To clone eight members of WRKY of transcription factor family in Camellia sinensis, and analyze their bioinformatics and expression under abiotic stress. Methods: Eight WRKY transcription factor genes were cloned from Tieguanyin cultivar by RT-PCR, and the physicochemical properties of the eight WRKY protein were analyzed by bioinformatics Methods:. At the same time, the establishment of phylogenetic tree, comparison of multiple sequences, and analysis of conserved motifs were carried out by comparing WRKY of C. sinensis with homologous genes of Arabidopsis thaliana. Quantitative real-time PCR (qRT-PCR) was used to detect the expression of eight WRKY genes under low temperature, drought, and ABA stress treatment. Results: The ORF lengths of eight WRKY genes were 1 407, 2 208, 1 302, 849, 978, 879, 1 443, and 810 bp, encoding 468, 735, 433, 282, 325, 292, 480, and 269 amino acids, respectively. GenBank accession numbers were MG298951, MG298952, MG298955, MG298956, MG298957, MG298959, MG298960, and MG298963, respectively. Phylogenetic tree and sequence alignment analysis showed that eight CsWRKYs could be divided into two groups and contained WRKYGQK conserved domain and zinc finger structures, except that CsWRKY39 lacked zinc finger structure. The expression pattern of CsWRKYs was induced under the condition of low temperature, drought, and ABA stress. The expression of CsWRKY2, CsWRKY21, CsWRKY23, CsWRKY44 and CsWRKY65 increased to more than 2 after low temperature treatment with significant response to low temperature stress. The expression of CsWRKY21, CsWRKY23, CsWRKY3,9 and CsWRKY65 was up-regulated under 12 h of drought stress and 6 h of ABA treatment. This result indicated that CsWRKYs might be closely related to stress response in C. sinensis. Conclusion: Eight CsWRKY genes from different groups were cloned, and this result indicated that CsWRKYs might be closely related to stress response in C. sinensis.
ABSTRACT
Objective To clone the UGD-glucose-dehydrogenase (CsUGD) gene involved in the polysaccharide metabolic pathway, and to analyze by bioinformatics analysis, tissue expression specificity analysis, and determination of polysaccharide content in different organs of Camellia sinensis. Methods The sequence of homologous gene was obtained by transcriptome. The bioinformatics analysis was carried out by using ProtParam, TMpred, signalP, NetPhos, SMART, SSPro 4.0 and so on. Three-dimensional structure of CsUGD protein was edited by VMD; Jalview software was used for multiple sequence alignment; MEGA5.0 was used for phylogenetic tree construction. Gene expression analysis in difference organs was performed by Real-time PCR and the determination of polysaccharide content in different organs was done by anthrone sulfuric acid colorimetric method. Results The cloned CsUGD gene (GenBank accession number MG366591) had a full length of 1 866 bp encoding a predicted protein of 480 amino acids. The results of bioinformatics showed that the protein encoded by CsUGD gene belongs to the stable hydrophilic protein with transmembrane structure but no signal peptide; Phylogenetic tree analysis showed that CsUGD keeps closest genetic relationship with Diospyros kaki. The highest expression was observed in lateral roots by RT-PCR. Determination of polysaccharides in different organs of C. sinensis by colorimetric method of anthrone and sulphuric acid showed that the content of tea polysaccharide (TPS) in lateral root was higher than other parts of C. sinensis. Conclusion The CsUGD gene was cloned from the tea plant for the first time and its important role in the growth and development of the tea tree was clarified. It also played a key role in the pathways of synthesis of C. sinensis polysaccharides, which provided a scientific basis for quality breeding of C. sinensis and improving the medicinal value of tea.