Molecular cloning, location and promoter analysis of NDH dehydrogenase gene from Santalum album

Abstract

In order to investigate the function and regulation mechanism of NDH dehydrogenase gene in Santalum album, the technique of RACE was used to amplify the full-length sequence of SaNDH6 with heartwood as material. The technique of quantitative real-time fluorescence PCR (RT-qPCR) was employed to analyze its expression in different tissues and after hormone induction. The subcellular location was determined by Arabidopsis thaliana protoplast transient expression. 2 kb cis-acting element upstream of start codon ATG was analyzed by PlantCARE online service, and the transcription factors which could bind the cis-acting elements was predicted by PlantRegMap software. The results were as follows: (1) SaNDH6 encoded 303 amino acids. It was a hydrophobin and located in chloroplast. (2) The phylogenetic tree analysis indicated that SaNDH6 had a more closely evolutionary relationship with NDH6 from woody plants. (3) Plant care analysis showed that the promoter sequence of SaNDH6 contained a large number of light responsive cis-acting elements such as ACE, AE-box, Box 4, G-Box and GT1-motif. It also contained abscisic acid (ABA) responsive element ABRE, jasmonic acid methyl ester (MeJA) responsive elements CGTCA-motif and TGACG-motif, gibberellin (GA3) responsive elements P-box, ARE cis-acting regulatory element essential for the anaerobic induction, and TC-rich repeats element involved in defense and stress responsiveness. (4) The results of plantRegMap analysis showed that there were 76 transcription factors that could bind to the SaNDH6 promoter, and among which, ERF transcription factor was the most (up to 40 TFs). (5) SaNDH6 can be expressed in the tissues of roots, heartwoods, calluses and leaves, but had a higher expression level in the tissue of leaves; under 1×10-4 mol·L-1 MeJA and GA3 treatments, the expression level of SaNDH6 were significantly elevated after 3 h when compared with 0 h, respectively. In conclusion, SaNDH6 was a nucleus gene encoding protein, its expression was induced by light and some hormones, and it might be involved in against some defense and stress processes in S. album.

Keywords

• santalum album
• chloroplast
• ndh dehydrogenase
• subcellular location
• expression regulation