Abstract：Sucrose phosphate synthase (SPS, EC 2.4.1.14) is a key enzyme for sucrose metabolism in plant. In sugarcane, at least 5 genes were classified into 4 subfamilies of SPS. Of the 5 SPS genes, SPSⅢ has high-level expression in the mature stem tissue of sugarcane, which is considered to be a Poaceae-specific member. Therefore, the study of binding protein genes of Cis-element in promoter region of SPSⅢ would provide initial insight for regulation of sucrose metabolism. In this study, to specific analyze the function of ATCT-motif and CAT-box from SPSⅢ in sugarcane (Saccharum spp.) for potential Cis-element of light response ATCT-motif and meristem-specific, the fragment containing ATCT-motif and CAT-box from SPSⅢ promoter region (from －1 320 to－1 210 bp) were cloned into T-vector, then, yeast one-hybrid bait vector pAbAi-SPSⅢ-AC were constructed and transformed to yeast. Subsequently, the ds-cDNA from mature leave of sugarcane were transformed to the yeast one-hybrid bait vector for constructing the Yeast One-hybrid Library System, and the insert sizes of the library were identified to be ranged from 0.25 kb to 1.5 kb. Fifty four positive clones of the candidate binding protein cDNA sequences were obtained from the Yeast One-hybrid Library, of them, 14 sequences were observed to be unique based on sequencing. Blast analysis showed that, besides E1-3, E9-1, and E0-3, the other 11 cDNA sequences had high homology to the close relatives of sugarcane with identities above 90%. E0-3 was found no Blast hit in the NCBI, which could be undiscovered gene or sugarcane specific gene. Furthermore, the potential Cis-element sequences were analyzed through SMART and SBASE. The results showed that five of the sequences, E0-3, E2-3, F2-1, F4-2 and G8-2, contained regulated motif domain. E2-3 was assumed to be a light-regulated element for SPSⅢ-AC due to the photo morphogenesis relative domain transcript presented in the corresponding amino acid sequence. BURP domain was proved to take part in the regulation of photosynthesis in bacterial. Based on the analysis of conserved domain by Blastx, a BURP domain with CH-X(10)-CH-X(25-27)-CH-X(25-26)-CH motif was found in sequence F2-1. Both of E0-3 and F4-2 contained AT-hook motif, which is DNA binding element in plant and was demonstrated to play roles in photosynthesis and the regulation of relative target genes. G8-2 belongs to super gene families of P-loop_NTPase with a conserved phosphate binding motif. The other ten sequences were found no homologous sequence relative to transcript factors. This study provides a frame for further investigating sucrose mechanism and gene engineering for sugar improvement of sugarcane.
