| Sugarcane is a commercially important sugar crop grown in tropical and sub-tropical regions. To achieve the ultimate aim of increasing the sucrose content in sugarcane stem, it is necessary to research the principal rate limiting steps in the entire sucrose accumulation processes. In the pathway of sucrose synthesis, sucrose phosphate synthase (SPS) is a key rate-limiting enzyme in sucrose formation and believed to exercise the most influence on sucrose accumulation and carbon partitioning processes.The research mainly focused on molecular cloning and 5' flanking promoter region deletion analysis of SPS gene from sugarcane. The main results of the research are as follows:1. The DNA framents(EU278617, EU278618) and cDNA framents coding entire SPSâ…¢were isolated from sugarcane. Analysis of the nucleotide sequence indicated that the gene contained thirteen exons, twelve introns, and the reading frame encoded a protein of 964 amino acids.2. On the basis of the SPSâ…¢cDNA sequences, plant expression vectors were constructed. The recombinant was transformed into Arabidopsis thaliana by Agrobacterium-mediated method.3. The 5'-flanking sequence of SPSâ…¢was cloned from sugarcane genome DNA. Transcriptional factor binding analysis indicated that, this novel sequence contained TATA box as well as other DNA elements involved in light-response and tissue-specific regulation.4. These 5'-flanking sequences of SPSâ…¢were fused to the coding sequence of GUS gene in different lengths to construct chimeric genes. All chimeric genes were bombarded into sugarcane embryogenic calli for transient GUS expression, and transformed into Arabidopsis thaliana by floral-dip method for stable GUS expression. The results confirmed the promoter activtity of 5'-flanking sequence of SPSâ…¢.5. A cDNA frament (EU269038) coding entire SPSâ…¡was isolated from sugarcane, which contains a 3183 bp open reading frame (ORF), encoding a 1060 amino acids. |
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