Cloning For Full-Length Pyruvate Orthophosphate Dikinase Of Sugarcane

Compared with C3 plant, C₄ plant has an efficient photosynthesis, and photorespiration is negligible. C₄ plant has evident growth advantage, and higher rates of water and nutrition using, especially in the condition of high light intensities and high temperatures. So C₄ plant has higher bio-yield than C3 plant.In this article, we want to clone full-length pyruvate, orthophosphate dikinase (PPDK) gene of sugarcane, preparing for transferring it into C3 plant. DNA was extracted from sugarcane leaves, primers were designed by the cDNA sequence of PPDK gene from GenBank. Then amplified via LA-PCR(Long Acute PCR) method, ligated into vectors, transformed E.coli. JM109, sequenced. At last, the full-length PPDK gene was splitted to two parts, each introduced a digest site, transformed E.coli. JM109. The strains were storaged in our lab. For checking the validity of experiment result, total RNA of sugarcane was extracted. Then amplified via RT-PCR, ligated into vectors, transformed E.coli. JM109, sequenced. By comparing our cDNA sequence and full-lenth DNA sequence results with GenBank cDNA sequence, we find that the cDNA sequence of PPDK gene in the experiment has 12bp false priming, the cDNA sequence of PPDK gene on GenBank has 29bp false priming, the exon parts of full-length PPDK gene have 10bp false priming.Full-length PPDK gene sequence of sugarcane is obtained, which has been submitted to the GenBank. Full-length PPDK gene sequence of sugarcane is 13551bp in length, including 19 exons, which were separated by 18 introns. The sequences of exon part are very short less than several hundreds base pairs. But the intron parts are different, the first and the third introns are bigger than the others. Especially the first intron is unusually long, 7164bp in length, the others were very short between 78bp and 253bp.