Inorganic pyrophosphatase hydrolyzes inorganic pyrophosphate (PPi) to two ions of phosphate radical (Pi) in vivo. The timely degradation of PPi has been recognized as a key ratelimiting step in the process of disaccharide synthesis, especially sucrose. However, maintaining certain concentration of PPi in vascular bundle cells is required for the translocation of sucrose. Therefore, it could be the best resolution to overexpress inorganic pyrophosphatase gene only in mesophyll tissue, which would feed back and accelerate photosynthesis and produce more carbohydrate. In this research, we firstly analyzed 30 inorganic pyrophosphatase genes predicted in rice by a series of bioinformatics softwares, such as protein clusters, amino acids sequence alignment, protein subcellular localization prediction and ciselements analysis. Secondly, one gene (Os04g0687100) that was considered encoding soluble cytoplasmic inorganic pyrophosphatase, named as OsIP1, was cloned and placed under the rice mesophyllspecific promoter cyFBPase from a cytosolic fructose1,6bisphosphatase gene. Thirdly, we transformed this chimeric gene, cyFBPase:OsIP1, into two different sourcesink type rice varieties using Agrobacteriummediated method, and obtained a total of 48 independent positive transgenic plants.
ZHANG Yafang, YU Yongqi, ZUO Shimin, LOU Lijuan, CHEN Zongxiang, PAN Xuebiao*
. Cloning of an Inorganic Pyrophosphatase Gene OsIP1 in Rice Based on Bioinformatics Analysis and its Genetic Transformation[J]. Chinese Journal OF Rice Science, 2013
, 27(1)
: 17
-25
.
DOI: 10.3969/j.issn.10017216.2013.01.003
\[1\]王延枝. 植物液泡膜上的焦磷酸酶. 植物生理学通讯, 1990, 4: 7376.
\[2\]王异星, 李明启. 焦磷酸在植物细胞能量代谢中的作用. 热带亚热带植物学报, 1997, 5(4): 8590.
\[3\]张宁, 司怀军, 柳娜, 等. 马铃薯无机焦磷酸酶基因cDNA 在大肠杆菌中的表达及蛋白质特征分析. 分子植物育种, 2008, 6(6): 11071110.
\[4\]李小园, 曾日中, 校现周. 植物焦磷酸酶(PPase)的研究进展. 生命科学研究, 2004, 8(4): 8387.
\[5\]司丽珍, 储成才. 植物蔗糖合成的分子机制. 中国生物工程杂志, 2003, 23(1): 1116.
\[6\]Sonnewald U. Expression of E. coli inorganic pyrophosphatase in transgenic plants alters photoassimilate partitioning. Plant J, 1992, 2(4): 571581.
\[7\]Lerchl J, Geigenberger P, Stitt M,et al. Impaired photoassimilate partitioning caused by phloemspecific removal of pyrophosphate can be complemented by a phloemspecific cytosolic yeastderived invertase in transgenic plants. Plant Cell, 1995(7): 259270.
\[8\]Si L Z, Cao S Y, Chu C C. Isolation of a 1195bp 5′Flanking region of rice cytosolic fructose1,6bisphosphatase and analysis of its expression in transgenic rice. Acta Bot Sin, 2003, 45(3): 359364.
\[9\]廖志勇. 外源无机焦磷酸化酶基因（PPa）转化水稻的效应研究\[学位论文\]. 扬州： 扬州大学, 2006: 4041.
\[10\] 张亚芳, 陈宗祥, 娄丽娟. 几个水稻品种源库关系分析. 扬州大学学报：农业与生命科学版, 2011, 32(1): 69.
\[11\] 曹显祖, 朱庆森. 水稻品种的源库特征及其类型划分的研究.作物学报, 1987, 13(4): 265272.
\[12\] Hiei Y, Ohta S, KomarionT, et al. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the TDNA. Plant J, 1994, 6(2): 271282.
\[13\] 黄健秋, 卫志明, 安海龙, 等. 根癌土壤杆菌介导的水稻高效转化和转基因植株的高频再生. 植物生理学报, 2000, 42(11): 11721178.
\[14\] 张亚芳, 左示敏, 王力, 等. 一个籼稻推广品种优良组培特性的发现. 扬州大学学报：农业与生命科学版, 2005, 26(4): 4751.
\[15\]Porebski S, Bailey L G, Baum B R. Modification of a CTAB DNA extraction protocol for plants containing high polysaccharide and polyphenol components. Plant Mol Biol Rep, 1997, 15(1): 815.
\[16\]Schulze S, Mant A, Kossmann J, et al. Identification of an Arabidopsis inorganic pyrophasphatase capable of being imported into chloroplasts. FEBS Lett, 2004, 565(1/3): 101105.
\[17\]Koroleva O A, Tomlinson M L, Leader D, et al. Highthroughput protein localization in Arabidopsis using Agrobacteriummediated transient expression of GFPORF fusions. Plant J, 2005, 41(1): 162174.
\[18\] May A, Berger S, Hertel T, et al. The Arabidopsis thaliana phosphate starvation responsive gene AtPPsPase1 encodes a novel type of inorganic pyrophosphatase. Biochim Biophys Acta, 2011, 1810(2): 178185.
\[19\] Drozdowicz Y M, Rea P A. Vacuolar H(+) pyrophosphatases: From the evolutionary backwaters into the mainstream.Trends Plant Sci, 2001, 6(5): 206211.
\[20\] Drozdowicz Y M, Lu Y P, Patel V, et al. A thermostable vacuolartype membrane pyrophosphatase from the archaeon Pyrobaculum aerophilum: Implications for the origins of pyrophosphateenergized pumps. FEBS Lett, 1999, 460(3):505512.
\[21\] Gilmartin P M, Sarokin L, Memelink J, et al. Molecular light switches for plant genes. Plant Cell, 1990, 2(5):369378.
\[22\] Gowik U, Burscheidt J, Akyildiz M, et al. Cisregulatory elements for mesophyIIspecific gene expression in the C4 plant Flaveria trinervia, the promoter of the C4 phosphoenolpyruvate carboxylase gene. Plant Cell, 2004, 16(5): 10771090.
\[23\] Degenhardt J, Tobin E M. A DNA binding activity for one of two closely defined phytochrome regulatory elements in an Lhcb promoter is more abundant in etiolated than in green plants. Plant Cell, 1996, 8(1): 3141.
\[24\] Zhou D X. Regulatory mechanism of plant gene transcription by GTelements and GTfactors. Trends Plant Sci, 1999, 4 (6): 210214.
\[25\] Schunmann P H, Richardson A E, Vickers C E, et al. Promoter analysis of the barley Pht1;1 phosphate transporter gene identifies regions controlling root expression and responsiveness to phosphate deprivation. Plant Physiol, 2004, 136(4): 42054214.
\[26\] 张春凤. 转PPase基因马铃薯块茎休眠和发芽特性的分析与评价 \[学位论文\]. 兰州：甘肃农业大学, 2009: 5152.
\[27\] 黄东杰, 张树珍, 冯翠莲, 等.无机焦磷酸化酶基因的克隆与植物表达载体的构建. 热带作物学报, 2007, 28(2): 6973.
\[28\] 赵丽宏, 王俊刚, 杨本鹏, 等.无机焦磷酸化酶基因转化甘蔗的遗传研究. 生物技术通报, 2009, 9: 7377.
