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研究生:林建吏
研究生(外文):Chien-Li Lin
論文名稱:C4光合基因轉殖水稻的氣體交換能力及C4光合酵素活性之分析
論文名稱(外文):Gas exchange analyses and activities of C4 photosynthetic enzymes in transgenic rice
指導教授:林正宏林正宏引用關係
指導教授(外文):Jenq-Horng Lin
學位類別:碩士
校院名稱:國立中興大學
系所名稱:植物學系
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
中文關鍵詞:水稻C4光合作用C4光合酵素
外文關鍵詞:ricePEPCPPDKNADP-MEC4-photosynthesis
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利用農桿菌基因轉殖法將玉米ppc、pdk及Me-2基因轉入C3糧食作物水稻中,分別得到兩組ppc (k2, k3)、pdk (k4, k5)、Me-2 (k6, k7)基因轉殖品系及三組PEPC/PPDK雜交品系 (k8, k9, k10)。
本實驗針對各品系轉殖水稻的葉片進行酵素活性分析,結果顯示大部分轉殖水稻株的光合酵素活性皆提高,其中k9品系的PEPC活性比未轉殖對照組高出15倍以上;且ppc基因轉殖株中的PEPC與玉米相似,酵素活性均受G6P保護以抵抗malate的抑制。轉殖水稻箭葉的氣體交換實驗結果顯示,各品系間的光合作用速率並無明顯差異,不過PEPC活性大量提高的品系其光飽和點增加了,且蒸散速率及氣孔導度也較未轉殖對照組為高。酵素活性與氣體交換實驗結果的相關性分析顯示,轉殖株的PEPC活性與箭葉的蒸散速率及氣孔導度略呈正相關,表示PEPC活性對植株的氣孔開閉有正面性的影響;而氣體交換實驗與穗重的分析結果則顯示,轉殖株的蒸散速率及氣孔導度與穗重呈現一定程度的正相關,表示高蒸散及高氣孔導度性狀與水稻的高產量有關;針對轉殖株的PEPC活性與穗重進行相關性分析後發現,PEPC活性與穗重之間亦呈一定程度之正相關,表示提高PEPC活性對轉殖株的穗重有促進的效果。至於PPDK及NADP-ME的活性對轉殖株的穗重則不產生明顯影響。針對轉殖水稻葉片組織進行碳同位素比的分析,結果顯示轉殖水稻的δ13C值集中在-29 0/00 ~ -30.3 0/00之間,表示轉殖水稻的光合固碳形式皆仍維持原來的C3-type,不因C4光合酵素活性提高而改變。
由上述結果可知,在水稻中轉入完整的玉米C4光合酵素基因可提高C4光合酵素活性,而提高的PEPC活性可促進水稻轉殖株的氣孔開度,並對提高水稻產量有所助益。本實驗中得到幾株具高PEPC活性、高氣孔導度或高產量等優良性狀的水稻轉殖株,將留種進行後續研究。

Three C4 photosynthetic genes of maize, ppc, pdk and Me-2 were constructed and introduced into rice (Oryza sativa. var Kitaake) using Agrobacterium-mediated transformation system. Two lots of T6-PEPC (k2, k3), T4-PPDK (k4, k5) and T4-NADP-ME (k6, k7) rice seeds were obtained as single gene transformants, while 3 lots of T3-PEPC/PPDK as hybrid transformants (k8, k9, k10). Rice seedings were planted in Wagner’s pots(1/5000 a) in green house. The 3rd fully-expanded leaves were used for assaying enzyme activities.
Transgenic rice plants showed higher enzyme activities than that of untransformed line. In k9 line, the PEPC activity increased 15-fold over that of untransformed line. G6P provided protection against malate inhibition of PEPC in ppc gene transgenic rice plants. Gas exchange measurements of flag leaves of transgenic rice plants showed that there were no significant difference in photosynthetic rates among transgenic lines. The light saturation point, transpiration rate and stomatal conductance of the lines with higher PEPC-activity were higher than untransformed line. The relationship between the gas exchange measurements and the enzyme activities of transgenic rice plants showed that PEPC-activity had a positive correlation with transpiration rate and stomatal conductance. The relationship between the tassel weight and gas exchange measurements showed that tassel weight had a positive correlation with transpiration rate and stomatal conductance of rice plants. The relationship between PEPC-activities and the tassel weight of transgenic rice plants showed that PEPC-activities had a positive correlation with the tassel weight of transgenic rice plants. The activities of PPDK and NADP-ME showed no significant difference among transgenic rice plants. The carbon isotope composition of leaf tissue of transgenic rice plants showed that all transgenic lines were still assimilating CO2 by C3-photosynthesis.
These results suggest that introducing the intact C4-photosynthetic gene into rice can increase the activities of C4- photosynthetic enzymes in rice plants, and the raising of PEPC-activity in rice plants can enhance the stomatal conductance and promote the yield of rice plants. In this experiment, we have selected some transgenic rice plants with high PEPC-activity, high stomatal conductance or high yield, and will keep their seeds for future studies.

中文摘要 …………………………………………………………… vi
英文摘要 …………………………………………………………… vii
壹、 前言 ………………………………………………………… 1
貳、 材料與方法 ………………………………………………… 11
一、實驗材料 ……………………………………………….. 11
二、實驗方法 ……………………………………………….. 12
(一) 氣體交換實驗 …………………………………... 12
1. 不同光度下之光合作用速率測定 ………….. 12
2. 最大光合作用速率、蒸散速率、氣孔導度、及
水分利用效率之測定 ………………………… 13
(二) 光合酵素活性測定 ………………………………. 13
1. 酵素萃取液之蛋白質濃度之測定 …………… 13
2. Rubisco酵素活性之測定 …………………….. 14
3. PEPC酵素活性之測定 ….…………………… 14
4. pH值及G6P、malate對PEPC酵素活性之影
響 ………….……………………………………… 15
5. PPDK酵素活性之測定 ………………………. 16
6. NADP-ME酵素活性之測定 …………………. 16
(三) 碳同位素比之測定 ………………………………. 17
(四) 穗重之測定 …………………………………….… 17
參、 結果 …………………………………………………………. 18
一、轉殖水稻的栽培與生長………………………………. 18
二、轉殖株之光合酵素活性………………………………. 18
三、pH值及G6P、malate對PEPC酵素活性的影
響………………………………………………………. 20
四、不同光度下轉殖株的光合作用速率…………………. 20
五、轉殖水稻之光合特性:最大光合作用速率、蒸散速率
與氣孔導度……………………………………………. 21
六、光合酵素與光合特性之相關性………………………. 22
七、光合特性與穗重之相關性……………………………. 23
八、碳同位素比的分析結果………………………………. 24
肆、 討論 …………………………………………………………… 42
一、水稻C4光合基因轉殖株的酵素活性之探討 ………… 42
二、水稻轉殖株的光合特性之探討 ………………………. 44
三、水稻轉殖株產量與光合特性相關性之探討 ………… 48
四、水稻轉植株之固碳形式 ……………………………… 49
伍、 參考文獻……………………………………………………… 52

李彥芬 (1999) 轉殖玉米C4光合酵素基因啟動子表現的研究。中興大學植物學研究所碩士論文。61頁。
Andreo CS, Gonzalez DH and Iglesias AA (1987) Higher plant phosphoenolpyruvate carboxylase. Structure and regulation. FEBS Lett 213:1-8
Aoyagi K, Bassham JA and Greene FC (1984) Pyruvate, orthophosphate dikinase in C3 seeds and leaves as compared to the enzyme from maize. Plant Physiol 75:387-392
Asai N, Nakajima N, Kondo N and Kamada H (1999) The effect of osmotic stress on the solutes in guard cell of Vicia faba L..Plant Cell Phyisol 40: 843-849
Asai N, Nakajima N, Tamaoki M, Kamada H and Kondo N (2000) Role of synthesis mediate by phosphoenolpyruvate carboxylase in guard cells in the regulation of stomatal movement. Plant Cell Physiol 41: 10-15
Brooks A and Farquhar GD (1985) Effect of temperature on the CO2/O2 specificity of ribulose-1,5-bisphosphate carboxylase/oxygenase and the rate of respiration in the light. Planta 165: 397-406
Brown RH, Byrd GT and Black CC (1991) Assessing the degree of C4 photosynthesis in C3-C4 species using an inhibitor of phosphoenol- pyruvate caboxylase. Plant Physiol 97: 985-989
Casati P, Spampinato CP and Andreo CS (1997) Characteristics and physiological function of NADP-malic enzyme from wheat. Plant Cell Physiol 38: 928-934
Chen Z and Spreitzer RJ (1992) How various factors influence the CO2/O2 specificity of ribulose-1,5-bisphosphate carboxylase /oxygenase. Photosynth. Res 31: 157-164
Chollet R, Vidal J and O’Leary MH (1996) Phosphoenolpyruvate carboxylase: a ubiquitous, highly regulated enzyme in plants. Annu Rev Plant Physiol. Plant Mol Biol 47: 273-298
Chu C, Dai Z, Ku MSB and Edwards GE (1990) Induction of crassulacean acid metabolism in the facultative halophyte Mesembryanthemum crystallinum by abscisic acid. Plant Physiol 93: 1253-1260
Cotelle V, Pierre JN and Vavasseur A (1999) Potential strong regulation of guard cell phosphoenolpyruvate caboxylase through phosphorylation. J Exp Bot 50: 777-783
Dai Z, Ku MSB and Edwards GE (1993) C4 photosynthesis:The CO2 concentrating mechanism and photorespiration. Plant Physiol. 103:83-90
Drincovich MF, Casati P, Andreo CS, Chessin SJ, Franceschi VR, Edwards GE and Ku MSB (1998) Evolution of C4 photosynthesis in Flaveria species: isoforms of NADP-malic enzyme. Plant Physiol 117 : 733-744
Edwards GE and Andreo CS (1992) NADP-malic enzyme from plant. Phytochem. 31: 1845-1857
Ehleringer JR, Sage RF, Flanagan LB and Pearce RW (1991) Climate change and the evolution of C4 photosynthesis. Trends Ecol Evol 6:95-99
Farguhar GD, Ehleringer JR and Hubick KT (1989) Carbon isotope discrimination and photosynthesis. Annu. Rev. Plant Physiol. Plant Mol Biol 40:503 - 537
Fukayama H, Tsushida H, Agarie S, Nomura M, Onodera H, Ono K, Lee BH, Hirose S, Toki S, Ku MSB, Makino A, Matsuoka M and Miyao M (2001) Significant accumulation of C4-specific pyruvate,orthophosphate dikinase in C3 plant, rice. Plant Physiol 127:1136-1146
Furbank RT and Taylor WC (1995) Regulation of photosynthesis in C3 and C4 plants:a molecular approach. Plant Cell 7:797-807
Gupta SK, Ku MSB, Lin JH, Zhang D, Edwards GE (1994) Light/dark modulation of phosphoenolpyruvate carboxylase in C3 and C4 species. Photo Res 42:133-143
Hata S and Matsuoka M (1987) Immunological studies of pyruvate,orthophosphate dikinase in C3 plant. Plant Cell Physiol 28:635-641
Hath MD (1987) C4 photosynthesis:a unique blend of modified biochemistry,anatomy and ultrastructure. Biochim Biophys Acta 895:81-106
Hudspeth RL, Glackin CA, Bonner J and Grula JW (1986) Genomic and cDNA clones for maize phosphoenolpyruvate carboxylase and pyruvate,orthophosphate dikinase:expression of different gene-family members in leaves and roots. Proc Natl Acad Sci USA 83:2884-2888
Hudspeth RL, Glackin CA, Dai Z, Edwards GE and Ku MSB (1992) Expression of maize phosphoenolpyruvate carboxylase in:effects on biochemistry and physiology. Plant Physiol 98:458-464
Hudspeth RL and Grula JW (1989) Structure and expression of the maize gene encoding the phosphoenolpyruvate carboxylase isozyme involved in C4 photosynthesis. Plant Mol Biol 12: 579-589
Iglesias AA and Andreo CS (1989) Purification of NADP-malic enzyme and phosphoenolpyruvate carboxylase from sugar cane leaves. Plant Cell Physiol 30(3):399-405
Jenkins CLD (1989) Effects of the phosphoenolpyruvate carboxylase inhibitor 3,3-dichloro-2dihydroxyphosphinoylmethyl-2-propenoate on photosynthesis: C4 selectivity and studies on C4 photosynthesis. Plant Physiol 89: 1231-1237
Jenkins CLD, Harris RLA and McFadden H.G (1987) 3,3-dichloro-2dihydroxyphosphinoylmethyl-2-propenoate, a new, specific inhibitor of phosphoenolpyruvate caboxylase. Biochem Int 14: 219-226
Kao W-Y and Chang K-W (1998) Stable carbon isotope ratio and nutrient contents of the Kandelia candel mangrove population of different growth forms. Bot Bull Acad Sin 39:39-45
Ku MSB, Agarie S, Nomura M, Fukayama H, Tsuchida H, Ono K, Hirose S, Toki S, Miyao M and Matsuoka M (1999) High-level expression of maize phosphoenolpyruvate carboxylase in transgenic rice plants. Nature Biotech17:76-80
Ku MSB, Murakami YK and Matsuoka M (1996) Evolution and expression of C4 photosynthesis gene. Plant Physiol 111: 949-957
Ku MSB, Wu J, Dai Z, Scott RA, Chu C and Edwards GE (1996) Photo-synthetic and photorespiratory characteristics of Flaveria species. Plant Physiol 96: 518-528
Leegood RC (1997) The regulation of C4 photosynthesis.Adv Bot Res 26:256-316
Lepiniec L, Keryer E, Philippe H, Gadal P and Cretin C (1993) Sorghum phosphoenolpyruvate carboxylase gene family:structure, regulation and molecular evolution. Plant Mol Biol 21:487-502
Lipka V, Hausler RE, Rademacher T, Li J, Hirsch H-J and Kreuzaler F (1999) Solanum tuberosum double transgenic expressing phosphoenolpyruvate carboxylase and NADP-malic enzyme display reduced electron requirement for CO2 fixation. Plant Sci 144: 93-105
Long JJ and Berry JO (1996) Tissue-specific and light-mediate expression of the C4 photosynthetic NAD-dependent malic enzyme of Amaranth mitochondria. Plant Physiol 112: 473-482
Lu ZM, Percy RG, Quanlse CO and Zeiger E (1998) Stomatal conductance predicts yields in irrigated Pima cotton and breed weat grown at high temperatures. J Exp Bot 49:453 — 460
Magnin NC, Cooley BA, Reiskind J and Bowes G (1997) Regulation and localization of key enzymes during the induction of Kranz-Less, C4-type photosynthesis in Hydrilla verticillata. Plant Physiol 115: 1681-1689
Maurino VG, Drincovich MF and Andreo CS (1996) NADP-malic enzyme isoform in maize leaves. Biochemistry and Molecular Biology International 38: 239-250
Maurino VG, Drincovich MF, Casati P, Andreo CS, Edwards GE, Ku MSB, Gupta SK and Franceschi VR (1997) NADP-malic enzyme: immunolocalization in different tissues of the C4 plant maize and the C3 plant wheat. J Exp Bot 48: 799-811
Moore P (1982) Evolution of photosynthetic pathway in flowering plant.Nature 295:647-648
O’Leary MH (1982) Phosphoenolpyruvate carboxylase:An enzymologist’s view. Ann Rev Plant Physiol 33:297-315
O’Leary MH (1988) Carbon isotopes in photosynthesis. BioScience 38:328-333
Parvathi K and Raghavendra AS (1997) Both rubisco and phosphoenopyruvate caboxylase beneficial for stomatal function in epidermal strips of Commelina benghalensis. Plant Sci 124: 153-157
Poetsh W, Hermans J and Westhoff P (1991) Multiple cDNA of phosphoenolpyruvate carboxylase in C4 dicot Flaveria trinervia. FEBS Lett 292:133-136
Pupillo P and Bossi Patrizia (1979) Two forms of NADP-dependent enzyme in expending maize leaves. Planta 144:283-289
Rosche E, Streubel M and Westhoff P (1994) Primary structure of the pyruvate,orthophosphate dikinase of the C3 plant Flaveria pringlei and expression analysis of pyruvate,orthophosphate dikinase sequence in C3 , C3- C4 and C4 Flaveria species. Plant Mol Biol 26:763-769
Sage RF and Monson RK (1998) Modeling C4 photosynthesis. C4 Plant Biology PP:173~211 The Academic Press. San Diego, California.
Spencer WE, Wetzel G and Teeri J (1996) Photosynthetic phenotype plasticity and the role of phosphoenolpyruvate carboxylase in Hydrilla verticillata. Plant Sci 118: 1-9
Sugiyama T (1973) Purification, molecular, and catalytic properties of pyruvate,orthophosphate dikinase from maize leaf. Biochemistry 12:2862-2868
Taiz L and Zeiger E (1998) Photosynthesis: carbon metabolism. Plant Physiology PP: 219-248 The Benjamin/Cummings Publishing Company, Inc. Redwood City, Clifosrnia.
Tarczynski MC and Outlaw WHJr (1993) The interactive effects of pH, L-malate, and glucose-6-phosphate on guard cell phosphoenolpyruvate caboxylase. Plant Physiol 103: 1189-1194
Tagu D, Cretin C, Bergounious C, Lepiniec L and Gadal P (1991) Trnscription of a Sorghum phosphoenolpyruvate caboxylase gene in transgenic tobacco leaves:maturation of monocot pre-mRNA by dicot cells. Plant Cell Rep 9:688-690
Tsushida H, Tamai T, Fukayama H, Agarie S, Nomura M, Onodera H, Ono K, Nishizawa Y, Lee BH, Hirose S, Toki S, Ku MSB, Matsuoka M and Miyao M (2001) High level expression of C4-specific NADP-malic enzyme in leaves and impairment of photoautotrophic growth in a C3 plant, rice. Plant Cell Physiol 42(2):138-145
Usuda H, Ku MSB and Edwards GE (1985) Influence of light intensity during growth on photosynthetic enzymes in a C4 plant(Zea mays). Plant Physiol 63:65-70
Voznesenskaya EW, Franceschi VR, Kiirats O, Freitag H and Edwards GE (2001) Kranz anatomy is not essential for terrestrial C4 plant photosynthesis. Nature 414:543-546
Yen HE, Edwards GE and Grimes HD (1994) Characterization of a salt-responsive glycoprotein in the halophyte Mesembryanthemum crystallinum. Plant Physiol. 105:1179-1187

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