[Objective] This study aimed to analyze the effects of temperature on the expression of AGPase isoform genes in rice endosperm during milk stage. [Method] Different temperature treatments （33 and 25 ℃ of daily mean ...[Objective] This study aimed to analyze the effects of temperature on the expression of AGPase isoform genes in rice endosperm during milk stage. [Method] Different temperature treatments （33 and 25 ℃ of daily mean temperature for high and normal temperature treatments, respectively） and the real-time fluorescence quantitative PCR （ FQPCR） were used to analyze the expression patterns of seven isoforms （AGPS1, AGPS2a, AGPS2b, AGPL1, AGPL2, AGPL3 and AGPL4） of ADPglucose pyrophosphorylase （AGPase） which was the key enzyme in starch synthesis and metabolism in rice endosperm of two rice varieties Teqing and Thai Fragrant Rice. [Result] The AGPase isoforms AGPS2b, AGPL2 and AGPL3 had much higher expression than the other four isoforms, thus they were thought to be the main expression patterns of AGPase in rice endosperm. The relative expressions of AGPL2 was the highest among all the isoforms. The relative expressions of AGPS2b, AGPL2 and AGPL3 were higher in the normal temperature treatment than in the high temperature treatment in both rice varieties. The relative expression of the three enzyme genes in milk stages in Teqing was higher than those in Thai Fragrant Rice under different temperature treatments. [Conclusion] This study provides a theoretical basis for further use of molecular biology techniques to cultivate stable high-quality rice varieties.展开更多
GDP-D-mannose pyrophosphorylase （GMPase） catalyses the synthesis of GDP-D-mannose and represents the first committed step in the synthesis of ascorbate. In the present study, the GMPase gene of tomato was introduced...GDP-D-mannose pyrophosphorylase （GMPase） catalyses the synthesis of GDP-D-mannose and represents the first committed step in the synthesis of ascorbate. In the present study, the GMPase gene of tomato was introduced into potato by Agrobacterium-mediated transformation. Two transgenic lines with higher GMPase expression were selected using qPCR and protein blot analyses. The results showed that the content of L-ascorbic acid （AsA） and the ratio of AsA/ DHA （dehydroascorbate） significantly increased in both leaves and tubers of transgenic potato plants. Both pigment content and photosynthetic rate were much higher in transgenic plants than in wild-type plants. Transgenic plants showed a distinguishable change in phenotype from the wild-type plants. Furthermore, transgenic plants showed delayed senescence.展开更多
A method based on degenerate Oligo primed polymerase chain reaction (PCR) and random amplification of cDNA end (RACE) PCR for cloning a full length cDNA is described. An Amorpha fruticosa cDNA clone encoding UDP gluco...A method based on degenerate Oligo primed polymerase chain reaction (PCR) and random amplification of cDNA end (RACE) PCR for cloning a full length cDNA is described. An Amorpha fruticosa cDNA clone encoding UDP glucose pyrophosphorylase (UGP), a key enzyme producing UDP glucose in the synthesis of sucrose and cellulose, is cloned by using this method. We design 5’ RACE primers based on UGPA1 fragment, which obtains from degenerate PCR. Inverse PCR and nested PCR enable cloning of the remainder 5’ and 3’ end fragments of the gene. The deduced amino acid sequence exhibits significant homology with the other UGP genes cloned. This method is more simple and inexpensive than screening cDNA library, and can be easily adapted to clone other genes.展开更多
Dear Editor,L-Fucose(6-deoxy-L-galactose,fucose)is the basic compone nt of a variety of glyca n structures.The fucosylated oligosaccharides participate in a variety of cellular activities,like the cell-cell recognitio...Dear Editor,L-Fucose(6-deoxy-L-galactose,fucose)is the basic compone nt of a variety of glyca n structures.The fucosylated oligosaccharides participate in a variety of cellular activities,like the cell-cell recognition,selectin-mediated leukocyteendothelial adhesion and the formation of Lewis blood group antigens(Ma et al.,2006).GDP-fucose is an important fucose donor in the process of fucosylated oligosaccharides formation.Two pathways of GDP-fucose synthesis are present in the cytosol of mammalian cells,including the de novo pathway and the salvage pathway(Becker et al.,2003).In the salvage pathway,cells use fucose from the extracellular or lysosomal sources to synthesize GDP-fucose.展开更多
Dynamic changes of starch, amylose, sucrose contents and the activities of starch synthesis enzymes under shading treatments alter flowering were studied using two rice varieties IR72 （indica） and Nipponbare （japon...Dynamic changes of starch, amylose, sucrose contents and the activities of starch synthesis enzymes under shading treatments alter flowering were studied using two rice varieties IR72 （indica） and Nipponbare （japonica） as materials. Under shading treatments, the starch, amylose and sucrose contents decreased, while ADP-glucose pyrophosphorylase （ADPGPPase） activity only changed a little, soluble starch synthase activity and granule bound starch synthase activity decreased, soluble starch branching enzyme （SSBE, Q-enzyme） activity and granule bound starch branching enzyme （GBSBE, Q-enzyme） activity increased, and starch debranching enzyme （DBE, R-enzyme） activity varied with varieties. Correlation analyses showed that the changes of starch content were positively and significantly correlated with the changes of sucrose content in the weak light. Both ADPGPPase activity and SSBE activity were positively and significantly correlated with starch accumulation rate. It was implied that the decline of starch synthase activities was related to the decrease of starch content and the increase of the activity of starch branching enzyme played an important role in the decrease of the ratio of amylose to the total starch under the weak light.展开更多
L-Ascorbic acid （AsA） plays an important role in plants and animals. In plants, GDP-D-mannose pyrophosphorylase （GMP） is essential in the AsA biosynthetic pathway. However, little is known about the genes encoding...L-Ascorbic acid （AsA） plays an important role in plants and animals. In plants, GDP-D-mannose pyrophosphorylase （GMP） is essential in the AsA biosynthetic pathway. However, little is known about the genes encoding GMP in soybean and here we report genetic and functional analysis of the GmGMP1 （Glycine max GDP-D-mannose pyrophosphorylase 1） gene in this species. GmGMP1 encoded a GDP-mannose pyrophosphorylase and exhibited higher transcript levels in the leaf than in the root, stem, flower, and seed. Transcript of this gene was ubiquitous in the vegetative and reproductive organs, and was induced by abiotic stress and light. Increasing expression of GmGMP1 in Arabidopsis and soybean through an overexpressing approach caused pronounced enhancement of AsA content, and was implicated in lowering the superoxide anion radical content and lipid peroxidation levels in Arabidopsis, and conferring tolerance to osmotic and high salt stresses during seed germination. The present study represents the first systematic determination of soybean genes encoding GDP-mannose pyrophosphorylase and provides useful evidence for the functional involvement of GmGMP1 in control of AsA content and conferring tolerance to osmotic and salt stress.展开更多
Under-development of pea (Pisum sativum L.) seeds in normally developed pods, resulting in “unfilled pods”, has been a serious problem in the greenhouse cultivation in Wakayama Prefecture, one of the major regions o...Under-development of pea (Pisum sativum L.) seeds in normally developed pods, resulting in “unfilled pods”, has been a serious problem in the greenhouse cultivation in Wakayama Prefecture, one of the major regions of pea cultivation in Japan. This phenomenon appears to be induced by low temperature and low solar irradiation during pea growing season. This study examined the relationship between this seed under-development and sucrose-to-starch metabolism in developing seeds and pods after flowering. The starch content, ADP-glucose pyrophosphorylase (AGPase) and sucrose synthase activities of a pea cultivar, Kishuusui, were monitored through seed development in shading-treatment plot and in control plot. Results showed that the present treatment induced the depression of starch accumulation and AGPase activity in developing seeds compared with control, which might cause the occurrence of under-development of seeds and unfilled pods. Surplus carbohydrates from source organs might be stored in pod walls, as expected from higher starch level and AGPase activity in treated pod walls. The necessity of thermostable AGPase variants in pea to prevent the unfilled-pod problem was discussed.展开更多
Ammonium （NH4^＋） is an important form of nitrogen nutrient for most plants, yet is also a stressor for many of them. However, the primary events of NH4^＋ toxicity at the cellular level are still unclear. Here, we ...Ammonium （NH4^＋） is an important form of nitrogen nutrient for most plants, yet is also a stressor for many of them. However, the primary events of NH4^＋ toxicity at the cellular level are still unclear. Here, we showed that NH4^＋ toxicity can induce the root cell death in a temporal pattern which primarily occurs in the cells of root maturation and elongation zones, and then spreads to the ceils in the meristem and root cap. The results from the NH4^＋-hypersensitive mutant hsn1 further confirmed our findings. Taken together,NH4^＋ toxicity inhibits primary root growth by inhibiting cell elongation and division and inducing root cell death.展开更多
The dynamic changes of the activities of enzymes involving in starch biosynthesis, including ADP-glucose pyrophosphorylase (AGPase), soluble starch synthases (SSS), starch branching enzyme (SBE) and starch debranching...The dynamic changes of the activities of enzymes involving in starch biosynthesis, including ADP-glucose pyrophosphorylase (AGPase), soluble starch synthases (SSS), starch branching enzyme (SBE) and starch debranching enzymes (DBE) were studied, and changes of fine structure of amy- lopectin were characterized by isoamylase treatment during rice grain development, using trans anti-waxy gene rice plants. The relationships between the activities of those key enzymes were also analyzed. The amylose synthesis was significantly inhibited in transgenic Wanjing 9522, but the total starch content and final grain weight were less affected as compared with those of non-transgenic Wanjing 9522 rice cultivar. Analyses on the changes of activities of enzymes involving in starch bio- synthesis showed that different enzyme activities were expressed differently during rice endosperm development. Soluble starch synthase is relatively highly expressed in earlier stage of endosperm de- velopment, whilst maximal expression of granule-bound starch synthase (GBSS) occurred in mid-stage of endosperm development. No obvious differences in changes of the activities of AGPase and SBE between two rice cultivars investigated, except the DBEs. Distribution patterns of branches of amy- lopectin changed continually during the development of rice grains and varied between two rice culti- vars. It was suggested that amylopectin synthesis be prior to the synthesis of amylose and different enzymes have different roles in controlling syntheses of branches of amylopectin.展开更多
基金Supported by Special Project for Breeding and Cultivation of GMO Varieties of Ministry of Agriculture (2011ZX08001-001, 2011ZX08001-004)Major Science and Technology Program of Hunan, China (2011FJ1002-2)+1 种基金Natural Science Foundation of Hunan, China (09JJ3046 )Science and Technology Innovation Program of Hunan Academy of Agricultural Sciences (2009hnnkycx17)~~
文摘[Objective] This study aimed to analyze the effects of temperature on the expression of AGPase isoform genes in rice endosperm during milk stage. [Method] Different temperature treatments （33 and 25 ℃ of daily mean temperature for high and normal temperature treatments, respectively） and the real-time fluorescence quantitative PCR （ FQPCR） were used to analyze the expression patterns of seven isoforms （AGPS1, AGPS2a, AGPS2b, AGPL1, AGPL2, AGPL3 and AGPL4） of ADPglucose pyrophosphorylase （AGPase） which was the key enzyme in starch synthesis and metabolism in rice endosperm of two rice varieties Teqing and Thai Fragrant Rice. [Result] The AGPase isoforms AGPS2b, AGPL2 and AGPL3 had much higher expression than the other four isoforms, thus they were thought to be the main expression patterns of AGPase in rice endosperm. The relative expressions of AGPL2 was the highest among all the isoforms. The relative expressions of AGPS2b, AGPL2 and AGPL3 were higher in the normal temperature treatment than in the high temperature treatment in both rice varieties. The relative expression of the three enzyme genes in milk stages in Teqing was higher than those in Thai Fragrant Rice under different temperature treatments. [Conclusion] This study provides a theoretical basis for further use of molecular biology techniques to cultivate stable high-quality rice varieties.
基金supported by the National Basic Re-search Program of China (2009CB119000)the National Natural Science Foundation of China (30771473)funded by Key Laboratory of Horticultural Crops Ge-netic Improvement, Ministry of Agriculture of China
文摘GDP-D-mannose pyrophosphorylase （GMPase） catalyses the synthesis of GDP-D-mannose and represents the first committed step in the synthesis of ascorbate. In the present study, the GMPase gene of tomato was introduced into potato by Agrobacterium-mediated transformation. Two transgenic lines with higher GMPase expression were selected using qPCR and protein blot analyses. The results showed that the content of L-ascorbic acid （AsA） and the ratio of AsA/ DHA （dehydroascorbate） significantly increased in both leaves and tubers of transgenic potato plants. Both pigment content and photosynthetic rate were much higher in transgenic plants than in wild-type plants. Transgenic plants showed a distinguishable change in phenotype from the wild-type plants. Furthermore, transgenic plants showed delayed senescence.
文摘A method based on degenerate Oligo primed polymerase chain reaction (PCR) and random amplification of cDNA end (RACE) PCR for cloning a full length cDNA is described. An Amorpha fruticosa cDNA clone encoding UDP glucose pyrophosphorylase (UGP), a key enzyme producing UDP glucose in the synthesis of sucrose and cellulose, is cloned by using this method. We design 5’ RACE primers based on UGPA1 fragment, which obtains from degenerate PCR. Inverse PCR and nested PCR enable cloning of the remainder 5’ and 3’ end fragments of the gene. The deduced amino acid sequence exhibits significant homology with the other UGP genes cloned. This method is more simple and inexpensive than screening cDNA library, and can be easily adapted to clone other genes.
文摘Dear Editor,L-Fucose(6-deoxy-L-galactose,fucose)is the basic compone nt of a variety of glyca n structures.The fucosylated oligosaccharides participate in a variety of cellular activities,like the cell-cell recognition,selectin-mediated leukocyteendothelial adhesion and the formation of Lewis blood group antigens(Ma et al.,2006).GDP-fucose is an important fucose donor in the process of fucosylated oligosaccharides formation.Two pathways of GDP-fucose synthesis are present in the cytosol of mammalian cells,including the de novo pathway and the salvage pathway(Becker et al.,2003).In the salvage pathway,cells use fucose from the extracellular or lysosomal sources to synthesize GDP-fucose.
文摘Dynamic changes of starch, amylose, sucrose contents and the activities of starch synthesis enzymes under shading treatments alter flowering were studied using two rice varieties IR72 （indica） and Nipponbare （japonica） as materials. Under shading treatments, the starch, amylose and sucrose contents decreased, while ADP-glucose pyrophosphorylase （ADPGPPase） activity only changed a little, soluble starch synthase activity and granule bound starch synthase activity decreased, soluble starch branching enzyme （SSBE, Q-enzyme） activity and granule bound starch branching enzyme （GBSBE, Q-enzyme） activity increased, and starch debranching enzyme （DBE, R-enzyme） activity varied with varieties. Correlation analyses showed that the changes of starch content were positively and significantly correlated with the changes of sucrose content in the weak light. Both ADPGPPase activity and SSBE activity were positively and significantly correlated with starch accumulation rate. It was implied that the decline of starch synthase activities was related to the decrease of starch content and the increase of the activity of starch branching enzyme played an important role in the decrease of the ratio of amylose to the total starch under the weak light.
基金supported by the Genetically Modified Organisms Breeding Major Projects, China (2016ZX08004)the earmarked fund for China Agriculture Research System (CARS-004-PS10)the Program for Changjiang Scholars and Innovative Research Team in University, China (PCSIRT13073)
文摘L-Ascorbic acid （AsA） plays an important role in plants and animals. In plants, GDP-D-mannose pyrophosphorylase （GMP） is essential in the AsA biosynthetic pathway. However, little is known about the genes encoding GMP in soybean and here we report genetic and functional analysis of the GmGMP1 （Glycine max GDP-D-mannose pyrophosphorylase 1） gene in this species. GmGMP1 encoded a GDP-mannose pyrophosphorylase and exhibited higher transcript levels in the leaf than in the root, stem, flower, and seed. Transcript of this gene was ubiquitous in the vegetative and reproductive organs, and was induced by abiotic stress and light. Increasing expression of GmGMP1 in Arabidopsis and soybean through an overexpressing approach caused pronounced enhancement of AsA content, and was implicated in lowering the superoxide anion radical content and lipid peroxidation levels in Arabidopsis, and conferring tolerance to osmotic and high salt stresses during seed germination. The present study represents the first systematic determination of soybean genes encoding GDP-mannose pyrophosphorylase and provides useful evidence for the functional involvement of GmGMP1 in control of AsA content and conferring tolerance to osmotic and salt stress.
文摘Under-development of pea (Pisum sativum L.) seeds in normally developed pods, resulting in “unfilled pods”, has been a serious problem in the greenhouse cultivation in Wakayama Prefecture, one of the major regions of pea cultivation in Japan. This phenomenon appears to be induced by low temperature and low solar irradiation during pea growing season. This study examined the relationship between this seed under-development and sucrose-to-starch metabolism in developing seeds and pods after flowering. The starch content, ADP-glucose pyrophosphorylase (AGPase) and sucrose synthase activities of a pea cultivar, Kishuusui, were monitored through seed development in shading-treatment plot and in control plot. Results showed that the present treatment induced the depression of starch accumulation and AGPase activity in developing seeds compared with control, which might cause the occurrence of under-development of seeds and unfilled pods. Surplus carbohydrates from source organs might be stored in pod walls, as expected from higher starch level and AGPase activity in treated pod walls. The necessity of thermostable AGPase variants in pea to prevent the unfilled-pod problem was discussed.
基金Project supported by the National Basic Research Program (973) of China (No.2005CB120901)the China Postdoctoral Science Foundation (No.20090451463),the China Postdoctoral Special Foundation (No.201003729)
文摘Ammonium （NH4^＋） is an important form of nitrogen nutrient for most plants, yet is also a stressor for many of them. However, the primary events of NH4^＋ toxicity at the cellular level are still unclear. Here, we showed that NH4^＋ toxicity can induce the root cell death in a temporal pattern which primarily occurs in the cells of root maturation and elongation zones, and then spreads to the ceils in the meristem and root cap. The results from the NH4^＋-hypersensitive mutant hsn1 further confirmed our findings. Taken together,NH4^＋ toxicity inhibits primary root growth by inhibiting cell elongation and division and inducing root cell death.
基金the National Key Research and Development Program of China(GrantNo.G199810100)
文摘The dynamic changes of the activities of enzymes involving in starch biosynthesis, including ADP-glucose pyrophosphorylase (AGPase), soluble starch synthases (SSS), starch branching enzyme (SBE) and starch debranching enzymes (DBE) were studied, and changes of fine structure of amy- lopectin were characterized by isoamylase treatment during rice grain development, using trans anti-waxy gene rice plants. The relationships between the activities of those key enzymes were also analyzed. The amylose synthesis was significantly inhibited in transgenic Wanjing 9522, but the total starch content and final grain weight were less affected as compared with those of non-transgenic Wanjing 9522 rice cultivar. Analyses on the changes of activities of enzymes involving in starch bio- synthesis showed that different enzyme activities were expressed differently during rice endosperm development. Soluble starch synthase is relatively highly expressed in earlier stage of endosperm de- velopment, whilst maximal expression of granule-bound starch synthase (GBSS) occurred in mid-stage of endosperm development. No obvious differences in changes of the activities of AGPase and SBE between two rice cultivars investigated, except the DBEs. Distribution patterns of branches of amy- lopectin changed continually during the development of rice grains and varied between two rice culti- vars. It was suggested that amylopectin synthesis be prior to the synthesis of amylose and different enzymes have different roles in controlling syntheses of branches of amylopectin.
