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1.
Seeds of yellow lupine (Lupinus luteus L. cv. Juno) were collected throughout their development on the mother plant to determine whether the ability to germinate and to tolerate desiccation is related to the level of free radicals and the changes in the redox state of ascorbate and glutathione as well as the activities of antioxidative enzymes. Electron paramagnetic resonance (EPR)-based analyses showed that development of lupine seed was accompanied by generation of free radicals with g1 and g2 values of 2.0049 ± 0.0004 and 2.0029 ± 0.0003, respectively. Free radical level increased significantly 25 DAF and decreased thereafter. The amount of hydrogen peroxide was high in fresh immature seeds and decreased during maturation drying. Ascorbate accumulated in lupine embryos during early seed filling stage whereas glutathione content increased during late seed filling phase. During maturation drying the redox state of both ascorbate and glutathione pools shifted towards the oxidized forms. While superoxide dismutase (SOD, EC 1.15.1.1), and ascorbate peroxidase (APX, EC 1.11.1.11) activities remained high at the early seed filling stage the activities of both dehydroascorbate reductase (DHAR, EC 1.8.5.1) and glutathione reductase (GR, EC 1.6.4.2) and that of catalase (CAT, EC 1.11.1.6) increased before seeds reached physiological maturity and decreased thereafter. The changes of isoform patterns of antioxidative enzymes were observed during seed maturation. Immature lupine seeds tested immediately after harvest acquired the ability to germinate when less than half-filled and reached high tolerance to desiccation just after physiological maturity. The physiological implications of the changes in antioxidative machinery for the acquisition of desiccation tolerance and seeds germinability are discussed.  相似文献   

2.
Plant homeobox genes play an important role in plant development, including embryogenesis. Recently, the function of a class I homeobox of knox 3 gene, HBK3, has been characterized in the conifer Picea abies (L.) Karst (Norway spruce) [8]. During somatic embryogenesis, expression of HBK3 is required for the proper differentiation of proembryogenic masses into somatic embryos. This transition, fundamental for the overall embryogenic process, is accelerated in sense lines over-expressing HBK3 (HBK3-S) but precluded in antisense lines (HBK3-AS) where the expression of this gene is experimentally reduced. Altered HBK3 expression resulted in major changes of ascorbate and glutathione metabolism. During the initial phases of embryogeny the level of reduced GSH was higher in the HBK3-S lines compared to their control counterpart. An opposite profile was observed for the HBK3-AS lines where the glutathione redox state, i.e. GSH/GSH + GSSG, switched towards its oxidized form, i.e. GSSG. Very similar metabolic fluctuations were also measured for ascorbate, especially during the transition of proembryogenic masses into somatic embryos (7 days into hormone-free medium). At this stage the level of reduced ascorbate (ASC) in the HBK3-AS lines was about 75% lower compare to the untransformed line causing a switch of the ascorbate redox state, i.e. ASC/ASC + DHA + AFR, towards its oxidized forms, i.e. DHA + AFR. Changes in activities of several ascorbate and glutathione redox enzymes, including dehydroascorbate reductase (EC 1.8.5.1), ascorbate free radical reductase (EC 1.6.5.4) and glutathione reductase (GR; EC 1.6.4.2) were responsible for these metabolic differences. Data presented here suggest that HBK3 expression might regulate somatic embryo yield through alterations in glutathione and ascorbate metabolism, which have been previously implicated in controlling embryo development and maturation both in vivo and in vitro.  相似文献   

3.
The ascorbate and glutathione systems have been studied during the first stages of germination in orthodox seeds of the gymnosperm Pinus pinea L. (pine). The results indicate that remarkable changes in the content and redox balance of these metabolites occur in both the embryo and endosperm; even if with different patterns for the two redox pairs. Dry seeds are devoid of the ascorbate reduced form (ASC) and contain only dehydroascorbic acid (DHA). By contrast, glutathione is present both in the reduced (GSH) and in the oxidized (GSSG) forms. During imbibition the increase in ASC seems to be mainly caused by the reactivation of its biosynthesis. On the other hand, the GSH rise occurring during the first 24 h seems to be largely due to GSSG reduction, even if GSH biosynthesis is still active in the seeds. The enzymes of the ascorbate--glutathione cycle also change during germination, but in different ways. ASC peroxidase (EC 1.11.1.11) and glutathione reductase (EC 1.6.4.2) activities progressively rise both in the embryo and in endosperm. These changes are probably required for counteracting production of reactive oxygen species caused by recovery of oxidative metabolism. The two enzymes involved in the ascorbate recycling, ascorbate free radical (AFR) reductase (EC 1.6.5.4) and DHA reductase (EC 1.8.5.1), show different behaviour: the DHA reductase activity decreases, while that of AFR reductase remains unchanged. The relationship between ascorbate and glutathione metabolism and their relevance in the germination of orthodox seeds are also discussed.  相似文献   

4.
The ascorbate-glutathione system was studied during development and maturation of beech (Fagus sylvatica L.) seeds, the classification of which in the orthodox category is controversial. This study revealed an increase in glutathione content after acquisition of desiccation tolerance, which was more intensive in embryonic axes than in cotyledons. During seed maturation, the redox status of glutathione markedly changed toward the more reducing state, especially in cotyledons. Ascorbic acid content decreased during maturation, mostly in cotyledons. Activities of the enzymes of the ascorbate-glutathione cycle—ascorbate peroxidase (APX, EC 1.11.1.11), monodehydroascorbate reductase (MR, EC 1.6.5.4), dehydroascorbate reductase (DHAR, EC 1.8.5.1) and glutathione reductase (GR, EC 1.6.4.2)—were markedly higher in embryonic axes than in cotyledons throughout the study period. In the course of seed maturation, the activities of these enzymes decreased. Importance of the ascorbate-glutathione cycle in desiccation tolerance of beech seeds was discussed in relation to results for typical orthodox and recalcitrant seeds of other broadleaved species.  相似文献   

5.
以携带谷胱甘肽转移酶(GST)和过氧化氢酶(CAT1)的转基因水稻和非转基因水稻(Oryza sativa L.) 品种'中花11'的根系为材料, 比较分析了二者在PEG 6000、38℃及PEG 6000和38℃复合胁迫下抗氧化系统特别是抗坏血酸-谷胱甘肽循环系统的变化.结果显示, 6% PEG处理时,转基因水稻的CAT、GST、超氧化物歧化酶(SOD)、谷胱甘肽还原酶(GR)和脱氢抗坏血酸还原酶(DHAR)的活性都显著高于非转基因水稻;38℃处理时,前者的CAT、GST、SOD和GR的活性则显著低于后者;6% PEG和38℃复合处理时,前者的CAT、GST、抗坏血酸过氧化物酶(APX)和DHAR的活性也都显著高于后者,但前者的SOD和GR活性则显著低于后者.6%PEG 诱导的转基因水稻根系的抗坏血酸氧还状态显著低于非转基因水稻,但二者的谷胱甘肽氧还状态无显著差异; 而6% PEG和38℃同时处理时,转基因水稻的谷胱甘肽氧还状态则显著高于非转基因水稻,但二者的抗坏血酸氧还状态差异不显著.研究发现,干旱和高温复合胁迫时,转基因水稻和非转基因水稻的抗氧化组分的变化均不等于这2种单一胁迫的叠加;GST和CAT1基因的转入对水稻抗氧化系统内源功能相关组分尤其是抗坏血酸-谷胱甘肽循环系统产生了一定的影响,两种水稻的根系可能利用不同的抗氧化组分调节机制对这些胁迫做出应答.  相似文献   

6.
The presence of the enzymes of the ascorbate-glutathione cycle was investigated in mitochondria and peroxisomes purified from pea (Pisum sativum L.) leaves. All four enzymes, ascorbate peroxidase (APX; EC 1.11.1.11), monodehydroascorbate reductase (EC 1.6.5.4), dehydroascorbate reductase (EC 1.8.5.1), and glutathione reductase (EC 1.6.4.2), were present in mitochondria and peroxisomes, as well as in the antioxidants ascorbate and glutathione. The activity of the ascorbate-glutathione cycle enzymes was higher in mitochondria than in peroxisomes, except for APX, which was more active in peroxisomes than in mitochondria. Intact mitochondria and peroxisomes had no latent APX activity, and this remained in the membrane fraction after solubilization assays with 0.2 M KCl. Monodehydroascorbate reductase was highly latent in intact mitochondria and peroxisomes and was membrane-bound, suggesting that the electron acceptor and donor sites of this redox protein are not on the external side of the mitochondrial and peroxisomal membranes. Dehydroascorbate reductase was found mainly in the soluble peroxisomal and mitochondrial fractions. Glutathione reductase had a high latency in mitochondria and peroxisomes and was present in the soluble fractions of both organelles. In intact peroxisomes and mitochondria, the presence of reduced ascorbate and glutathione and the oxidized forms of ascorbate and glutathione were demonstrated by high-performance liquid chromatography analysis. The ascorbate-glutathione cycle of mitochondria and peroxisomes could represent an important antioxidant protection system against H2O2 generated in both plant organelles.  相似文献   

7.
Antioxidant defences of the apoplast   总被引:1,自引:0,他引:1  
Summary The apoplast of barley and oat leaves contained superoxide dismutase (SOD), catalase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, and glutathione reductase activities. The activities of these enzymes in the apoplastic extracts were greatly modified 24 h after inoculation with the biotrophic fungal pathogenBlumeria graminis. The quantum efficiency of photosystem II, which is related to photosynthetic electron transport flux, was comparable in inoculated and healthy leaves during this period. Apoplastic soluble acid invertase activity was also modified in inoculated leaves. Inoculation-dependent increases in apoplastic SOD activity were observed in all lines. Major bands of SOD activity, observed in apoplastic protein extracts by activity staining of gels following isoelectric focusing, were similar to those observed in whole leaves but two additional minor bands were found in the apoplastic fraction. The apoplastic extracts contained substantial amounts of dehydroascorbate (DHA) but little or no glutathione (GSH). Biotic stress decreased apoplastic ascorbate and DHA but increased apoplastic GSH in resistant lines. The antioxidant cycle enzymes may function to remove apoplastic H2O2 with ascorbate and GSH derived from the cytoplasm. DHA and oxidized glutathione may be reduced in the apoplast or returned to the cytosol for rereduction.Abbreviations AA reduced ascorbate - APX ascorbate peroxidase - DHA dehydroascorbate (oxidised ascorbate) - DHAR dehydroascorbate reductase - G6PDH glucose-6-phosphate dehydrogenase - GSH reduced glutathione - GSSG glutathione disulphide - GR glutathione reductase - MDHA monodehydroascorbate - MDHAR monodehydroascorbate reductase - SOD superoxide dismutase  相似文献   

8.
Kuzniak E  Skłodowska M 《Planta》2005,222(1):192-200
Peroxisomes, being one of the main organelles where reactive oxygen species (ROS) are both generated and detoxified, have been suggested to be instrumental in redox-mediated plant cell defence against oxidative stress. We studied the involvement of tomato (Lycopersicon esculentum Mill.) leaf peroxisomes in defence response to oxidative stress generated upon Botrytis cinerea Pers. infection. The peroxisomal antioxidant potential expressed as superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6) and glutathione peroxidase (GSH-Px, EC 1.11.1.19) as well as the ascorbate-glutathione (AA-GSH) cycle activities was monitored. The initial infection-induced increase in SOD, CAT and GSH-Px indicating antioxidant defence activation was followed by a progressive inhibition concomitant with disease symptom development. Likewise, the activities of AA-GSH cycle enzymes: ascorbate peroxidase (APX, EC 1.11.1.11), monodehydroascorbate reductase (MDHAR, EC 1.6.5.4), dehydroascorbate reductase (DHAR, EC 1.8.5.1) and glutathione reductase (GR, EC 1.6.4.2) as well as ascorbate and glutathione concentrations and redox ratios were significantly decreased. However, the rate and timing of these events differed. Our results indicate that B. cinerea triggers significant changes in the peroxisomal antioxidant system leading to a collapse of the protective mechanism at advanced stage of infection. These changes appear to be partly the effect of pathogen-promoted leaf senescence.  相似文献   

9.
The response of the antioxidant system to salt stress was studied in the roots of the cultivated tomato Lycopersicon esculentum Mill. cv. M82 (Lem) and its wild salt-tolerant relative L. pennellii (Corr.) D'Arcy accession Atico (Lpa). Roots of control and salt (100 m M NaCl)-stressed plants were sampled at various times after commencement of salinization. A gradual increase in the membrane lipid peroxidation in salt-stressed root of Lem was accompanied with decreased activities of the antioxidant enzymes: superoxide dismutase (SOD; EC 1.15.1.1), catalase (CAT; EC 1.11.1.6), ascorbate peroxidase (APX; EC 1.11.1.11) and decreased contents of the antioxidants ascorbate and glutathione and their redox states. In contrast, increased activities of the SOD, CAT, APX, monodehydroascorbate reductase (MDHAR; EC 1.6.5.4), and increased contents of the reduced forms of ascorbate and glutathione and their redox states were found in salt-stressed roots of Lpa, in which the level of membrane lipid peroxidation remained unchanged. It seems that the better protection of Lpa roots from salt-induced oxidative damage results, at least partially, from the increased activity of their antioxidative system.  相似文献   

10.
Root growth and modifications of ROS-scavenging systems were investigated in transgenic rice (Oryza sativa L., cv. Zhonghua no. 11) co-expressing glutathione-S-transferase (GST, EC. 2.5.1.18) and catalase 1 (CAT1, EC 1.11.1.6) and nontransgenic rice exposed to just salt or heat and their combination. The higher number of adventitious roots but the lower root to shoot ratio were observed in the stressed transgenics as compared with nontransgenics. Most antioxidant enzymes, such as CAT, GST, ascorbate peroxidase (APX, EC 1.11.1.11), glutathione reductase (GR, EC.1.6.4.2), dehydroascorbate reductase (DHAR, EC.1.8.5.1), and the redox state of glutathione and ascorbate, measured in the transformant roots, were significantly different from those in nontransformant roots following the three types of stress. The variations of root growth and antioxidant systems in the stressed transgenic rice may be attributed to not only the GST and CAT1 transgenes but also the coordination of the ascorbate-glutathione cycle.  相似文献   

11.
12.
Compared to non-embryogenic callus, proembryonic mass, globular, and heart-shaped embryos of Eleutherococcus senticosus had higher levels of endogenous reduced glutathione (GSH). GSH content declined during the course of the embryo development (torpedo and cotyledon). Similarly, glutathione reductase that is involved in the recycling of GSH providing a constant intracellular level of GSH was also higher in globular and heart-shaped embryos. The transient increase in GSH contents also correlated with the changes in measured γ-glutamylcysteine synthetase activity over the same period. The endogenous levels of oxidized glutathione showed similar trend during development of the somatic embryos, whereas it declined in maturing somatic embryos. A pronounced increase in glutathione-S-transferase, glutathione peroxidase, catalase, and guaiacol peroxidase activity was observed during somatic embryo maturation. Ascorbate-glutathione cycle enzymes (ascorbate peroxidase; dehydroascorbate reductase and monodehydroascorbate reductase) activities also induced indicated that antioxidant enzymes played an important role during embryo development. These results suggested that the coordinated up-regulations of the antioxidant enzymes and glutathione redox system provide protection during somatic embryo development in E. senticosus. Antioxidant responses through alterations of the glutathione redox systems, have been described in the present studies have a significant role in somatic embryo development.  相似文献   

13.
14.
To explore the significance of the ascorbate–glutathione cycle under drought stress, the leaves of 2-year-old potted apple (Malus domestica Borkh.) plants were used to investigate the changes of each component of the ascorbate–glutathione cycle as well as the gene expression of dehydroascorbate reductase (DHAR, EC 1.8.5.1), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) under drought stress. The results showed that the malondialdehyde (MDA) and H2O2 concentrations in apple leaves increased during drought stress and began to decrease after re-watering. The contents of total ascorbate, reduced ascorbic acid (AsA), total glutathione and glutathione (GSH) were obviously upregulated in apple leaves when the soil water content was 40–45%. With further increase of the drought level, the contents of the antioxidants and especially redox state of AsA and GSH declined. However, levels of them increased again after re-watering. Moreover, drought stress induced significant increase of the activities of enzymes such as APX, scavenging H2O2, and also of monodehydroascorbate reductase (MDHAR, EC 1.6.5.4), DHAR and GR used to regenerate AsA and GSH, especially when the soil water content was above 40–45%. During severe drought stress, activities of the enzymes were decreased and after re-watering increased again. Gene expression of cytoplasmic DHAR, cytoplasmic APX and cytoplasmic GR showed similar changes as the enzyme activities, respectively. The results suggest that the ascorbate–glutathione cycle is up-regulated in response to drought stress, but cannot be regulated at severe drought stress conditions.  相似文献   

15.
Crop yield is severely affected by soil salinity, as salt levels that are harmful to plant growth occur in large terrestrial areas of the world. The present investigation describes the studies of enzymatic activities, in-gel assays, gene expression of some of the major antioxidative enzymes, tocopherol accumulation, lipid peroxidation, ascorbate and dehydroascorbate contents in a salt-sensitive rice genotype PB1, and a relatively salt-tolerant cultivar CSR10 in response to 200 mM NaCl. Salt solution was added to the roots of hydroponically grown 5-day-old etiolated rice seedlings, 12 h prior to transfer to cool white fluorescent?+?incandescent light (100 μmol photons m?2 s?1). Total tocopherol and ascorbate contents declined in salt-stressed rice seedlings. Among antioxidative enzymes, an increase in the activities of superoxide dismutase (EC 1.15.1.1), catalase (EC 1.11.1.6), ascorbate peroxidase (EC 1.11.1.11), glutathione reductase (EC 1.6.4.2), and their gene expression was observed in both cultivars in response to salt stress. The salt-tolerant cultivar CSR10 resisted stress due to its early preparedness to combat oxidative stress via upregulation of gene expression and enzymatic activities of antioxidative enzymes and a higher redox status of the antioxidant ascorbate even in a non-stressed environment.  相似文献   

16.
The effects of arsenite treatment on generation of reactive oxygen species, induction of oxidative stress, response of antioxidative system, and synthesis of phytochelatins were investigated in two indica rice (Oryza sativa L.) cvs. Malviya-36 and Pant-12 grown in sand cultures for a period of 5–20 days. Arsenite (As2O3; 25 and 50 μM) treatment resulted in increased formation of superoxide anion (O2.−), elevated levels of H2O2 and thiobarbituric acid reactive substances, showing enhanced lipid peroxidation. An enhanced level of ascorbate (AA) and glutathione (GSH) was observed irrespective of the variation in the level of dehydroascorbate (DHA) and oxidized glutathione (GSSG) which in turn influenced redox ratios AA/DHA and GSH/GSSG. With progressive arsenite treatment, synthesis of total acid soluble thiols and phytochelatins (PC) increased in the seedlings. Among antioxidative enzymes, the activities of superoxide dismutase (EC 1.15.1.1), catalase (EC 1.11.1.6), total ascorbate peroxidase (APX, EC 1.11.1.11), chloroplastic ascorbate peroxidase, guaiacol peroxidase (EC 1.11.1.7), monodehydroascorbate reductase (EC 1.6.5.4), and glutathione reductase (EC 1.6.4.2) increased in arsenite treated seedlings, while dehyroascorbate reductase (EC 1.8.5.1) activity declined initially during 5–10 days and increased thereafter. Results suggest that arsenite treatment causes oxidative stress in rice seedlings, increases the levels of many enzymatic and non-enzymatic antioxidants, and induces synthesis of thiols and PCs, which may serve as important components in mitigating arsenite-induced oxidative damage.  相似文献   

17.
Antioxidative responses of Calendula officinalis under salinity conditions.   总被引:10,自引:0,他引:10  
To gain a better insight into long-term salt-induced oxidative stress, some physiological parameters in marigold (Calendula officinalis L.) under 0, 50 and 100 mM NaCl were investigated. Salinity affected most of the considered parameters. High salinity caused reduction in growth parameters, lipid peroxidation and hydrogen peroxide accumulation. Under high salinity stress, a decrease in total glutathione and an increase in total ascorbate (AsA + DHA), accompanied with enhanced glutathione reductase (GR, EC 1.6.4.2) and ascorbate peroxidase (APX, EC 1.11.1.11) activities, were observed in leaves. In addition, salinity induced a decrease in superoxide dismutase (SOD, EC 1.15.1.1) and peroxidase (POX, EC 1.11.1.7) activities. The decrease in dehydroascorbate reductase (DHAR, EC 1.8.5.1) and monodehydroascorbate reductase (MDHAR, EC 1.6.5.4) activities suggests that other mechanisms play a major role in the regeneration of reduced ascorbate. The changes in catalase (CAT, EC 1.11.1.6) activities, both in roots and in leaves, may be important in H2O2 homeostasis.  相似文献   

18.
Ascorbate–glutathione systems were studied during desiccation of recalcitrant seeds of the silver maple (Acer saccharinum L.). The desiccated seeds gradually lost their germination capacity and this was strongly correlated with an increase in electrolyte leakage from seeds. Simultaneously the increase of reactive oxygen species (ROS) (superoxide radical – O2 and hydrogen peroxide – H2O2) production was observed. The results indicate that remarkable changes in the concentrations and redox status of ascorbate and glutathione occur in embryo axes and cotyledons. After shedding, concentrations of ascorbic acid (ASA) and the reduced form of glutathione (GSH) are higher in embryo axes than in cotyledons and their redox status is high in both embryo parts. Cotyledons in freshly shed seeds are devoid of GSH. At the first stages of desiccation, up to a level of 43% of moisture content, ASA content in embryo axes and GSH content in cotyledons increased. Below this level of moisture content, the antioxidant contents as well as their redox status rapidly decreased. The enzymes of the ascorbate–glutathione pathway: ascorbate peroxidase (APX) (EC 1.11.1.11), monodehydroascorbate reductase (MR) (EC 1.6.5.4), dehydroascorbate reductase (DHAR) (EC 1.8.5.1) and glutathione reductase (GR) (EC 1.6.4.2) increased their activity during desiccation, but mainly in embryonic axes. The changes are probably required for counteracting the production of ROS during desiccation. The relationship between ascorbate and glutathione metabolism and their relevance during desiccation of recalcitrant Acer saccharinum seeds is discussed.  相似文献   

19.
Ascorbate content and the activities of some key enzymes involved in the detoxification from reactive oxygen species were investigated in germinated embryos of two Zea mays L. inbred lines (B73 and Mo17) and of their heterotic F1 hybrid (B73×Mo17). The F1 hybrid showed a higher ascorbate biosynthetic capability owing to a higher activity of l -galactono- Γ -lactone dehydrogenase (EC 1.6.5.4), the last enzyme in ascorbate biosynthesis. Ascorbate peroxidase (EC 1.11.1.11), ascorbate free radical reductase (EC 1.6.5.4) and dehydroascorbate reductase (EC 1.8.5.1) activities were much higher in the F1 hybrid than in either inbred line, whereas catalase (EC 1.11.1.6) activity was similar in the three genotypes. Native polyacrylamide gel electrophoresis (PAGE) analysis showed three forms of cytosolic ascorbate peroxidase, both in parental lines and in the F1 hybrid. On the other hand, a complex pattern of proteins with dehydroascorbate reductase activity was observed, with the hybrid combining the different dehydroascorbate-reducing proteins expressed by the inbred lines. The possible involvement of the enzymes of the ascorbate system in the phenomenon of hybrid vigour is discussed.  相似文献   

20.
It is shown in experiments is vivo that development of experimental metabolic alkalosis in rats is followed by changes in redox processes in the eye retina and tunic. For the first two months of the experiment the number of sulphydryl group decreases, while that of disulphide ones of water-soluble proteins and low-molecular compounds increases. The amount of oxidized metabolites of glycolysis and of a cycle of tricarboxylic acids (pyruvate, oxaloacetate, alpha-ketoglutarate) increases relative to the reduced ones (lactate, isocitrate, malate), as well as activities of hexokinase, pyruvate kinase, NAD-dependent malate dehydrogenase, while activities of fructose diphosphatase, glucoso-6-phosphate dehydrogenase, glutathione peroxidase and glutathione reductase fall. The content of malonic dialdehyde increases. 90 days later disorders of certain compensatory mechanisms of the metabolic system of alkalosis regulation probably occurred in the eye retina and tunic tissues: hexokinase and pyruvate kinase activity fell to the control values, while that of NAD-dependent malate dehydrogenase--below the control level; the content of lactate increased. Activity of glutathione-dependent enzymes remained low and the amount of malonic dialdehyde grew much more than in the previous terms.  相似文献   

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