Arabinose is one of the most dynamic cell wall glycosyl residues released during fruit ripening, alpha-L-arabinofuranosidase (alpha-Arab) are major glycosidases that may remove arabinose units from fruit cell wall polysaccharides. To find out whether alpha-Arab plays important roles in banana fruit softening, the enzyme activities in peel and pulp, fruit firmness, respiration rate and ethylene release rate were assayed during banana softening. The results showed that alpha-Arab activities in banana pulp and peel increased slightly at the beginning of storage and reached their maxima when the fruit firmness decreased drastically, alpha-Arab activity increased by more than ten folds in both pulp and peel during ripening and alpha-Arab activities were higher in pulp than in peel. Treatment of banana fruits with ethylene absorbent postponed the time of reaching of its maxima of respiration and ethylene, enhanced the firmness of pup and decreased alpha-Arab activity in the peel and pulp. These results suggest that alpha-Arab induced the decrease of fruit firmness and played an important role in banana fruit softening, and its activity was regulated by ethylene.
Gas exchange and chlorophyll a fluorescence in soybean plants were investigated to explore the effects of iron deficiency on photosynthesis and photosystem II function in vivo. Iron deficiency induced a drastic decrease in net photosynthesis (Pn). Compared with normal plants, the maximal quantum yield of PSII photochemistry (psipo) in iron-deficient plants was only slightly lower; whereas, the efficiency with which a trapped exciton can move an electron into the electron transport chain further than QA-(Psio) and quantum yield of electron transport beyond QA (psiEo) were significantly depressed. Iron deficiency also caused a clear enhancement of the relative variable fluorescence at K step (VK). When exposed to light, iron-deficient plants had considerably lower efficiency of excitation energy capture by open PSII reaction centers (Fv'/Fm'), quantum yield of PSII electron transport (PhiPSII), and photochemical quenching coefficient (qP), but markedly higher non-photochemical quenching (NPQ). In addition, post-illumination transient increase in chlorophyll fluorescence was clearly enhanced in iron-deficient plants. Basing on these data, we suggest that both the donor and the acceptor sides of PSII complex were damaged by iron deficiency; cyclic electron transport around PSI in iron-deficient soybean plants might play an important role in inducing the excitation energy dissipation and meeting the demand for extra ATP as a compensation for the loss of phosphorylation capability.
Plant seed oil is the major source of many fatty acids for human nutrition, and also one of industrial feedstocks. Recent advances in understanding of the basic biochemistry of seed oil biosynthesis, coupled with cloning of the genes encoding the enzymes involved in fatty acid modification and oil accumulation, have set the stage for the metabolic engineering of oilseed crops that produce "designer" plant seed oils with the improved nutritional values for human being. In this review we provide an overview of seed oil biosynthesis/regulation and highlight the key enzymatic steps that are targets for gene manipulation. The strategies of metabolic engineering of fatty acids in oilseeds, including overexpression or suppression of genes encoding single or multi-step biosynthetic pathways and assembling the complete pathway for the synthesis of long-chain polyunsaturated fatty acids (e.g. arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid) are described in detail. The current "bottlenecks" in using common oilseeds as "bioreactors" for commercial production of high-value fatty acids are analyzed. It is also discussed that the future research focuses of oilseed metabolic engineering and the prospects in creating renewable sources and promoting the sustainable development of human society and economy.
H(2)O(2), plasma membrane H(+)-ATPase (PM H(+)-ATPase) and salicylic acid (SA) play important roles in sensing external stimulation and activating defense responses in plants. However, it remains uncertain whether they are involved and interrelated in response to heat acclimation. Experiments were performed by pharmacological methods, and the relationship and the connection between endogenous H(2)O(2), free SA and PM H(+)-ATPase were investigated in pea plants (Pisum sativum L.) during heat acclimation. The results showed that an accumulation peaks of H(2)O(2), free SA and PM H(+)-ATPase, were detected during heat acclimation at 37 degrees C for 2 h and H(2)O(2) burst appeared before SA accumulation that followed by increase of PM H(+)-ATPase activity (Fig.1). Pretreatments with either scavengers of active oxygen species (dimethyl sulfoxide and ascorbic acid) or antioxidant (reduced glutathione) inhibited the increases in both H(2)O(2) and free SA contents as a part of heat acclimation (Fig.2). Additionally, changes in activity of plasma membrane NADPH oxidase paralleled with H(2)O(2) level during heat acclimation (Figs.1 and 3), implicating that H(2)O(2) might be generated by plasma membrane NADPH oxidase. Moreover, pretreatments with either diphenylene iodonium (DPI), a suicide substrate inhibitor of plasma membrane NADPH oxidase, or dimethylthiourea (DMTU), a quencher of H(2)O(2), could block the increase in free SA content and activity of plasma membrane NADPH oxidase as a part of heat acclimation (Fig.4). According to the assay described above, it is suggested that both H(2)O(2) and PM H(+)-ATPase participate in SA signaling that leads to the development of thermotolerance in pea plant, and H(2)O(2) functions upstream and PM H(+)-ATPase functions downstream of the SA signal. Also, the regulation mechanism of PM H(+)-ATPase activity was investigated, which showed that during heat acclimation, increase of PM H(+)-ATPase activity was independent of PM H(+)-ATPase amount and the enzyme activity may be modulated at post-translational level that may involve in reversible protein phosphorylation (Fig.5).
L-arginine is an important and unique amino acid in plants. It serves not only as an important nitrogen reserve and recycling, but also as a precursor of the biosynthesis of polyamines, nitric oxide and so on. Polyamines and nitric oxide are important messengers involved in almost all physiological and biochemical processes, growth & development, and adaptation of plants to stress. Arginine decarboxylase, arginase and nitric oxide synthase are the key enzymes in L-arginine catabolism, in which polyamines are formed through ADC or arginase-ODC pathway while nitric oxide is formed through the NOS pathway. The relative activity of these three enzymes can control the direction of arginine metabolism. Arginine content keeps higher level in roots during overwinter period. The arginine metabolism plays important role in perception and adaptation of plant to environmental disturbances.
When measured at a same CO(2) concentration, net photosynthetic rate is often significantly lower in long-term high CO(2)-grown plants than the ambient CO(2)-grown ones. This phenomenon is termed photosynthetic acclimation or down-regulation. Although there have been many reports and reviews, the mechanism(s) of the photosynthetic acclimation is not very clear. Combining the work of the authors' group, this paper briefly reviews the progress in studies on the mechanism(s) of the photosynthetic acclimation to elevated CO(2). It is suggested that besides the possible effects of respiration enhancement and excessive photosynthate accumulation, RuBP carboxylation limitation and RuBP regeneration limitation are probably the main factors leading to the photosynthetic acclimation.
Gamma-tubulin gene is a very important house-keeping gene and plays key roles in cell division, microtubule nucleation and cell cycle regulation in both plants and animals. In this paper, we studied the gamma-tubulin gene silencing in Nicotiana tabacum var. Samsun NN using the recombinant PVX vector with D fragment of gamma-tubulin cDNA. Special phenotype appeared from recombinant PVX-vector-infected plants, the different morphological leaves grew alternatively in leaf layers (Figs.3-5), and all the buds fell off gradually (Fig.4), resulting in failure of all the microspores to develop to the tetrad stage (Fig.8). The silencing initiated from the apical tissue and kept on until it was mature (Figs.3, 4). There was the increasing of target gene mRNA obviously during gene silencing, even to exceed the positive control (Fig.6). The PVX vector was also suppressed strongly and could fluctuate to some degree (Fig.9), which was probably correlated with target gene mRNA expression.
Microprojectile bombardment-mediated transformation method has been developed for onion (Allium cepa L.) using embryogenic calli, induced from stem discs, as target tissue. Zinc-finger protein gene OSISAP1 (Oryza sative subspecies indica stress-associated protein gene) was introduced into the open-pollinated onion cultivar (subs.) 'HG400B'. Bombardment parameters were optimized as: the pressure is 1,100 psi, the distance is 6 cm, two times, the ratio of mass between plasmid DNA and golden particles is 1:320. An efficient microprojectile bombardment-mediated transformation system of onion (Allium cepa L.) callus has been established. The binary vector used carried the nptII gene for kanamycin resistance and the GUS reporter gene. Transgenic cultures were screened for their ability to express the GUS reporter gene and to grow in the presence of kanamycin (150 mg/L). Transient expression of GUS reporter gene was observed through histochemical staining of embryogenic callus transformed by microprojectile bombardment. The putative transgenic plants were analysed at the molecular level using PCR, southern hybridization, and RT-PCR. The results confirmed that the OSISAP1 gene was integrated as one copy into the genome of onion and expression. Transgenic plants were produced efficiently with a transformation frequency of about 10%. Test of salinity-alkali stress showed that sodium chloride and sodium bicarbonate at 200 mmol/L effectively killed non-transgenic plants within 1 week of irrigation, while the transgenic plants were completely unaffected by salinity of 400 mmol/L. So transformation with the OSISAP1 gene raised the salinity-alkali-tolerance of the transgenic plants to a high level.
Some desiccation-tolerant plants can survive the loss of water even when the water content becomes as low as 0.3 g H2O/g dry weight, and can still repair quickly their cellular structures and function damages by desiccation when they get access to enough moisture again. While moderate dehydration, the tissue could accumulate some special proteins, small molecules, carbohydrate etc. which can protect the original structure and function of macromolecule and membrane through themselves' natural characteristics. Mean-while, water loss results in increased activities of reactive-oxygen-scavenging enzymes (SOD, CAT, APX and GR). For the acquisition of desiccation tolerance, water in the plants or seeds must be induced to enter the glassy state, a change which can be induced by many compounds. The crucial protective means also include the increased formation of endogenous antioxidants and the partitioning of amphiphilic substances in the lipid phase to protect the membrane. The resurrection plants and seeds are good materials for desiccation tolerance research.
To use different types of promoters in transgenic rice research, the 1.1 kb 5'-upstream regulation region of one of the tomato (Solanum tuberosum L.) Rubisco small subunit gene, rbcS3A, was cloned and its sequences were confirmed by comparison with the known sequences in GenBank. The cloned rbcS3A promoter was fused to the 5'-upstream of GUS (beta-glucuronidase) coding region in a binary vector, and introduced into an elite japonica rice variety by Agrogacterium-mediated transformation. The integration of the GUS fusion gene into the genome of transgenic rice was confirmed by both PCR and Southern blot analysis. The results of both histochemical staining and quantitative analysis of GUS activity showed that the expression level of GUS fusion gene was significantly stronger in stem, leaf blade and sheath than in other organs of transgenic rice plants, and showed highest in the stem, which implies that the tomato rbcS3A promoter can make tissue-specific, in particular in the stem, expression of foreign genes in transgenic rice. The results present here also demonstrate that light induction had no effect on the expression of the foreign gene when regulated by the tomato rbcS3A promoter in transgenic rice. Our results show that the cloned tomato rbcS3A promoter might be very useful for the expression of target genes in transgenic rice, with particularly high efficiency in stem tissues.
Selective microelectrode technique, known as an electrophysiological approach, can be used to measure directly specific information on ion or molecule distribution and movement both inside and outside of living organelle, biological cells, tissue and organs. It has several advantages over other methods in measuring ionic or molecular information, e.g. easy to handle, fast response, high sensitivity (10(-12) moles cm(-2) s(-1)) and non-invasive to the samples in addition to continuous measurement and automatic monitoring. Microscopic-scale selective electrode (with a tip diameter of 0.5-5 microm) can be used to measure net fluxes of ions or molecules outside of growing biological cells, tissues and organs, to measure activities of ions or molecules inside of growing organelle and biological cells. Thus, it has many applications in various research fields. The technical principle of design and use of selective microelectrode and its progress and development prospect in plant physiological research are summarized.
Poplar is among the most important deciduous tree species in plantations over the world and has been used as an important model system for molecular genetics of trees. The analysis of codon usage may improve the understanding of the mechanism of codon distribution and variation in poplar and the understanding of factors shaping the codon usage patterns. Here, an EN(c) (effective number of codons)-plot method and multivariate statistical method called correspondence analysis (COA) were used to examine the codon usage of 314 genes of poplar. The results show that the main trend was highly negative correlated with the gene expression level assessed by the ''Codon Adaptation Index'' value. Moreover, there were two significant correlations between axis 1 coordinates and GC3(s) content and gene length, we infer that gene nucleotide composition and gene length also play an important role in shaping the codon usage bias in poplar. The result of relative synonymous codon usage (RSCU) analysis shows a high bias of codon usage toward the codon with A or T ending. In addition, we compared the codon preferences among poplar, Arabidopsis thaliana, Oryza sative, Homo sapiens and Escherichia coli, and poplar was found to be most similar to A.thaliana and least similar to E.coli. In this paper, 10 codons defined firstly as optimal codons through an analysis of the high-expression codon in poplar may provide some useful information for genetic engineering of poplar.
The relationships between the rates of starch synthesis and the activities of enzymes responsible for starch biosynthesis in developing grains of normal, pop, sweet and waxy corns were investigated and compared throughout the grain filling period. The results indicated that the rates of starch synthesis and the activities of sucrose synthase (SS), soluble starch synthase (SSS), granule-bound starch synthase (GBSS), starch-branching enzyme (SBE) and starch-debranching enzyme (DBE) each exhibited a single peak during grain filling period. Normal corn showed significantly higher SS activity than other genotypes between 30 and 40 DAP. The mean and maximum activities of SSS were in the following order: normal corn>waxy corn>pop corn>sweet corn. GBSS activities were significantly higher in normal corn, and significantly lower in waxy corn at late filling period. SBE activity of waxy corn was significantly higher than other lines after 10 DAP. DBE activity of sweet corn was extremely low and completely lost at 40 DAP. The rates of starch synthesis had some correlation with the activities of SS, SSS, GBSS and SBE during the grain filling process. No correlation was found between the rates of starch synthesis and the activities of ADP-glucose pyrophosphorylase (AGPase) and DBE. SS activity appears to play a major role in starch biosynthesis in maize. GBSS is responsible for amylose synthesis especially in the later period. SSS and SBE are associated with amylopectin biosynthesis.
A full-length 1,043-base-pair cDNA clone encoding a chloroplast copper/zinc superoxide dismutase (Cu/Zn-SOD) of upland cotton was first isolated by rapid amplification of cDNA ends (RACE) from the leaves of the cotton (Gossypium hirsutum L.) variety 'CRI36'. Nucleotide sequence analysis of the clone revealed that it contained the complete coding sequence of the mature SOD isozyme subunit, along with a 60-amino acid transit peptide at N-terminal. The amino acid sequence predicted from the full-length clone showed 66%-74% homology with the amino acid sequences of Cu/Zn-SOD from several other plants. This gene was found to be expressed in the leaves and stems, but not in roots, flowers, and hypocotyls, indicating that the gene was expressed only in green tissues. Also, its expression was found to be most active at seedling stage and declined gradually in later development stages. Expression of this cotton Cu/Zn-SOD gene by using the pET-21a (+) expression vector in E. coli BL21 (DE3) led to the production of a novel 29 kD polypeptide with SOD enzyme activity, confirming that the cloned cotton Cu/Zn-SOD cDNA was indeed encoding a functioning SOD enzyme.
研究了不同施氮量对高蛋白含量植物杂交酸模（Rumex patientiaxR.tianschanicus）叶片中总光合电子流和分配在碳同化、光呼吸、Mehler反应以及氮代谢上的光合电子流的影响,并研究了不同施氮量对硝酸还原酶（NR）和谷氨酰胺合成酶（GS）的活性、叶片的蛋白质含量及叶绿素含量的影响。结果表明随着施氮量的增加,硝酸还原酶和谷氨酰胺合成酶的活性都显著提高,同时更多的光合电子流分配到氮代谢和光呼吸。氮代谢所需光合电子流约占总光合电子流的15%～21%。缺氮并没有造成光合电子流向Mehler反应分配的增加。
Vacuolated mesophyll protoplasts of Nicotiana rustica L.were electrically fused with evacuolated protoplasts of the same genus(N. tabacum cv.‘Gexin No.1’)during a 7-day space flight in the Chinese spacecraftSZ-4.The initial cell division leading to micro-callus formation took place after landing(Fig.1).Higher plating efficien- cies were observed in the flight samples than the control culture,but the frequency of plantlets re- generation reduced by about 20% of the control (Table 1).The hybrid characters were tested by chromosome counting,isozyme analysis and com- parison of morphological characteristics(Figs.2- 4).About 32% of the regenerates showed hybrid character.Leaf morphological modifications were found in 3 hybrids,i.e.,H23,H25 and H27.After backcrossing with N.rustica,alterations in flower color and leaf shape occurred in the somatic hy- brid H23(Fig.5).These results demonstrate that the hybrids formed under microgravity condition could regenerate fertile plants.
Arabinose is one of the most dynamic cell wall glycosyl residues released during fruit ripening, alpha-L-arabinofuranosidase (alpha-Arab) are major glycosidases that may remove arabinose units from fruit cell wall polysaccharides. To find out whether alpha-Arab plays important roles in banana fruit softening, the enzyme activities in peel and pulp, fruit firmness, respiration rate and ethylene release rate were assayed during banana softening. The results showed that alpha-Arab activities in banana pulp and peel increased slightly at the beginning of storage and reached their maxima when the fruit firmness decreased drastically, alpha-Arab activity increased by more than ten folds in both pulp and peel during ripening and alpha-Arab activities were higher in pulp than in peel. Treatment of banana fruits with ethylene absorbent postponed the time of reaching of its maxima of respiration and ethylene, enhanced the firmness of pup and decreased alpha-Arab activity in the peel and pulp. These results suggest that alpha-Arab induced the decrease of fruit firmness and played an important role in banana fruit softening, and its activity was regulated by ethylene.
Wild banana species are essential natural gene pools for banana improvement. In this study, six RGAs about 500 bp were obtained from leaves of Musa acuminata, a wild banana shown to be resistant to banana Fusarium wilt race 4, by PCR amplification with degenerate primers designed according to the conserved NBS motif and serine/threonine kinase domain of plant resistance (R) genes. Among these RGAs, the deduced amino acids of WNB1 and WNB2 contain NB-ARC domain and WNB1 can be translated into polypeptide uninterrupted by stop codons. The deduced amino acids of other four RGAs (WST1, WST2, WST3 and WST4) all contain the serine/threonine kinase domain and WST3 encodes a polypeptide homologous to that of bacterial blight resistance gene Xa21 of rice. At different time after inoculation with Fusarium oxysporum f. sp. cubense (FOC) race 4, the transcript patterns of WNB1 and WST3 was enhanced, which implied that the expression of WNB1 and WST3 may be related to the resistance of banana to Fusarium wilt.
With electron spin resonance and chemical assay, the production of superoxide anion in the Xanthomonas oryzae pv. oryzae strain OS-14 suspension is extracellularly localized. It was found that superoxide anion produced in the filtrate. It was confirmed that peptide or enzyme did not contribute to the production of superoxide anion in the filtrate of OS-14. Organic acids were shown to be the primary source of superoxide anion produced in the filtrate of OS-14. These results indicated that non-enzymic molecules might be another source of superoxide anion produced by Xanthomonas oryzae pv. oryzae, in addition to enzymes, which suggested the multiform generation of superoxide anion.
