LHC II protein phosphorylation in leaves of Arabidopsis thaliana mutants deficient in non-photochemical quenching

Phosphorylation of the light-harvesting chlorophyll a/b complex II (LHC II) proteins is induced in light via activation of the LHC II kinase by reduction of cytochrome b₆f complex in thylakoid membranes. We have recently shown that, besides this activation, the LHC II kinase can be regulated in vitro by a thioredoxin-like component, and H₂O₂ that inserts an inhibitory loop in the regulation of LHC II protein phosphorylation in the chloroplast. In order to disclose the complex network for LHC II protein phosphorylation in vivo, we studied phosphorylation of LHC II proteins in the leaves of npq1-2 and npq4-1 mutants of Arabidopis thaliana. In comparison to wild-type, these mutants showed reduced non-photochemical quenching and increased excitation pressure of Photosystem II (PS II) under physiological light intensities. Peculiar regulation of LHC II protein phosphorylation was observed in mutant leaves under illumination. The npq4-1 mutant was able to maintain a high amount of phosphorylated LHC II proteins in thylakoid membranes at light intensities that induced inhibition of phosphorylation in wild-type leaves. Light intensity-dependent changes in the level of LHC II protein phosphorylation were smaller in the npq1-2 mutant compared to the wild-type. No significant differences in leaf thickness, dry weight, chlorophyll content, or the amount of LHC II proteins were observed between the two mutant and wild-type lines. We propose that the reduced capacity of the mutant lines to dissipate excess excitation energy induces changes in the production of reactive oxygen species in chloroplasts, which consequently affects the regulation of LHC II protein phosphorylation.     

Development of an ELISA for estimation of the copper nutritional status of wheat and cotton

Studies were carried out with hydroponically grown wheat and cotton to develop the Cu-requiring protein phenolase as a biomarker of Cu nutrient status. Isozymes of phenolase whose levels were reduced by Cu deficiency were identified by Western blots. A competitive enzyme-linked, immunosorbent assay (ELISA) was developed that could detect as little as 25 ng of phenolase. The ELISA revealed that Cu-sufficient cotton leaves had about 4-fold more phenolase antigen than did Cu-sufficient wheat leaves. In both species, the level of phenolase was reduced by 2- to 5-fold in leaves of Cu-deficient plants. Because the immunoassay for phenolase protein is rapid, inexpensive, and can be carried out with small amounts of leaf material, it has potential as a tool for assessment of the Cu status of crop plants.Abbreviations ELISA enzyme-linked immunosorbent assay - HRP horseradish peroxidase - TBS Trisbuffered saline (20 mM Tris-HCl, pH 9.5, 150 mM NaCl) - TBST Tris-buffered saline containing 0.05% Tween-20     

Monoclonal antibodies to a higher-plant nitrate reductase: Differential inhibition of enzyme activities

A set of monoclonal antibodies has been raised against NADH-nitrate reductase (NR; EC 1.6.6.1) from spinach (Spinacea oleracea L.) leaves. Antibodies were screened by enzyme-linked immunosorbent assay and by their ability to inhibit various activities of the enzyme. The six monoclonals selected (AFRC MAC 74 to 79) are all gamma globulins; four (MAC 74 to 77) inhibit all terminal donating activities (NADH-NR; flavin mononucleotide, reduced form (FMNH₂)-NR; and methyl viologen, reduced form (MV)-NR) and two (MAC 78 and 79) inhibit the acceptor activities (NADH-NR, and NADH-cytochrome c reductase). MAC 74 to 77 inhibit the NADH-NR activity of crude extracts of a variety of species (mono- and dicotyledoneae) while MAC 78 and 79 are effective against spinach and marrow, but not oil-seed rape, cucumber, oats, wheat and barley.Abbreviations Cyt c Rase cytochrome c reductase - ELISA enzyme-linked immunosorbent assay - FAD(H₂) flavin adenine dinucleotide (reduced form) - FMN(H₂) flavin mononucleotide (reduced form) - McAb monoclonal antibody - MV methyl viologen reduced form - NR nitrate reductase     

Leaf senescence in a non-yellowing mutant of Festuca pratensis: Proteins of photosystem II

The senescence of leaves is characterized by yellowing as chlorophyll pigments are degraded. Proteins of the chloroplasts also decline during this phase of development. There exists a non-yellowing mutant genotype of Festuca pratensis Huds. which does not suffer a loss of chlorophyll during senescence. The fate of chloroplast membrane proteins was studied in mutant and wild-type plants by immune blotting and immuno-electron microscopy. Intrinsic proteins of photosystem II, exemplified by the light-harvesting chlorophyll a/b-binding protein (LHCP-2) and D1, were shown to be unusually stable in the mutant during senescence, whereas the extrinsic 33-kilodalton protein of the oxygen-evolving complex was equally lable in both genotypes. An ultrastructural study revealed that while the intrinsic proteins remained in the internal membranes of the chloroplasts, they ceased to display the heterogenous lateral distribution within the lamellae which was characteristic of nonsenescent chloroplasts. These observations are discussed in the light of possible mechanisms of protein turnover in chloroplasts.Abbreviations kDa kilodalton - LHCP-2 light-harvesting chlorophyll a/b-binding protein - M_r relative molecular mass - PSII photosystem II - SDS sodium dodecyl sulphate     

Immunochemical detection of ochratoxin A in black Aspergillus strains

One hundred and fifty-seven strains belonging to Aspergillus section Nigri were tested for ochratoxin A production using three different methods: a relatively new immunochemical method based on an enzyme-linked immunosorbent assay (ELISA), thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). The monoclonal antibody-based ELISA technique was successfully used to screen for low levels of ochratoxin A in the black Aspergilli without concentrating the culture filtrates. The results were confirmed by TLC and HPLC analysis and chemical derivatization. These latter methods required concentrated filtrates. Ochratoxin A was detected in the culture filtrates of five of the 12 A. carbonarius strains, none of the 45 A.japonicus strains and three of the 100 isolates in the A. niger aggregate (A. foetidus, A. awamori and A. niger).Abbreviations ELISA enzyme-linked immunosorbent assay - HPLC high-performance liquid chromatography - OA ochratoxin A - TLC thin-layer chromatography     

Biochemical differentiation in the tobacco flower probed with monoclonal antibodies

We have isolated a series of monoclonal antibodies that react to antigens in flowers of Nicotiana tabacum L. (tobacco) displaying specificity or preferentiality in their cell and tissue distributions. We immunized mice with extracts from tobacco flowers and then screened the hybridomas by enzyme-linked immunosorbent assay (ELISA) against extracts from leaves, sepals, petals, stamens and pistils; twenty five were chosen from the total screened. The antigens detected by about half of the antibodies were periodate-sensitive, implying that the epitopes were carbohydrate. Competition ELISA assays were used to determine if any antibodies were reacting to the same epitopes. Western blot analysis showed that while some antibodies reacted to specific bands, the bulk either failed to react or reacted to multiple bands, consistent with a glyco-conjugate nature for many of the antigens. Analysis of the spatial pattern of antigen distribution within tobacco flowers by immunolocalization showed that some antibodies recognized epitopes that were limited to very specific cells and tissues. We used the immunolocalization technique to analyze a mutant with stigmoid anthers: an antibody recognizing a pistil transmitting-tract antigen also reacted to cells in stigmoid anthers. Our results with this antibody set imply that biochemical differentiation within the tobacco flower includes cell-and tissue-specific glyco-moeities, and also that similarities, at the biochemical level, exist between a normal floral organ and the abnormal organ in a phenotype with a developmental switch.Abbreviations ELISA enzyme-linked immunosorbent assay - Fg immunoglobulin - kDa kilodalton     

Monoclonal antibodies identify common and differentiation-specific antigens on the plasma membrane of guard cells of Vicia faba L.

Monoclonal antibodies were raised against the plasma membrane of Vicia faba L. guard cells by immunizing either with total membranes from purified guard-cell protoplasts or with sealed, predominantly right-side-out plasma-membrane vesicles prepared from abaxial epidermes of V. faba by aqueous two-phase partitioning. Hybridoma screening was performed by enzyme-linked immunosorbent assay using polystyrene-adsorbed plasma-membrane vesicles as solid phase and by indirect immunofluorescence analysis using unfixed, immobilized protoplasts in a microvolume Terasaki assay. A range of monoclonal antibodies was characterized and is reported here. One monoclonal antibody, G26-6-B2, is guard-cell-specific and does not react with mesophyll-cell protoplasts of the same species. It binds to a periodate-resistant but trypsin-labile epitope, probably a differentiation-specific plasma-membrane protein.Abbreviations ELISA enzyme-linked immunosorbent assay - FITC fluorescein isothiocyanate - GCP guard cell protoplast(s) - Ig immunoglobulin - MAB monoclonal antibody - MCP mesophyll-cell protoplast(s) - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis     

Studies on the regulation of 1-aminocyclopropane-1-carboxylate synthase in tomato using monoclonal antibodies

A partially purified preparation of 1-aminocyclopropane-1-carboxylate (ACC) synthase (EC 4.4.1.14) from tomato (Lycopersicon esculentum (Mill.) fruit tissue was used to generate monoclonal antibodies (MAb) specific for the two different MAbs yielded a 50-kDa polypeptide as shown by sodium dodecylsulfate-polyacrylamide gel electrophoresis. An enzyme-linked immunosorbent assay (ELISA) capable of detecting <1 ng of antigen was developed. The ELISA system was used to demonstrate that two of the MAbs recognized different epitopes on the ACC-synthase protein. Wound-induced increases in ACC-synthase activity in tomato fruit tissue were correlated with changes in ELISA-detectable protein. In-vivo labeling of wounded tissue with [³⁵S]methionine followed by extraction and immunopurification in the presence of various protease inhibitors yielded one major radioactive band of 50 kDa molecular mass. Pulse labeling with [³⁵S]methionine at various times after wounding indicated that the wound-induced increase in ACC-synthase activity involved de-novo synthesis of a rapidly turning over 50-kDa polypeptide.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - ELISA enzyme-linked immunosorbent assay - MAb monoclonal antibody - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis     

The redox state of the plastoquinone pool controls the level of the light-harvesting chlorophyll a/b binding protein complex II (LHC II) during photoacclimation

A cytochrome b ₆ f deficient mutant of Lemna perpusilla maintains a constant and lower level of the light-harvesting chl a/b-binding protein complex II (LHC II) as compared to the wild type plants at low-light intensities. Inhibition of the plastoquinone pool reduction increases the LHC II content of the mutant at both low- and high-light intensities but only at high-light intensity in the wild type plants. Proteolytic activity against LHC II appears during high-light photoacclimation of wild type plants. However, the acclimative protease is present in the mutant at both light intensities. These and additional results suggest that the plastoquinone redox state serves as the major signal-transducing component in the photoacclimation process affecting both, synthesis and degradation of LHC II and appearance of acclimative LHC II proteolysis. The plastoquinol pool cannot be oxidized by linear electron flow in the mutant plants which are locked in a ‘high light’ acclimation state. The cytochrome b ₆ f complex may be involved indirectly in the regulation of photoacclimation via 1) regulation of the plastoquinone redox state; 2) regulation of the redox-controlled thylakoid protein kinase allowing exposure of the dephosphorylated LHC II to acclimative proteolysis. This revised version was published online in June 2006 with corrections to the Cover Date.     

Molecular differentiation in pea powdery-mildew haustoria

Monoclonal antibodies have been raised against haustorial complexes isolated from pea (Pisum sativum L.) leaves infected by the biotrophic powdery mildew fungus Erysiphe pisi D.C. Immuno-localisation studies, using isolated haustorial complexes and infected pea leaf material, have shown that one of the antibodies, designated UB7, binds to fungal wall and plasma membranes present in both haustoria and mycelia. However, a second antibody, UB8, binds specifically to the haustorial plasma membrane, and does not label fungal plasma membranes in mycelia. Western blotting and antigen-modification techniques have shown that UB8 recognises a protein epitope of a 62-kDa antigen. A reduction in molecular weight of this component after endo-F treatment indicates that the antigen is an N-linked glycoprotein. UB7 also recognises a 62-kDa glycoprotein, which is susceptible to endo-F treatment, and the antibody binds to a carbohydrate epitope. Differences in molecular weights of the products after endo-F treatment of antigens show that the 62-kDa glycoproteins recognised by the antibodies are distinct molecules, in accordance with the localisation results. Overall, the results provide evidence for molecular differentiation associated with the development of haustoria in a biotrophic infection.Abbreviations ehm extrahaustorial membrane - ELISA enzyme-linked immunosorbent assay - HC haustorial complex - hpm haustorial plasma membrane - IIF indirect immunofluorescence - MAb monoclonal antibody - M_r apparent molecular weight - PMSF phenylmethylsulfonyl fluoride - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis We thank Mr. D. Mills and Mr. P. Stanley for help with the EM immunogold techniques. This work was supported by an Agricultural and Food Research Council grant and a studentship from the Science and Engineering Research Council.     

Cytochrome b 6 f complex: Dynamic molecular organization,function and acclimation

The cytochrome b ₆ f complex occupies a central position in photosynthetic electron transport and proton translocation by linking PS II to PS I in linear electron flow from water to NADP⁺, and around PS I for cyclic electron flow. Cytochrome b ₆ f complexes are uniquely located in three membrane domains: the appressed granal membranes, the non-appressed stroma thylakoids and end grana membranes, and also the non-appressed grana margins, in contrast to the marked lateral heterogeneity of the localization of all other thylakoid multiprotein complexes. In addition to its vital role in vectorial electron transfer and proton translocation across the membrane, cytochrome b ₆ f complex is also involved in the regulation of balanced light excitation energy distribution between the photosystems, since its redox state governs the activation of LHC II kinase (the kinase that phosphorylates the mobile peripheral fraction of the chlorophyll a/b-proteins of LHC II of PS II). Hence, cytochrome b ₆ f complex is the molecular link in the interactive co-regulation of light-harvesting and electron transfer.The importance of a highly dynamic, yet flexible organization of the thylakoid membranes of plants and green algae has been highlighted by the exciting discovery that a lateral reorganization of some cytochrome b ₆ f complexes occurs in the state transition mechanism both in vivo and in vitro (Vallon et al. 1991). The lateral redistribution of phosphorylated LHC II from stacked granal membrane regions is accompanied by a concomitant movement of some cytochrome b ₆ f complexes from the granal membranes out to the PS I-containing stroma thylakoids. Thus, the dynamic movement of cytochrome b ₆ f complex as a multiprotein complex is a molecular mechanism for short-term adaptation to changing light conditions. With the concept of different membrane domains for linear and cyclic electron flow gaining credence, it is thought that linear electron flow occurs in the granal compartments and cyclic electron flow is localised in the stroma thylakoids at non-limiting irradiances. It is postulated that dynamic lateral reversible redistribution of some cytochrome b ₆ f complexes are part of the molecular mechanism involved in the regulation of linear electron transfer (ATP and NADPH) and cyclic electron flow (ATP only). Finally, the molecular significance of the marked regulation of cytochrome b ₆ f complexes for long-term regulation and optimization of photosynthetic function under varying environmental conditions, particularly light acclimation, is discussed.Abbreviations Chl chlorophyll - cyt cytochrome - PS Photosystem     

Immunoassay and ultrastructural localization of isopentenyladenine and related cytokinins using monoclonal antibodies

Two hybridoma cell lines, J40-IV-A1 and J40-IV-C4 were obtained from a fusion of spleen cells of Balb/c mice immunized against an isopentenyladenosine-bovine serum albumin conjugate with X63. Ag 8.653 myeloma cells. These hybrids secrete monoclonal antibodies of the immunoglobulin G (IgG) class and share high affinities and specificities to isopentenyladenine and isopentenyladenosine suitable for the detection of femtomole amounts of these cytokinins in plant extracts by enzyme-linked immunosorbent assay (ELISA). One of the monoclonal antibodies (J40-IV-C4) has been employed to localize isopentenyladenine immunoreactivity in a cytokinin-over-producing mutant of the moss, Physcomitrella patens. After fixation and embedding at low temperature, immunoreactivity was visualized in protonemal filaments of the moss mutant by the use of indirect immunogold labelling. In the mutant, the labelling was predominantly in the wall of the protonemal cells. Neither the wild-type nor control treatments showed any labelling. The signficance of these observations is discussed with respect to the applicability of immunocytochemical techniques for the localization of low-molecular-weight compounds in plant tissue.Abbreviations ELISA enzyme linked immunosorbent assay - HPLC high-performance liquid chromatography - IP isopentenyladenine - IPA isopentenyladenosine - mAB monoclonal antibody - OVE cytokinin-over-producing mutant - RIA radioimmunoassay     

Subcellular distribution of arabinogalactan proteins in pollen grains and tubes as revealed with a monoclonal antibody raised against stylar arabinogalactan proteins

Summary Monoclonal antibody PCBC3, raised against stylar extracts fromNicotians, alata flowers, was deduced from enzyme-linked immunosorbent assays and inhibition of immuno-gold labelling on tissue sections to bind specifically to carbohydrate epitopes on arabinogalactan proteins (AGPs) but not to other arabinose-containing cell wall polysaccharides. When pollen grains ofN. tabacum were hydrated in fixative, PCBC3 bound to vesicles in the vicinity of the endoplasmic reticulum but, when grains were hydrated for 20 min in culture medium before fixation, binding was restricted to the plasma membrane. The generative-cell plasma membrane was also labelled in grains ofLycopersicon peruvianum. In pollen tubes ofN. tabacum grown in liquid culture, the AGPs detected by PCBC3 were located in several regions, including the plasma membrane, tubular-vesicular structures (plasmalemmasomes) at and under the plasma membrane, and multilamellar bodies within vacuoles, features generally associated with endocytosis. Labelling was not evident in secretory vesicles or the plasma membrane at the pollen-tube tip. The AGPs detected with PCBC3 were also present in pollen-tube walls, near the interface between the inner, callosic layer and the outer, fibrillar, pectic layer. Pollen tubes ofN. tabacum grown in medium lacking added CuSO₄ produce a wall with an abnormally thickened fibrillar layer, and this layer was uniformly labelled with PCBC3. The disposition of wall AGPs thus changes in pollen tubes of different morphologies.Abbreviations AGP arabinogalactan protein - -L-Araf -L-arabinofuranose - ELISA enzyme-linked immunosorbent assay - MAb monoclonal antibody - PBS phosphate-buffered saline     

Discrimination between the red- and far-red-absorbing forms of phytochrome from Avena sativa L. by monoclonal antibodies

A set of rat monoclonal antibodies (ARC MAC 48 to 52 and 54 to 56), raised to phytochrome from dark-grown seedlings of Avena sativa L. was tested for the ability to discriminate between the red-absorbing (Pr) and far-red-absorbing (Pfr) forms of phytochrome by indirect enzyme-linked immunosorbent assay. MAC 50 bound more strongly to Pfr and MAC 49 and 52 showed preferential binding to Pr from extracts of dark-grown Avena seedlings; MAC 50 also bound more strongly to Pfr from brushite-purified phytochrome. The remainder of the monoclonal antibodies and a rabbit polyclonal antiphytochrome preparation did not discriminate between Pr and Pfr. The results provide evidence for conformational changes in defined regions of the phytochrome apoprotein upon photoconversion.Abbreviations ELISA enzyme-linked immunosorbent assay - FR far-red light - McAb monoclonal antibody(ies) - PBS phosphate-buffered saline - Pfr far-red-absorbing form of phytochrome - Pr red-absorbing form of phytochrome - R red light - PMSF phenylmethylsulphonylfluoride     

Characterization of rapeseed myrosinase-binding protein

Myrosinase-binding proteins (MBPs) were purified from seeds of Brassica napus L. (oilseed rape). The proteins were characterized with respect to amino-acid composition, peptide sequence and isoelectric points. Gel electrophoresis and Western blotting of protein extracts from mature seeds showed the existence of at least ten proteins reacting with a monoclonal anti-MBP antibody and ranging in molecular size from 110 to 30 kDa. Proteins other than MBP reacting with the anti-MBP antibody were assigned as myrosinase-binding protein-related proteins (MBPRPs). Two MBPRPs were purified by immunoaffinity chromatography and characterized with respect to partial amino-acid sequence. Sequence identities were found between MBP and MBPRP. Western blot analysis of protein extracts from different tissues of B. napus showed that MBPRP is present in the whole plant, whereas MBP mostly occurs in the mature seed. A double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) was used to investigate the occurrence of MBP and MBPRP in developing seeds of some species in the Brassicaceae family.Abbreviations FPLC fast protein liquid chromatography - MBP myrosinase-binding protein - MBPRP myrosinase-bindingprotein-telated protein - PBS phosphate-buffered saline     

Mannan and D-arabinitol concentrations in serum from a patient with Candida albicans endocarditis

In an attempt to clarify the comparative values of serological and microbiological examinations for the early diagnosis of systemic candidiasis, antibodies against Candida albicans, serum mannan, and the D-arabinitol creatinine ratio were investigated in a patient with aortic valve endocarditis associated with carcinoma of the bile duct. Candida precipitins and the antibody titer against Candida cell wall mannan were examined by an immunodiffusion technique and hemagglutination test, respectively. Serum mannan was tested by enzyme-linked immunosorbent assay (ELISA) using the biotin-streptavidin procedure. The upper limit of negativity of the assay was determined by adding 0.06 to the absorbance of pooled serum from healthy laboratory workers. This value ws about 0.8 ng/ml with ELISA. The D-arabinitol concentration in serum was examined by an enzymatic fluorometric method. Rising antibody titers against C. albicans, mannan antigenemia, and an elevated D-arabinitol creatinine ratio were first observed between the 11th and 12th hospital days. Blood cultures obtained on 8th, 9th, and 11th hospital days grew C. albicans after 3 to 4 days of incubation. Of 11 serum samples, 5 were positive for mannan, whereas D-arabinitol creatinine ratio was positive in 7 of 9 samples. Blood cultures was the earliest evidence of Candida infections in our cases. However, because of saprophytic nature of Candida species, tests for antibodies, antigenemia, and the D-arabinitol creatinine ratio in combination with blood cultures are necessary to confirm systemic candidiasis at an early stage of infection.Abbreviations ELISA enzyme-linked immunosorbent assay     

Immunocytochemical localization of callose in the germinated pollen ofCamellia japonica

Summary A polyclonal antibody against -1,3-glucan, callose, extracted from the pollen tube wall ofCamellia japonica was raised in mice and, using it as a probe, the localization of callose in the germinated pollen was studied. By confocal laser scanning microscopy, callose was found in the tip region of the pollen tube and the tube wall; the immuno-fluorescence in the tube wall was less toward the base of the tube. In contrast, the tip region did not fluoresce although the whole of the tube wall did strongly with aniline blue, the specific dye for callose. Immuno-electron microscopy showed that callose was also found in Golgi vesicles which concentrated in the tip region of the pollen tube, the inner layer of the tube wall, callose plugs, and Golgi vesicles in the pollen grain. Immuno-gold labeling was often detected on the fibrous structures in Golgi vesicles and callose plugs. Based on these results, the participation of Golgi vesicles in the formation of the tube wall and callose plugs was discussed.Abbreviation TBS Tris-buffered saline - Tris Tris(hydroxy-methyl)-aminomethane - PBS phosphate-buffered saline - BSA bovine serum albumin - ELISA enzyme-linked immunosorbent assay - CLSM confocal laser scanning microscopy - DP degree of polymerization     

Photosystem II in a mutant of Chlamydomonas reinhardtii lacking the 23 kDa psbP protein shows increased sensitivity to photoinhibition in the absence of chloride

The psbP gene product, the so called 23 kDa extrinsic protein, is involved in water oxidation carried out by Photosystem II. However, the protein is not absolutely required for water oxidation. Here we have studied Photosystem II mediated electron transfer in a mutant of Chlamydomonas reinhardtii, the FUD 39 mutant, that lacks the psbP protein. When grown in dim light the Photosystem II content in thylakoid membranes of FUD 39 is approximately similar to that in the wild-type. The oxygen evolution is dependent on the presence of chloride as a cofactor, which activates the water oxidation with a dissociation constant of about 4 mM. In the mutant, the oxygen evolution is very sensitive to photoinhibition when assayed at low chloride concentrations while chloride protects against photoinhibition with a dissociation constant of about 5 mM. The photoinhibition is irreversible as oxygen evolution cannot be restored by the addition of chloride to inhibited samples. In addition the inhibition seems to be targeted primarily to the Mn-cluster in Photosystem II as the electron transfer through the remaining part of Photosystem II is photoinhibited with slower kinetics. Thus, this mutant provides an experimental system in which effects of photoinhibition induced by lesions at the donor side of Photosystem II can be studied in vivo.Abbreviations Chl chlorophyll - DCIP 2,6-dichlorophenolindophenol - DPC 2,2-diphenylcarbonic dihydrazide - HEPES 4-(2-hydroxyethyl)-1-piperazinethanesulfonic acid - P₆₈₀ the primary electron donor to PS II - PpBQ phenyl-p-benzoquinone - PS II Photosystem II - Q_A the first quinone acceptor of PS II - Q_B the second quinone acceptor of PS II - SDS sodium dodecyl sulfate - Tris tris(hydroxymethyl)aminomethane - Tyr_D accessory electron donor on the D₂-protein - Tyr_Z tyrosine residue, acting as electron carrier between P₆₈₀ and the water oxidizing system     

肺炎克雷伯菌荚膜糖蛋白ELISA检测方法的研究

以肺炎克雷伯菌荚膜糖蛋白为免疫原,获得了兔抗肺炎克雷伯菌荚膜糖蛋白抗血清,通过硫酸铵分级沉淀与吸附法相结合的方法对抗血清进行纯化,并在此基础上建立了肺炎克雷伯菌荚膜糖蛋白间接ELISA检测方法.该检测方法的检测灵敏度为0.031 mg/L,线性检测范围为2.5～30.0 mg/L,批内误差为1.04％～3.22％,批间误差为2.61％～8.35％.     

Sid: a Mendelian locus controlling thylakoid membrane disassembly in senescing leaves of Festuca pratensis

Summary A spontaneous mutation arising in Festuca pratensis has the effect of stabilizing the pigmentproteolipid complexes of thylakoid membranes so that leaf tissue does not turn yellow during senescence. Inheritance of the non-yellowing character was analysed in crosses between the wild-type cultivar Rossa and a mutant line Bf 993. Electrophoretic variants of cytoplasmic phosphoglucoisomerase coded by alleles of the nuclear gene Pgi-2 were used to identify hybrids during intercrossing. About 96% of the F₁ progeny were heterozygous and all were phenotypically yellowing. In the F₂ generation yellow green segregated in a ratio of 2.141, not significantly different from 31. In the backcross between F₁ and Bf 993 the ratio was 11 yellow green. There was no indication of linkage to Pgi-2. Senescence of detached Bf 993 and Rossa leaves was compared with that of the F₁ hybrid. The hybrid behaved in an essentially identical fashion to the wildtype parent, and in marked contrast to the mutant, in all aspects of the senescence syndrome investigated, including loss of chlorophyll, carotenoids and the light-harvesting chlorophyll-protein of thylakoid membranes, and elevation of the particulate protein chlorophyll ratio in the terminal stages. It is concluded that there exists in Festuca pratensis a nuclear gene, designated Sid (senescence-induced degradation) which regulates turnover of hydrophobic components of photosynthetic membranes in ageing leaf tissue and which occurs in at least two allelic forms, y (yellow) dominant over g (green).Abbreviations PGI phosphoglucoisomerase - Pgi nuclear gene coding for PGI - TSH tris buffer containing 2-mercaptoethanol - SDS sodium dodecyl sulphate - Chl chlorophyll - LHCP light-harvesting chlorophyll a/b binding protein - ELISA enzyme-linked immunosorbent assay