Purification and characterization of glutathione reductase from corn mesophyll chloroplasts

Differences in the apparent molecular weights of the subunits of glutathione reductase (EC 1.6.4.2) from pea chloroplasts and corn mesophyll chloroplasts have been recently reported. In order to more fully describe the differences between the enzymes from these two sources, glutathione reductase from the mesophyll chloroplasts of corn seedlings ( Zea mays L. cv. G-4507) has been purified 200-fold by affinity chromatography using adenosine 2',5'-disphosphate agarose. The purified enzyme had a specific activity of 26 μmol NADPH oxidized (mg protein)^-1 min^-1. The native enzyme had a relative molecular weight of 190 ± 30 kDa and exhibited polypeptides of 65, 63, 34, and 32 kDa when separated on sodium dodecylsulfate-polyacrylamide gels. Comparisons of the results from electroblotting, native molecular weight and subunit molecular weight analyses suggest that the enzyme exists as a heterotetramer. Optimal enzyme activity was obtained at pH 8 in N-2-hydroxyethyl-piperazine-N'-2-ethanesulfonic acid (HEPES-NaOH) buffer. The sulfhydryl reagent, n -ethylmaleimide, inhibited enzymatic activity when incubated in the presence of NADPH while no inhibition was detected with oxidized glutathione in the incubation mixture. Reduced glutathione (5 m M ) inactivated the enzyme by 50%. This inactivation followed first order kinetics with a rate constant of 0.0028 s^-1. The enzyme was also inactivated by NADPH. The inactivation reached ca 90% within 30 min and followed first order kinetics with a rate constant of 0.0015 s^-1.     

Rapid immunofluorescent determination of cells in the S phase in pea root meristems: An alternative to autoradiography

Photosystem II (PS II) activity and the localization of ribulose-l,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39) were studied in primary leaves of young maize plants ( Zea mays L. cv. Fronica) by tetra-nitro-blue-tetrazoliumchloride reduction and immunolocalization, respectively. In tissue of 3-day-old plants all chloroplasts were structurally identical. From day 4 they developed into their typical appearance of mesophyll and bundle sheath chloroplasts. First PS II-activity was present in both types of chloroplasts. From day 4 it disappeared in bundle sheath chloroplasts concomitant with the loss of grana. RuBP carboxylase on the other hand was only present in bundle sheath chloroplasts at all stages of development. Thus, the control of the development of the photosystems and the Calvin cycle enzymes seem to differ.     

Synthesis of ribulose 1,5-bisphosphate carboxylase by isolatedSorghum mesophyll chloroplasts

Chloroplasts isolated fromSorghum vulgare are active in light-dependent, organelle protein synthesis. Intact chloroplasts can use light as an energy source; photosynthetically inactive chloroplasts require the addition of ATP for this protein synthesis. Preincubation of chloroplasts in light at 25°C for 1 h depleted the endogenous templates completely; such preincubated chloroplasts translated exogenously added heterologous templates efficiently. When total cellular RNA fromChlorella protothecoides, a C₃ plant, was used as template for translation in a cell-free light-dependent system of isolated mesophyll chloroplasts fromSorghum vulgare, a C₄ type plant, polypeptides of 55 kDa (large subunit) and 15 kDa (small subunit) were detectable in the fluorographic profile of the newly synthesized proteins; these polypeptides were absent in the products obtained with endogenous RNA. Evidence for the fidelity of the system was obtained by immunological analysis of ribulose 1, 5-bisphosphate carboxylase obtained by the translation ofChlorella cellular RNAs.     

Measurement of light within thin plant tissues with fiber optic microprobes

Vogelmann, T. C., Knapp, A. K., McClean, T. M. and Smith, W. K. 1988. Measurement of light within thin plant tissues with fiber optic microprobes. - Physiol. Plant. 72: 623–630.
The measurement of light with fiber optic microprobes has been extended to thin (200–300 μm) plant tissue samples. To test the method, light measurements were made in thin aqueous films and paradermal sections from 10-day-old etiolated Cucurbita pepo L. cv. Fordhook cotyledons. The measurements obtained were highly reproducible. Paradermal sections of spongy mesophyll that were irradiated with collimated light scattered light more effectively than the palisade layer of intact cotyledons. These results demonstrate that different plant tissues have different light scattering characteristics. The successful extension of the fiber optic microprobe technique to thin systems makes it possible to examine the optical properties of different cell layers within leaves and other plant organs.     

桑树叶肉原生质体培养再生植株

近年来,木本植物原生质作诱导再生植株的研究越来越受到国内外学者关注。但在林木树种中,迄今成功的种类仍然不多,在文     

Measurement of chlorophyll fluorescence within leaves using a fibreoptic microprobe

Abstract. The distribution of chlorophyll fluorescence was measured within leaves of Medicago saliva with a fibre optic microprobe. Leaves were irradiated with broad band blue light (1000 μmol m⁻²s⁻¹) and chlorophyll fluorescence was measured at 688 nm. The amount of fluorescence measured within the leaf depended upon the direction in which the probe was inserted. When the probe was advanced directly through the leaf from the shaded towards the irradiated surface, the maximum amount of detected fluorescence occurred near the boundary between the palisade and spongy mesophyll. When the probe was advanced through the leaf from the opposite direction maximum detected fluorescence was at the boundary between the epidermis and palisade. These results appear to be a consequence of the blue light gradient, which declined exponentially within the palisade but was counterbalanced by increasing chlorophyll content within the leaf. Modelling indicates that the measured distribution of chlorophyll fluorescence can be explained by relatively uniform emission of fluorescence throughout the palisade layer, indicating that the chloroplasts may be photosynthetically specialized to their light environment within the leaf.     

Transformation of Nicotiana tabacum,N. debneyi,and N. rustica: Inheritance and protoplast expression of antibiotic resistance

Agrobacterium tumefaciens strains harbouring plasmid vectors pBCAT1, pVU1011 or pMON806 were used to transform leaf explants of Nicotiana tabacum cultivars Delgold and Candel, N. debneyi, and N. rustica var. NRT. Transgenic plants resistant to the selective agents kanamycin, hygromycin or methotrexate were regenerated and used as sources of leaf mesophyll protoplasts. Protoplasts divided and regenerated plants in the presence of selective agents at levels inhibitory to protoplasts of non-transformed plants. Cross-resistance of protoplasts to more than one selective agent was not observed in this study which suggests that this approach may lead to an efficient interspecific somatic hybrid selection system.     

Beneficial interaction between photosynthesis and respiration in mesophyll protoplasts of pea during short light-dark cycles

The respiratory uptake or photosynthetic evolution of oxygen by mesophyll protoplasts of pea ( Pisum sativum L. cv. Arkel) were monitored during successive short. (3–5 min) cycles of darkness and illumination. The rate of respiration was nearly doubled after 3–4 short periods of illumination while there was a 15–20% enhancement in photosynthesis with cycles of illumination and darkness preceding illumination. Such interaction between photosynthesis and respiration was statistically significant when bicarbonate was present in the reaction medium. The inhibitors of photosynthesis [3(3,4–dichlorophenyl)-l,l-dimethylurea (DCMU), glyceraldehyde] decreased respiration after periods of illumination, whereas inhibitors of respiratory electron transport (Rotenone, antimycin A, NaN₃) suppressed photosynthesis, as well. We suggest that a rapid beneficial interaction exists between photosynthesis and respiration in protoplasts, even during short cycles of light and darkness.     

水分胁迫对小麦叶片光合作用的影响及其与抗旱性的关系

在水分胁迫初期,两个小麦品种叶片光合速率,气孔导度和细胞间隙CO_2浓度降低,气孔限制值增加,光合速率的降低主要是气孔因素的限制。中度到严重水分胁迫使叶片光合速率、气孔导度和气孔限制值降低,细胞间隙CO_2浓度明显增加,且叶圆片放氧能力,叶绿体Hill反应、叶绿素荧光强度和表观量子产额降低,此时光合速率的降低主要是叶肉细胞光合活性的下降引起的。抗旱性弱的郑引一号叶肉细胞光合活性比抗旱性强的丰抗13更容易受到水分胁迫的影响。