Identification of somatic hybrids in plant protoplast fusions with monoclonal antibodies to plasma-membrane antigens

Monoclonal antibodies generated by immunization with a plasma-membrane preparation from suspension-cultured cells of Nicotiana glutinosa L. were used in combination with fluoresceinor rhodamine-labeled goat anti-mouse immunoglobulins to identify heterokaryons in protoplast fusion procedures. Antibody labeling did not inhibit callus formation nor plantlet regeneration. The antibodies are non-invasive and surface labeling provides clear optical discrimination of true heterokaryons from unfused aggregates as well as from parental protoplasts and homokaryons. Labeling is stable throughout fusion and hence by pre-labeling parental protoplast populations the strategy is both versatile and of general applicability.     

Spinach ferredoxin is a calcium-binding protein

Spinach-leaf ferredoxin was identified as a calcium-binding protein by ⁴⁵Ca autoradiography on nitrocellulose membranes and with the cationic carbocyanine dye 1-ethyl-2-[3-(1-ethylnaphtho[1,2-d]thiazolin-2-ylidene)-2-methylpropenyl] naphtho[1,2-d]thiazolium bromide (stains-all). Binding of ⁴⁵Ca was observed at pH 6.8 and pH 7.8 and in the presence of 5 mM and 20 mM MgCl₂. At the higher MgCl₂ concentration the Ca²⁺-binding capacity is reduced. Only micromolar concentrations of LaCl₃, however, are required to achieve a similar effect. Both the oxidized and reduced forms of ferredoxin bind calcium.Abbreviations PAGE polyacrylamide gel electrophoresis - SDS sodium dodecyl sulfate - stains-all 1-ethyl-2-[3-(1-ethylnaphtho[1,2-d]thiazolin-2-ylidene)-2-methylpropenyl] naptho[1,2-d]thiazolium bromide     

Expression of the β-subunit of β-conglycinin in seeds of transgenic plants

Soybean (Glycine max (L.) Merr.) seeds contain the storage protein -conglycinin, encoded by a multigene family. -Conglycinin consists of three subunits; , , and . A genomic clone for a -subunit of -conglycinin has been characterized by restriction-enzyme mapping and hybrid selected in-vitro translation followed by immunoprecipitation. In order to determine the developmental regulation of this -subunit gene, its expression was studied in seeds of transgenic petunia (Petunia hybrida) and tobacco (Nicotiana tabacum L.) plants. The -subunit expressed in seeds of petunia and tobacco was recognized by anti--conglycinin serum at a relative molecular mass of 53 000, equivalent to that of the native protein. Separation of the petunia-seed proteins by isoelectric focusing followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis showed that multiple isoelectric forms of the -subunit were produced. There was approximately a twofold variation in the accumulation of the -subunit protein in the mature seeds of transgenic petunia plants, each containing a single -subunit gene. However, the level of protein accumulation in mature seeds and the amount of -subunit mRNA in developing seeds was not correlated. Accumulation of the -subunit protein in transgenic seeds was less than the -subunit protein that accumulated in transgenic petunia seeds containing a single -subunit gene and less than the amount of the -subunit in mature soybean seeds which contain 8–13 -subunit genes. In transgenic tobacco plants, the accumulation of the -subunit protein in seeds was generally well correlated with the number of genes that were incorporated in the different transformants.Abbreviations kb kilobase - kDa kilodalton - M_r relative molecular mass - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis     

A monoclonal antibody specific for the red-absorbing form of phytochrome

Characterisation of a new monoclonal antibody (mAb), designated LAS 41, directed against 124-kilodalton (kDa) etiolated-oat (Avena sativa L.) phytochrome, indicates that it recognises an epitope unique to the red-light-absorbing form, Pr. In a solid-phase enzyme-linked immunosorbent assay (ELISA), LAS 41 exhibits a seven- to eight-fold higher affinity for Pr than for the far-red-light-absorbing form of phytochrome, Pfr. In addition, in immunoprecipitation assays LAS 41 effectively precipitates 100% of phytochrome presented as Pr but only precipitates a maximum of 24.5% of phytochrome presented as Pfr. These values are indicative of binding exclusively to Pr. Peptide-mapping studies show that LAS 41 recognises and epitope located within a region 6–10 kDa from the aminoterminus of the phytochrome molecule. Since binding of LAS 41 to Pr induces alterations in the spectral properties of Pr, this indicates that at least part of the 4 kDa domain to which the antibody binds is essential for protein-chromophore interaction. Subsequent photoconversion of LAS 41-Pr complexes produces native Pfr spectra, with concomitant production of free antibody and antigen, as shown by a modified ELISA. The specificity of LAS 41 for Pr has facilitated the purification of Pfr which is free of contaminating Pr. This has enabled direct determination of the mole fraction of Pfr established by red light to be 0.874.Abbreviations ELISA enzyme-linked immunsorbent assay - kDa kilodalton - mAb monoclonal antibody - Pfr far-red-absorbing form of phytochrome - Pr red-absorbing form of phytochrome - SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis - (A) difference in absorbance (A ₆₆₅ ^Pr –A ₇₃₀ ^Pr )-(A ₆₆₅ ^Pfr –A ₇₃₀ ^Pfr ) - Ar/Afr spectral change ratio (SCR) - max mole fraction of Pfr following saturating red light     

NAD(P)H oscillation in relation to the rhythmic contraction in thePhysarum plasmodium

Summary ThePhysarum plasmodium shows rhythmic contractile activities with a period of a few min. Phases of the oscillation in the plasmodium migrating unindirectionally agreed sideways throughout at the frontal part. So, time course of an intracellular chemical component was determined by analyzing small pieces cut off successively from the frontal part of the large plasmodium. Intracellular NAD(P)H concentration oscillated with the same period as the rhythmic contraction but with a different phase advancing about 1/3 of the period. UV irradiation suppressed the rhythmic contraction without affecting the rhythmic variation of NAD(P)H. Thus, the NAD(P)H oscillator works independently of the rhythmic contractile system, but seems entraining with each other.Abbreviations UV ultraviolet - NADH nicotinamide adenine dinucleotide, reduced form - NADPH nicotinamide adenine dinucleotide phosphate, reduced form - ATP adenosine 5-triphosphate - cAMP cyclic adenosine 3, 5-monophosphate - FMNH₂ flavin mononucleotide, reduced form - TCA tricarboxylic acid - BSA bovine serum albumin - DTT dithiothreitol     

Fertile revertants from S-type male-sterile maize grown in vitro

Summary Plants were regenerated from callus cultures of maize inbred W182BN with the S(USDA) type of cytoplasmic male sterility (cms). Some regenerates from 16 of 18 separate cultures had fertile tassels. Many other regenerates, whose fertility could not be scored accurately because of abnormal plant morphology, produced fertile progeny after pollination with N cytoplasm W182BN. Revertant plants and/or progeny were obtained from all 18 cultures, which included the CA, D, LBN, and S sources of cmsS. More revertants were recovered from cultures maintained as callus for 12 months than from 3–4 month old cultures. Several types of evidence (absence of segregation for fertility after selfing or pollination of revertants with standard W182BN, pollen viability counts, failure of revertants to restore sterile cmsS lines to fertility, mitochondrial DNA analyses) indicated that the reversion to fertility involved cytoplasmic rather than nuclear alterations. All revertants examined lacked the S1 and S2 plasmid-like DNAs characteristic of the mitochondrial genome of sterile cmsS lines. Most callus cultures lost S1 and S2 after 13–20 months in vitro. No revertants were seen among thousands of W182BN cmsS plants grown from seed in the field or among plants from tissue cultures of W182BN with the C or T types of cms. The cytoplasmic revertants recovered from culture may be useful for the molecular analysis of cmsS.     

五种雀形目鸟类核型的比较研究

染色体核型的比较研究,对于了解物种的特性,探索物种的遗传、进化、系统发育及分类地位都有一定的意义。有关鸟类的核型研究,国外已有不少报道(Castrovicja,1969; Hammar,1966,1975; Ray-Chaudhuri et al.,1969;Shidds,1982;Ta—kagi,1972等)国内在这方面的工作开展较迟,已作过核型分析的鸟类为数不多,仅见王应祥等(1982)的报道。迄今,作过核型研究的鸟类已达500多种,其中雀形目有200种。本文比较分析了五种野生雀形目鸟类的核型,它们是黑头蜡嘴雀,斑鸫,黄腹山雀,红尾伯劳和灰背椋鸟。     

Food composition of Clarias gariepinus (= C. lazera) (Cypriniformes,Clariidae) in Lake Kinneret (Israel)

The feeding behaviour of the freshwater piscivore, Clarias gariepinus (C. lazera) (C & V 1840) was studied over two periods: 1973–1975 and 1981–1982, in Lake Kinneret (Israel). The total number of fish analysed was 264 and their sizes (SL) and weights varied between 238 and 830 mm (146 to 5728 g). More than fifty species of plants and animals from the plankton, benthos and nekton of Lake Kinneret were identified in the intestines of C. gariepinus. Preyed fish were the most abundant food component (81%) and constituted the highest biomass, with Mirogrex terraesanctae representing the majority (although other species were also found). The potential impact of piscivory in the Kinneret ecosystem is considered.     

Subcellular Location and Neuronal Release of Diazepam Binding Inhibitor

Diazepam binding inhibitor (DBI), a peptide located in CNS neurons, blocks the binding of benzodiazepines and beta-carbolines to the allosteric modulatory sites of gamma-aminobutyric acid (GABAA) receptors. Subcellular fractionation studies of rat brain indicate that DBI is compartmentalized. DBI-like immunoreactivity is highly enriched in synaptosomes obtained by differential centrifugation in isotonic sucrose followed by a Percoll gradient. In synaptosomal lysate, DBI-like immunoreactivity is primarily associated with synaptic vesicles partially purified by differential centrifugation and continuous sucrose gradient. Depolarization induced by high K+ levels (50 mM) or veratridine (50 microM) released DBI stored in neurons of superfused slices of hypothalamus, hippocampus, striatum, and cerebral cortex. The high K+ level-induced release is Ca2+ dependent, and the release induced by veratridine is blocked by 1.7 microM tetrodotoxin. Depolarization released GABA and Met5-enkephalin-Arg6-Phe7 together with DBI. DBI is also released by veratridine depolarization, in a tetrodotoxin-sensitive fashion, from primary cultures of cerebral cortical neurons, but not from cortical astrocytes. Depolarization fails to release DBI from slices of liver and other peripheral organs. These data support the view that DBI may be released as a putative neuromodulatory substance from rat brain neurons.     

Position of the Peptide Linkage in Glutamyl-Taurine from Calf Brain Synaptic Vesicles

To elucidate the position of the peptide bond in glutamyl-taurine this dipeptide was extracted from calf brain synaptic vesicles and subjected to paper electrophoresis. It was analyzed further in an automatic amino acid analyzer prior and subsequent to acid hydrolysis. Both alpha- and gamma-forms were found to be present in approximately equal amounts.