In vitro shoot regeneration from leaf mesophyll protoplasts of hybrid poplar (Populus nigra x P. maximowiczii)

Summary Protoplasts were isolated from leaf mesophyll of hybrid poplar (Populus nigra X P. maximowiczii) with a mean yield of 10.4 x 10⁶ protoplasts per g fresh weight using 2.0% Cellulase Onozuka R-10, 0.8% Macerozyme R-10, 1.2% Hemicellulase, 2.0% Driselase, and 0.05% Pectolyase Y-23 with CPW salts solution containing 0.6 M mannitol, 0.002 M DTT, 3 mM MES at pH 5.6. A liquid plating method produced the highest frequency of dividing protoplasts (48.6%) using an MS medium without NH₄NO₃. The highest percent of colony formation was 22.8%, produced with fabric supported semi-solid (0.5% w/v) agar plating method using the same culture medium. Growing cell colonies and/or micro-calli were transferred to a fresh semisolid agar medium containing 0.44 M BAP and 9.0 M 2,4-D. Multiple shoots were produced from protoplast-derived callus after culture on MS medium containing 6.8 M zeatin. After root induction on half-strength MS medium that lacked growth regulators, shoots were transferred to pots containing artificial soil mix.Abbreviations CPW Cell and Potoplast Wash solution - LPM Liquid Plating Method - LDM Liquid Drop Method - HDM Hanging Drop Method - FSPM Fabric supported Semi-solid agar Plating Method - DTT Dithiothreitol - MES 2-(N-morpholino) ethane sulfonic acid - BAP 6-benzylaminopurine - 2,4-D 2,4-dichlorophenoxy acetic acid - NAA -naphthalene acetic acid - MS Murashige and Skoog (1962)     

Metacercarial infections of Paragonimus westermani in freshwater crabs sold in markets in Seoul

Status of metacercarial infections of Paragonimus westermani was observed in freshwater crabs, which were purchased at 3 markets in its peak season of 1990. All of 85 crabs were Eriocheir japonicus. No other species of Eriocheir were found. When crushed muscle and viscera was examined individually, the infection rate was 11.8%; and mean number of metacercariae was 2.1 per infected crab. Unless adequately cooked, freshwater crabs are still potential sources of human paragonimiasis.     

Component proteins in crude extract of adult Paragonimus westermani purified by immunoaffinity chromatography using monoclonal antibodies.

The nature of 2 component proteins in crude saline extract of adult Paragonimus westermani was investigated. By immunoaffinity chromatography using monoclonal antibodies (MAb) as ligands, the proteins were purified from the crude extract. Band 1 protein in disc-polyacrylamide gel electrophoresis (PAGE) was purified by PFCK-136 MAb. The protein, known to have molecular mass of 440 kDa, was composed of 23, 46 and 92 kDa subunits when observed by reducing SDS-PAGE and SDS-PAGE/immunoblot. This protein was originated from eggs of the worm as revealed by immunohistochemical staining with PFCK-136 Mab. Another affinity purified protein utilizing PFCK-44 MAb was the band 4 protein of 17 kDa in disc-PAGE. This was a monomer protein in reducing SDS-PAGE and SDS-PAGE/immunoblot. The protein was produced at intestinal epithelium of the worm.     

Triterpene glycosides from Schefflera octophylla.

In addition to 3-epi-betulinic acid, three triterpene glycosides were isolated from leaves of Schefflera octophylla. The structures of the glycosides have been determined as 28-O-[alpha-L-rhamnopyranosyl(1----4)-O-beta-D-glucopyranosyl(1----6)-be ta-D- glucopyranosides of 3 alpha-hydroxy-lup-20(29)-ene-23,28-dioic acid, 3 alpha,11 alpha- dihydroxy-lup20(29)-ene-23,28-dioic acid and 3-epi-betulinic acid by spectroscopic data and chemical transformations. The last two compounds were found for the first time in the plant kingdom.     

Application of a stability statistic to international maize yield trials

Summary Genotype x environment (GE) interaction encountered in experiments complicates genotype selection and varietal recommendation. The integration of yield and stability of genotypes into a single parameter may make selection and recommendation easier. Kang developed a rank-sum method that allows selection for both yield and the stability variance statistics ( _i ² or s _i ² ) of Shukla. The objective of this research was to compare the rank-sum selection method to selection based on yield alone in five international maize (Zea mays L.) yield trials. Ranks were assigned for yield (the highest mean yield received a rank of 1) and for _i ² and s _i ² (the lowest value received a rank of 1). The yield and _i ² ranks and/or the yield and s _i ² ranks for each genotype were summed. Each trial contained two reference entries (REs). Yield rank or rank-sum of each genotype was compared to yield rank or rank-sum of the best RE (BRE). GE interaction was significant for all trials. Heterogeneity in the GE interaction due to the linear effect of a covariate (differences in fertility and/or cultural practices) was significant in Trials 1, 2, and 5. Overall, in all trials, 29 genotypes were selected on the basis of yield alone. On the basis of _i ² and yield rank-sum, 32 genotypes were identified, with 11 being lower yielding than the 29 yield-based selections. On the basis of s _i ² and yield rank-sum, 31 genotypes were selected, with 11 being lower yielding than the yield-bases selections. Obviously, yield is sacrificed when the rank-sum method is used in the selection process. However, selection based on yield alone may not be adequate when GE interaction is significant because of testing in diverse environments.     

垂体后叶素和加压素对离体心肌的直接作用

本实验采用大鼠离体右心房和右心室肌条模型,观察了垂体后叶素和加压素对右心房和右心室肌的直接作用。结果表明:垂体后叶素对右心房的自主性收缩频率和幅度及右心室肌的收缩幅度均有剂量依赖性抑制作用;加压素对右心房和右心室肌收缩幅度也有剂量依赖性抑制作用,但对右心房自主节律无影响;催产素对右心房的收缩频率和幅度则均无影响。加压素V_1、V_2受体拮抗剂d(CH_2)_5Tyr(Me)AVP和d(CH_2)_5(D-Ile~2,Ile~4,Ala(NH_2)~9)AVP对垂体后叶素的负性变力作用具有不同程度的阻断作用,但对垂体后叶素的负性变时作用无阻断作用。以上结果提示,垂体后叶素的负性变力作用主要是由加压素产生的,加压素对心肌有直接的负性变力作用;垂体后叶素的负性变时作用可能是非加压素和催产素成分的作用结果。     

Regulation of protein phosphorylation in pancreatic acini by cyclic AMP-mediated secretagogues: interaction with carbamylcholine

The effects on protein phosphorylation in mouse pancreatic acini of cyclic AMP-mediated secretagogues and the Ca2+-mediated agonist carbamylcholine were compared. Under the conditions adopted for the study of protein phosphorylation, carbamylcholine (3 microM) stimulated amylase release from pancreatic acini 6-fold, whereas vasoactive intestinal polypeptide (VIP) (100 nM) and the cyclic AMP analogue 8-bromo-cyclic AMP (1 mM) caused little or no increase in secretion. However, VIP and 8-bromo-cyclic AMP, when added in combination with carbamylcholine, potentiated the stimulation of amylase release to 170-180% of that caused by carbamylcholine alone. As assessed by two-dimensional gel electrophoresis, VIP reproduced four of the ten changes in protein phosphorylation elicited by carbamylcholine, these changes being the increased phosphorylation of one soluble protein and the decreased phosphorylation of three soluble proteins. VIP enhanced the carbamylcholine-induced changes in phosphorylation for three proteins. In addition, VIP increased the phosphorylation of a unique protein of Mr 52,000 and pI 5.66 which was not affected by carbamylcholine. All of the effects on protein phosphorylation exerted by VIP in the presence or absence of carbamylcholine were mimicked by 8-bromo-cyclic AMP. Secretin also reproduced most of the changes in protein phosphorylation caused by VIP, although concentrations of secretin of at least 100-fold higher were required to elicit a maximal response. It is concluded that cyclic AMP-mediated secretagogues alter the phosphorylation of a unique protein as well as of several pancreatic proteins affected by carbamylcholine. Moreover, these effects appear to be mediated primarily by VIP-preferring receptors and may be involved in the synergistic action of VIP to promote carbamylcholine-induced amylase release.     

Reversible dissociation of active octamer of cyanase to inactive dimer promoted by alteration of the sulfhydryl group

Cyanase is an inducible enzyme in Escherichia coli that catalyzes the reaction of cyanate with bicarbonate resulting in the decomposition of cyanate to ammonia and bicarbonate. In this study, the role of the single sulfhydryl group in each of the eight identical subunits of cyanase was investigated. Tetranitromethane, methyl methanethiosulfonate, N-ethylmaleimide, and Hg2+ all reacted with the sulfhydryl group to give derivatives which had reduced activities and which dissociated reversibly to inactive dimer. Association of inactive dimer to active octamer was facilitated by the presence of azide (cyanate analog) and bicarbonate, increased temperature and enzyme concentration, and presence of phosphate. Nitration of tyrosine residues by tetranitromethane occurred only in the absence of azide and bicarbonate, suggesting that at least some of the tyrosine residues become exposed when octamer dissociates to dimer. Site-directed mutagenesis was used to prepare a mutant enzyme in which serine was substituted for cysteine. The mutant enzyme was catalytically active and had properties very similar to native enzyme, except that it was less stable to treatment with urea and to high temperatures. These results establish that in native cyanase the sulfhydryl group per se is not required for catalytic activity, but it may play a role in stabilizing octameric structure, and that octameric structure is required for catalytic activity.     

Membrane protein phosphorylation during stomatocyte-echinocyte transformation of human erythrocytes

The normal, discoid shape of red blood cells represents an equilibrium between two opposing factors, i.e., stomatocytic and echinocytic transformations. Most stomatocytic agents were found to be inhibitors of calmodulin, a regulator of the phosphorylation of membrane proteins. We determined whether red cell shape transformations could be caused by changes in phosphorylation of membrane proteins, specifically the cAMP-dependent phosphorylation of ankyrin and band 4.1. Red blood cells were incubated with 32P and 100 microM chlorpromazine (stomatocytic transformation) or 30 mM sodium salicylate (echinocytic transformation) for various time intervals. Ghost membrane proteins were examined by polyacrylamide gel electrophoresis and autoradiography. Spectrin (beta-chain), ankyrin, band 3, band 4.1 and 4.9 were phosphorylated. No change was found in the degree and pattern of phosphorylation after stomatocytic transformation. Salicylate caused a reversible inhibition of transmembranous phosphate transport in both directions. The results indicate that the stomatocytic transformation induced by chlorpromazine and the echinocytic transformation induced by salicylate do not involve a change in phosphorylation, but that the echinocytic transformation induced by salicylate is associated with an inhibition of transmembranous transport of phosphate. Studies with salicylate suggest that the phosphorylation sites of band 3 are found mainly on the endofacial side of the membrane.