Optimum prediction of three-way crosses from single crosses in forage maize (Zea mays L.)

Summary Three-way cross means were predicted with formulae involving linear functions of general (GCA) and specific combining ability (SCA) effects estimated from single-cross factorials between genetically divergent populations. Data from an experiment with 66 single-cross and 66 three-way cross forage maize (Zea mays L.) hybrids was used for comparing the prediction formulae. The genotypic correlation (r) between observed and predicted three-way crosses increased with increasing , the weighting factor of SCA effects, for plant height and ear dry matter (DM) content. It displayed slightly convex curves for total and stover DM yield, ear percentage, and metabolizable energy content of stover. For Jenkins' method B, r was considerably less than 1.0 for all traits, indicating the presence of epistasis. The square root of heritability (h) of the predicted means decreased with increasing , the reduction being small with a greater number of test environments. Using the product r·h as a criterion of efficiency, none of the prediction methods was consistently superior and the differences among them were rather small (< 7.5%) for all traits, irrespective of the number of test environments. We recommend evaluating the GCA of a greater number of lines from each parent population in testcrosses with a small number of elite lines from the opposite population. All possible three-way or double crosses between both sets of lines should be predicted by Jenkins's method C. This procedure allows one to select with a higher intensity among the predicted hybrids and thus should increase the genetic gain.Extended version of a paper (Geiger et al. 1986) read at the sixth meeting of the EUCARPIA Section Biometrics in Plant Breeding held at Birmingham, UK, July 28–August 1, 1986     

Contact-dependent regulation of vinculin expression in cultured fibroblasts: a study with vinculin-specific cDNA probes.

Vinculin specific cDNA clones were isolated from chicken embryo fibroblast (CEF) cDNA library in lambda gt11. The clones, ranging in size from 2.8 to 5.0 kb, were initially selected by rabbit antibodies to vinculin. Their identity was further confirmed by their specific reactivities with a battery of different vinculin-specific monoclonal antibodies. Southern blot analysis of restriction enzyme digested chicken spleen DNA suggested that all the isolated cDNA clones correspond to the same gene(s). Northern blot hybridization revealed that the vinculin-specific cDNA clones react with a single 6.5 kb mRNA in total cellular RNA preparations of CEF, whole chicken embryos and chicken gizzard smooth muscle. Moreover, fractionation of CEF poly(A)+ RNA by sucrose gradient centrifugation followed by translation in cell free system indicated that the mRNA coding for vinculin has a size of about 6.0-7.0 kb. The identity of these clones was finally confirmed by selection hybridization assay. The isolated vinculin-specific cDNA probes were subsequently used in order to study the effect of substrate adhesiveness on the expression of vinculin. We show here that cells cultured on highly adhesive substrate, such as endothelial extracellular matrix (ECM), form large vinculin-rich focal contacts, while cells grown on poorly adhesive substrate poly(2-hydroxyethyl methacrylate) [poly(HEMA)] contain only small distorted vinculin spots. These morphological differences were accompanied by over 5-fold reduction in vinculin synthesis in cells growing on poly(HEMA), compared to those cultured on the ECM and over 7.5-fold decrease in the levels of vinculin-specific mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Tissue-type plasminogen activator and urokinase: differences in the reaction pattern with the active-site titrant 4-methylumbelliferyl-p-guanidinobenzoate hydrochloride

The applicability of 4-methylumbelliferyl-p-guanidinobenzoate hydrochloride (MUGB) as active-site titrant for tissue-type plasminogen activator (t-PA) was studied in comparison to urokinase. Although t-PA was capable of cleaving MUGB, active-site titration of t-PA (one-chain form as well as two-chain form) with MUGB was not possible, whereas MUGB titration of urokinase could be performed. We therefore studied the kinetics of the interaction of these two plasminogen activators with MUGB. The equilibrium dissociation constant, KS, for the interaction between MUGB and urokinase was 2.9 X 10(-6) M, and for the interaction with t-PA 3.13 X 10(-5) M. However, one main requirement for active-site titration, namely a stable acyl enzyme intermediate (ES'), was only fulfilled for MUGB urokinase but not for MUGB t-PA. Whereas for the reaction of MUGB and urokinase the first-order acylation rate constant k2 was found to be about 10(6)-times higher than the first-order deacylation rate constant k3 (k2 = 3.76 X 10(-1) s-1, k3 = 3.7 X 10(-7) s-1), the k2/k3 ratio for the reaction of MUGB and t-PA (one- and two-chain form) was 0.77 to 3.85. Therefore, urokinase and t-PA differ in their reaction with this fluorogenic substrate and MUGB cannot be used for active-site titration of tPA.     

Reversible thermal inactivation of the quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus. Ca2+ ions are necessary for re-activation.

The soluble form of the homogeneous quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus is reversibly inactivated at temperatures above 35 degrees C. An equilibrium is established between active and denatured enzyme, this depending on the protein concentration and the inactivation temperature used. Upon thermal inactivation the enzyme dissociates into the prosthetic group pyrroloquinoline quinone and the apo form of glucose dehydrogenase. After inactivation at 50 degrees C active enzyme is re-formed again at 25 degrees C. Ca2+ ions are necessary for the re-activation process. The velocity of re-activation depends on the protein concentration, the concentration of the prosthetic group pyrroloquinoline quinone and the Ca2+ concentration. The apo form of glucose dehydrogenase can be isolated, and in the presence of pyrroloquinoline quinone and Ca2+ active holoenzyme is formed. Even though native glucose dehydrogenase is not inactivated in the presence of EDTA or trans-1,2-diaminocyclohexane-NNN'NH-tetra-acetic acid, Ca2+ stabilizes the enzyme against thermal inactivation. Two Ca2+ ions are found per subunit of glucose dehydrogenase. The data suggest that pyrroloquinoline quinone is bound at the active site via a Ca2+ bridge. Mn2+ and Cd2+ can replace Ca2+ in the re-activation mixture.     

A new type of ultrasensitive bioluminogenic enzyme substrates. I. Enzyme substrates with D-luciferin as leaving group

Derivatives of D-luciferin, D-luciferin methyl ester, D-luciferin O-sulfate, D-luciferin O-phosphate, D-luciferyl-L-N alpha-arginine and D-luciferyl-L-phenylalanine were used as highly sensitive substrates for carboxylic esterase, arylsulfatase, alkaline phosphatase and carboxypeptidases A, B and N. Enzymatic cleavage of the compounds by enzymes leading to the release of D-luciferin was demonstrated. Kinetic constants have been determined for D-luciferin methyl ester and carboxylic esterase, for D-luciferin O-sulfate and arylsulfatase, for D-luciferin O-phosphate and alkaline phosphatase, for D-luciferyl-L-phenylalanine and carboxypeptidase A, and for carboxypeptidases B and N and D-luciferyl-L-N alpha-arginine. All compounds proved to be highly sensitive substrates for the respective enzymes, permitting a limit of detection for enzymes between 10 and 500 fg per assay.     

A Method for Calculating Sucrose Synthesis Rates throughout a Light Period in Sugar Beet Leaves

Sucrose synthesis rate in an exporting sugar beet (Beta vulgaris L.) leaf was calculated from simultaneous measurements of export and changes in leaf sucrose level. The amount of recently fixed carbon exported was determined from net carbon assimilated minus the tracer carbon accumulated in the leaf. The relative amount of ¹⁴C accumulated in the leaf supplied with ¹⁴CO₂ throughout an entire light period was recorded continuously with a Geiger-Mueller detector. To produce a continuous time course for tracer carbon accumulated in the leaf during the light period, the latter curve was superimposed on values for tracer carbon accumulated in leaves sampled at hourly intervals. Validity of the method requires that nearly all of the carbon that is exported be sucrose and that nearly all of the sucrose that is synthesized be either exported or accumulated as sucrose in the exporting leaves. These conditions appeared to be fulfilled in the situations where the method was applied. The method was used to study the effect of increasing atmospheric CO₂ concentration on the rate of sucrose synthesis. Further, the method can be used in conjunction with the gathering of other data such as gas exchange, metabolite levels, and enzyme activities in a set of leaves of a similar age on the same plant. This assemblage of data was found to be useful for understanding how rates of photosynthesis, sucrose synthesis, and translocation are regulated in relation to each other in an intact plant.     

Radioimmunoassay data handling and calculations with a graphics-statistics computer program.

This paper discusses the application of the data handling-graphics-statistics program Stata (Computing Resources Center, Santa Monica, CA) to radioimmunoassay. We have found that this program is more powerful and easier to use than a spreadsheet for analyzing various kinds of laboratory data generated from chromatography, radiolabeling experiments, enzyme-linked immunosorbent assays, and radioimmunoassays, to name several examples. Data from a radioimmunoassay procedure, originally analyzed using a spreadsheet, Lotus 1-2-3, have been processed with Stata. Simple programs (batch files) have been devised for computations and graphics. The original data and a comparison of results are presented.     

Dilazep-induced vasodilation is mediated through adenosine receptors

Administration of dilazep, an inhibitor of adenosine uptake, significantly reduced systemic arterial blood pressure and increased superior mesenteric arterial conductance without affecting the plasma adenosine levels of femoral arterial or portal venous blood. Administration of a bolus dose of 8-phenyltheophylline (8-PT), an antagonist of adenosine receptors, blocked adenosine-mediated autoregulation of the superior mesenteric artery. After the blockade of adenosine receptors by 8-PT, dilazep did not produce vasodilation. These data suggest that dilazep has a vasodilating effect in vivo that is mediated by adenosine.     

Carbon Assimilation and Leaf Water Status in Sugar Beet Leaves during a Simulated Natural Light Regimen

Carbon assimilation and leaf water status were studied in sugar beet (Beta vulgaris L., Klein E-type multigerm) leaves during a light period in which illumination either increased rapidly to full irradiance or changed gradually in a sinusoidal manner as generally occurs during a natural day. A light regimen that simulated the light of a natural day was produced by adjusting irradiance with a neutral-density filter under the control of a computer. Under this light regimen, photosynthesis, transpiration, and stomatal conductance followed the irradiance pattern very closely and ribulose bisphosphate carboxylase was nearly fully activated. When illumination was increased rapidly at the beginning of a light period, transpiration also increased quickly, causing leaves to wilt to some extent. The activation state of ribulose bisphosphate carboxylase increased to only 52%, but ribulose bisphosphate level was nearly twice as high as during the simulated natural day. In spite of the differences in activation state and ribulose bisphosphate levels, photosynthesis rates were very similar under both regimens. Nevertheless, differences in parameters between leaves under the two irradiance regimens can affect how a plant responds to internal or external factors, and therefore, the rate at which irradiance increases at the beginning of a light period is an important consideration when interpreting data.