Optimizing purebred selection for crossbred performance using QTL with different degrees of dominance

A method was developed to optimize simultaneous selection for a quantitative trait with a known QTL within a male and a female line to maximize crossbred performance from a two-way cross. Strategies to maximize cumulative discounted response in crossbred performance over ten generations were derived by optimizing weights in an index of a QTL and phenotype. Strategies were compared to selection on purebred phenotype. Extra responses were limited for QTL with additive and partial dominance effects, but substantial for QTL with over-dominance, for which optimal QTL selection resulted in differential selection in male and female lines to increase the frequency of heterozygotes and polygenic responses. For over-dominant QTL, maximization of crossbred performance one generation at a time resulted in similar responses as optimization across all generations and simultaneous optimal selection in a male and female line resulted in greater response than optimal selection within a single line without crossbreeding. Results show that strategic use of information on over-dominant QTL can enhance crossbred performance without crossbred testing.     

Secretion of extracellular proteins by Pseudomonas aeruginosa

Pseudomonas aeruginosa is a bacterial species of commercial value secreting numerous extracellular proteins, involved in pathogenesis. Most strains produce at least a lipase, a phospholipase, an alkaline phosphatase, an exotoxin and 2 proteases (elastase and alkaline protease). Various mechanisms for secretion of exoproteins appear to exist in P aeruginosa. Genetic analysis has led to the identification of 2 secretion pathways: i) a "general" secretion pathway, defined by the xcp mutations, which mediates secretion of most extracellular proteins, and; ii) an independent secretion pathway specific for alkaline protease. Our present knowledge on the pathways and components of the secretion machinery in P aeruginosa is reviewed in this article.     

Effect of postnatal development on calcium currents and slow charge movement in mammalian skeletal muscle

Single- (whole-cell patch) and two-electrode voltage-clamp techniques were used to measure transient (Ifast) and sustained (Islow) calcium currents, linear capacitance, and slow, voltage-dependent charge movements in freshly dissociated fibers of the flexor digitorum brevis (FDB) muscle of rats of various postnatal ages. Peak Ifast was largest in FDB fibers of neonatal (1-5 d) rats, having a magnitude in 10 mM external Ca of 1.4 +/- 0.9 pA/pF (mean +/- SD; current normalized by linear fiber capacitance). Peak Ifast was smaller in FDB fibers of older animals, and by approximately 3 wk postnatal, it was so small as to be unmeasurable. By contrast, the magnitudes of Islow and charge movement increased substantially during postnatal development. Peak Islow was 3.6 +/- 2.5 pA/pF in FDB fibers of 1-5-d rats and increased to 16.4 +/- 6.5 pA/pF in 45-50-d-old rats; for these same two age groups, Qmax, the total mobile charge measurable as charge movement, was 6.0 +/- 1.7 and 23.8 +/- 4.0 nC/microF, respectively. As both Islow and charge movement are thought to arise in the transverse-tubular system, linear capacitance normalized by the area of fiber surface was determined as an indirect measure of the membrane area of the t-system relative to that of the fiber surface. This parameter increased from 1.5 +/- 0.2 microF/cm2 in 2-d fibers to 2.9 +/- 0.4 microF/cm2 in 44-d fibers. The increases in peak Islow, Qmax, and normalized linear capacitance all had similar time courses. Although the function of Islow is unknown, the substantial postnatal increase in its magnitude suggests that it plays an important role in the physiology of skeletal muscle.     

The influence of amino-acid sequence on protein structure

On the basis of the known sequences and structures of myoglobin, and alpha and beta hemoglobin, a possible correlation between certain amino acids in the sequence and the location of the helical and non-helical parts of the structure is suggested. The presence in the sequence of four critical groups; proline, aspartic acid, glutamic acid, or histidine appears to be necessary (although the last three are not sufficient) for a helical disruption to form. Additional support for this correlation is obtained from analyses of proline replacement in mutant and variant proteins. A mechanism based on hydrophobic bonding is proposed as a rationale for the apparent behavior of these groups. On the basis of these rules and correlations, secondary structures can be proposed for lysozyme and tobacco mosaic virus protein which are consistent with several pieces of evidence.     

Mutational analysis of the RecA protein L1 region identifies this area as a probable part of the co-protease substrate binding site

Previous mutational analysis of the L1 region of the RecA protein suggested that Gly-157 and Glu-158 are 'hot-spots' for the occurrence of constitutive LexA co-protease mutants (coprt^c). In the present study, we clearly establish that position 157 is a hot-spot for the occurrence of such mutants, as 12 of 14 and 10 of 14 substitutions result in this phenotype for UmuD and LexA cleavage respectively. The frequency of such mutations at position 158 is somewhat lower, 8 of 13 and 5 of 13 for UmuD and LexA respectively. Comparison of the UmuD vs. LexA co-protease activity for all single mutants with substitutions at positions 154, 155, 156, 157 and 158 (47 in total) reveals that, although there is good agreement among most mutants regarding their ability to cleave both LexA and UmuD, there are two in particular (Glu-154→Asp and Glu-154→Gln) that show a clear preference for cleavage of UmuD. We also show that three second-site mutations that completely suppress coprt^c activity toward LexA have little or no effect on the coprt^c activity of the primary mutant toward UmuD. In addition, we observe a high frequency of second-site suppressor mutations, suggesting a functional interaction among side-chains in this region. Together, these results support the idea that the L1 region of RecA makes up part of the co-protease substrate-binding site.     

The secretion genes of Pseudomonas aeruginosa alkaline protease are functionally related to those of Erwinia chrysanthemi proteases and Escherichia coliα-haemolysin

The extracellular alkaline protease produced by Pseudomonas aeruginosa is secreted by a specific pathway, independent of the pathway used by most of the other extracellular proteins of this organism. Secretion of this protease is dependent on the presence of several genes located adjacent to the apr gene. Complementation studies have shown that PrtD, E, and F, the three secretion functions for Erwinia chrysanthemi proteases B and C (Létoffé et al., 1990), can mediate the secretion of the alkaline protease by Escherichia coli. The secretion functions involved in alpha-haemolysin secretion in E. coli (hlyB, hlyD, tolC) can also be used to complement alkaline protease secretion by E. coli, although less efficiently. These data indicate that protease secretion mechanisms in Pseudomonas and Erwinia are very similar and are homologous to that of E. coli alpha-haemolysin.     

Effects of tetracyclines on aldosterone- and insulin-mediated Na+ transport in the toad urinary bladder

The effect of oxytetracycline and demethylchlortetracycline on aldosterone- and insulin-mediated Na⁺ transport (short-circuit current) were examined in toad urinary bladders mounted in modified Ussing chambers. Oxytetracycline had little or no effect on either basal or aldosterone-mediated Na⁺ transport. In contrast, demethylchlortetracycline markedly inhibited both basal and aldosterone-mediated Na⁺ transport. Furthermore, demethylchlortetracycline inhibited the aldosterone response significantly out of proportion to its effects on basal Na⁺ transport. Neither of the drugs had an effect on insulin-mediated Na⁺ transport. Consequently, the natriuresis observed in certain patients treated with demethylchlortetracyline may be related to drug-induced renal resistance to the effects of aldosterone.     

A study on the minimum number of loci required for genetic evaluation using a finite locus model

For a finite locus model, Markov chain Monte Carlo (MCMC) methods can be used to estimate the conditional mean of genotypic values given phenotypes, which is also known as the best predictor (BP). When computationally feasible, this type of genetic prediction provides an elegant solution to the problem of genetic evaluation under non-additive inheritance, especially for crossbred data. Successful application of MCMC methods for genetic evaluation using finite locus models depends, among other factors, on the number of loci assumed in the model. The effect of the assumed number of loci on evaluations obtained by BP was investigated using data simulated with about 100 loci. For several small pedigrees, genetic evaluations obtained by best linear prediction (BLP) were compared to genetic evaluations obtained by BP. For BLP evaluation, used here as the standard of comparison, only the first and second moments of the joint distribution of the genotypic and phenotypic values must be known. These moments were calculated from the gene frequencies and genotypic effects used in the simulation model. BP evaluation requires the complete distribution to be known. For each model used for BP evaluation, the gene frequencies and genotypic effects, which completely specify the required distribution, were derived such that the genotypic mean, the additive variance, and the dominance variance were the same as in the simulation model. For lowly heritable traits, evaluations obtained by BP under models with up to three loci closely matched the evaluations obtained by BLP for both purebred and crossbred data. For highly heritable traits, models with up to six loci were needed to match the evaluations obtained by BLP.     

A focus on fixation

Three fixation issues related to immunostaining are discussed here: 1) Generally, a tissue block is fixed, then embedded and sectioned (pre-fixation). The type of fixative applied, crosslinking or coagulating, has an impact on selecting an epitope retrieval method. Individual antigens have a fixation–retrieval characteristic. 2) A long fixation time, especially with crosslinking fixatives, may compromise the result of immunostaining. This negative effect varies among different antigens and can be partially restored by applying a more sensitive/efficient detection system such as tyramide amplification. 3) Sections cut from a fresh frozen tissue block usually are acetone fixed (post-fixation). This was accepted as the “gold standard” for a long time. Post-fixation, however, may have serious consequences for preservation of small peptides leaking from the cut open cells, whereas this is not the case with pre-fixed intact cells. Consequently, the concept of an acetone post-fixed cryostat tissue section as “gold standard” no longer exists and a more appropriate use of the terms immunohistochemistry and immunocytochemistry therefore seems justified. For many antibodies, it is not known whether a formalin fixed, paraffin embedded tissue specimen is appropriate. Suggestions are made for creating a positive control cell block for testing such antibodies.