PubMed Health⌕ Search

Biomedical subjects

K W Siggens

Publications and source records attributed to K W Siggens.

12 recordsLinked to original sources
PubMed2006

Genetic diversity analysis using lowly polymorphic dominant markers: the example of AFLP in pigs.

DNA markers are commonly used for large-scale evaluation of genetic diversity in farm animals, as a component of the management of animal genetic resources. AFLP markers are useful for such studies as they can be generated relatively simply; however, challenges in analysis arise from their dominant scoring and the low level of polymorphism of some markers. This paper describes the results obtained with a set of AFLP markers in a study of 59 pig breeds. AFLP fingerprints were generated using four primer combinations (PC), yielding a total of 148 marker loci, and average harmonic mean of breed sample size was 37.3. The average proportion of monomorphic populations was 63% (range across loci: 3%-98%). The moment-based method of Hill and Weir (2004, Mol Ecol 13:895-908) was applied to estimate gene frequencies, gene diversity (F(ST)), and Reynolds genetic distances. A highly significant average F(ST) of 0.11 was estimated, together with highly significant PC effects on gene diversity. The variance of F(ST) across loci also significantly exceeded the variance expected under the hypothesis of AFLP neutrality, strongly suggesting the sensitivity of AFLP to selection or other forces. Moment estimates were compared to estimates derived from the square root estimation of gene frequency, as currently applied for dominant markers, and the biases incurred in the latter method were evaluated. The paper discusses the hypotheses underlying the moment estimations and various issues relating to the biallelic, dominant, and lowly polymorphic nature of this set of AFLP markers and to their use as compared to microsatellites for measuring genetic diversity.

Animals↗
PubMed2006

Genetic diversity within and between European pig breeds using microsatellite markers.

An important prerequisite for a conservation programme is a comprehensive description of genetic diversity. The aim of this study was to use anonymous genetic markers to assess the between- and the within-population components of genetic diversity for European pig breeds at the scale of the whole continent using microsatellites. Fifty-eight European pig breeds and lines were analysed including local breeds, national varieties of international breeds and commercial lines. A sample of the Chinese Meishan breed was also included. Eleven additional breeds from a previous project were added for some analyses. Approximately 50 individuals per breed were genotyped for a maximum of 50 microsatellite loci. Substantial within-breed variability was observed, with the average expected heterozygosity and observed number of alleles per locus being 0.56 [range 0.43-0.68] and 4.5 respectively. Genotypic frequencies departed from Hardy-Weinberg expectations (P < 0.01) in 15 European populations, with an excess of homozygotes in 12 of them. The European breeds were on average genetically very distinct, with a Wright F(ST) index value of 0.21. The Neighbour-Joining tree drawn from the Reynolds distances among the breeds showed that the national varieties of major breeds and the commercial lines were mostly clustered around their breeds of reference (Duroc, Hampshire, Landrace, Large White and Piétrain). In contrast, local breeds, with the exception of the Iberian breeds, exhibited a star-like topology. The results are discussed in the light of various forces, which may have driven the recent evolution of European pig breeds. This study has consequences for the interpretation of biodiversity results and will be of importance for future conservation programmes.

Alleles↗
PubMed1990

Detection of Listeria species and Listeria monocytogenes using polymerase chain reaction.

Five oligonucleotide sequences are described that were used as primers in the polymerase chain reaction (PCR) to amplify specific sequences from Listeria DNA. When all five primers were used in combination, three PCR products were possible; a Listeria specific product that occurs with DNA from any Listeria sp., a Listeria monocytogenes specific product that occurs only in the presence of DNA from this organism and a universal product that is found using DNA from any bacterial source. The occurrence of these PCR products was used as a diagnostic test on bacteria isolated from various food samples to detect Listeria sp. and L. monocytogenes.

Base Sequence↗
PubMed1987

Molecular cloning and characterization of the beta-amylase gene from Bacillus circulans.

A gene encoding the beta-amylase of Bacillus circulans was isolated from a lambda library and sequenced. The structural gene consists of a 1725 bp open reading frame encoding a polypeptide with a predicted molecular wt of 62830 Daltons. Two active forms of the enzyme were found when the gene was expressed in E. coli. The larger 60 kD form was approximately 3 kD larger than the mature beta-amylase secreted from B. circulans, suggesting that processing of this protein is different between the two species. The smaller 49 kD form is also present at a low level in B. circulans and may result from proteolytic cleavage. The enzyme has a temperature optimum of 50 degrees C. Two other genes, one encoding an alpha-amylase and one a pullulanase, were also isolated from the lambda library.

Amino Acid Sequence↗
PubMed1985

Interferon gamma mRNA "superinduction" in human lymphocytes.

Human peripheral blood mononuclear lymphocytes produce interferon gamma (IFN-gamma) in response to stimulation by mitogens. Previous studies on the kinetics of IFN-gamma mRNA production upon mitogen induction, showed that steady-state levels of mRNA increased to a maximum at 12-24 h post-induction after which they declined to levels not detectable by the assay used. We show here that in mitogen induced peripheral blood lymphocytes, inhibition of protein synthesis using three different inhibitors (cycloheximide, puromycin, pactamycin) resulted in an increase in the steady-state levels of IFN-gamma mRNA. The levels of mRNA in cells treated with inhibitor 16 h post-induction were up to 3-fold higher than in untreated cells. Superinduction was possible up to 40 h post-induction after which the steady-state levels of mRNA had declined to limits below detection; IFN-gamma mRNA was not superinduced by cycloheximide in the presence of actinomycin D.

Cells, Cultured↗
PubMed1985

Role of protein synthesis in induction of interferon-gamma by mitogens in human lymphocytes.

The effect of inhibition of protein synthesis on interferon gamma (IFN-gamma) mRNA induction has been examined in human peripheral blood leukocytes and growing T lymphoblasts. Inhibition of protein synthesis by cycloheximide or puromycin when lymphocytes were stimulated with mitogen alone had little effect on the steady-state levels of IFN-gamma mRNA induced. Activation of the IFN-gamma gene appears to occur independently of synthesis of protein factors. Synergistic induction of IFN-gamma by mitogen plus the phorbol ester mezerein was at least in part accounted for by increased levels of IFN-gamma mRNA in both fresh lymphocytes and growing lymphoblasts. Inhibition of protein synthesis abolished the synergistic effect on mRNA leading to levels similar to those observed in cells treated with mitogen alone. Synergistic induction is dependent upon synthesis of protein factors either within the cells or produced as soluble mediators; these factors are not simply lymphokines since addition of exogenous factors to the cultures did not reverse the block on synergy by the inhibition. These data suggest that protein factors though they may be important in exerting qualitative control on the level of production of IFN-gamma have no role in the initial activation of the gene.

Blood Proteins↗
PubMed1984

Differences in the expression of the human interferon-gamma gene in fresh lymphocytes and cultured lymphoblasts.

Fresh human peripheral blood mononuclear lymphocytes and lymphoblasts that had been grown for a period in T-cell growth-factor containing medium were stimulated with staphylococcal enterotoxin A plus mezerein to produce interferon-gamma (IFN-gamma). Growing lymphoblasts produced peak levels of IFN-gamma much earlier after induction than fresh lymphocytes. Quantitation of the steady-state levels of IFN-gamma mRNA showed these to differ markedly between the two cell types over a period of time post-induction. In fresh lymphocytes the steady-state levels of IFN-gamma mRNA increased to a peak level over a period of 4 days while in growing lymphoblasts the peak level occurred after 8 hours. These differences in IFN-gamma mRNA production were shown to be not the result of gross alteration of RNA metabolism following blast transformation.

Cells, Cultured↗
PubMed1983

Translation of mRNA for Limulus polyphemus haemocyanin polypeptides in vitro: studies on subunit heterogeneity.

The haemocyanin of Limulus polyphemus is composed of a number (possibly 10-15) of polypeptides and is believed to be synthesised in cells called cyanoblasts. In vitro translation in the rabbit reticulocyte haemolysate system and in Xenopus oocytes, of mRNA isolated from cyanoblast-containing tissue, allowed the detection of several haemocyanin polypeptides amongst the products of translation. At least seven polypeptides with molecular weights in the range 68 000-71 000 were identified by an immunological method followed by electrophoretic characterisation on two-dimensional polyacrylamide gels. Comparison of the polypeptide patterns of authentic haemocyanin, reticulocyte lysate translation products and Xenopus oocyte translation products led to the conclusion that the polypeptides are unlikely to undergo significant post-translational modification or to possess cleavable signal sequences. It is proposed that release of haemocyanin into the haemolymph in vivo may involve bursting of the cyanoblasts.

Animals↗