Modified Whole-Mount In situ Hybridization Protocol for the Detection of Transgene Expression in Electroporated Chick Embryos

hybridization. hybridization (WISH).Here we describe a modification to the WISH protocol that is essential to prevent DNA cross-hybridization and to specifically detect transgene mRNA transcripts in electroporated embryos. Our optimized WISH procedure can be applied not only to electroporated chick embryos but also to other embryos or adult tissues that have been transfected with large amounts of reporter- or expression construct DNA

Geographic clustering of testicular cancer incidence in the northern part of The Netherlands

Geographic variations in testicular cancer incidence may be caused by differences in environmental factors, genetic factors, or both. In the present study, geographic patterns of age-adjusted testicular cancer incidence rates (IRs) in 12 provinces in The Netherlands in the period 1989–1995 were analysed. In addition, the age-adjusted IR of testicular cancer by degree of urbanization was evaluated. Cancer incidence data were obtained from the Netherlands Cancer Registry. The overall annual age-adjusted IR of testicular cancer in The Netherlands in the period 1989–1995 was 4.4 per 100 000 men. The province Groningen in the north of the country showed the highest annual IR with 5.8 per 100 000 men, which was higher (P < 0.05) than the overall IR in The Netherlands (incidence rate ratio (IRR) 1.3, 95% confidence interval (CI) 1.1–1.6). The highest IR in Groningen was seen for both seminomas and non-seminomas. In addition, Groningen showed the highest age-specific IRs in all relevant younger age groups (15–29, 30–44 and 45–59 years), illustrating the consistency of data. The province Friesland, also situated in the northern part of the country, showed the second highest IR of testicular cancer with 5.3 cases per 100 000 men per year (IRR 1.2, 95% Cl 1.0–1.5, not significant). This mainly resulted from the high IR of seminoma in Friesland. Analysis of age-adjusted IRs of testicular cancer by degree of urbanization in The Netherlands showed no urban–rural differences at analysis of all histological types combined, or at separate analyses of seminomas and non-seminomas. Geographic clustering of testicular cancer seems to be present in the rural north of The Netherlands with some stable founder populations, which are likely to share a relatively high frequency of genes from common ancestors including genes possibly related to testicular cancer. Although this finding does not exclude the involvement of shared environmental factors in the aetiology of testicular cancer, it may also lend support to a genetic susceptibility to testicular cancer development. Testicular cancer cases in stable founder populations seem particularly suitable for searching for testicular cancer susceptibility genes because such genes are likely to be more frequent among affected men in such populations. © 1999 Cancer Research Campaig

Dietary perturbations alter the ecological significance of ingested Lactobacillus plantarum in the digestive tract

Host diet is a major determinant of the composition and function of the intestinal microbiome. Less understood is the importance of diet on ingested strains with probiotic significance. We investigated the population dynamics of exogenous Lactobacillus plantarum and its interactions with intestinal bacteria in mice undergoing switches between high-fat, high-sugar (HFHSD) and low-fat, plant-polysaccharide rich (LFPPD) diets. The survival and persistence of ingested L. plantarum WCFS1 was significantly improved during mouse consumption of HFHSD and was negatively associated with the numbers of indigenous Lactobacillus species. Diet also rapidly changed the composition of the indigenous microbiota, but with some taxa differentially affected between HFHSD periods. L. plantarum was not integrated into indigenous bacterial community networks according to co-occurrence patterns but still conferred distinct effects on bacterial species diversity and microbiota stability largely in a diet-dependent manner. Metagenome predictions supported the premise that L. plantarum dampens the effects of diet on the microbiome. This strain also consistently altered the predicted genetic content in the distal gut by enriching for genes encoding glyosyltransferases and bile salt hydrolases. Our findings demonstrate the interactions between ingested, transient probiotic bacteria and intestinal bacterial communities and how they can differ depending on host diet

The use of electroporation in developmental biology

Chapter 139-1During the formation of a complex organism, cells divide, die, migrate, and differentiate. Biologists have established tools to observe those phenomena but also to change their course, which subsequently enables to infer causal relationships between various events occurring in different cell groups. More precisely, present approaches mostly rely on modifications of gene expression. For instance, cells are labeled with fluorescent proteins and tracked within the embryo, molecular signals are switched on and off to perturb regulatory pathways. Importantly, in all those experiments, the exogenous genetic material must be delivered at the right place and with the appropriate timing: requirements that can both be fulfilled by electroporation. After 15 years of constant refinement, this technique has now superseded methods like viral infection, microinjection, and lipofection. Applications encompass a large number of model organisms, targeted anatomical structures, and molecular biology techniques