Biomedical subjects

Michael V Wiles

Publications and source records attributed to Michael V Wiles.

5 recordsLinked to original sources

PubMed2006

Know thy mouse.

In science it is imperative that the basic reagents and materials are defined and uniform. Unlike chemical reagents, which are uniform over time, mice, like all living creatures, have an intrinsic genetic drive to change, with mutations accumulating over time leading to increasing genetic variation and phenotypic change. Such changes compromise the reproducibility of experimental data over time and place. The use of the mouse has expanded rapidly in recent years and many scientists who have turned to the mouse as a research model might be unaware of the profound impact of changes in genetic background. Here we discuss the sources of genetic change and strategies to reduce them with the idea of strengthening international genetic standards for inbred mice.

Animals↗

PubMed2006

Peptide motifs of the single dominantly expressed class I molecule explain the striking MHC-determined response to Rous sarcoma virus in chickens.

Compared with the MHC of typical mammals, the chicken MHC is smaller and simpler, with only two class I genes found in the B12 haplotype. We make five points to show that there is a single-dominantly expressed class I molecule that can have a strong effect on MHC function. First, we find only one cDNA for two MHC haplotypes (B14 and B15) and cDNAs corresponding to two genes for the other six (B2, B4, B6, B12, B19, and B21). Second, we find, for the B4, B12, and B15 haplotypes, that one cDNA is at least 10-fold more abundant than the other. Third, we use 2D gel electrophoresis of class I molecules from pulse-labeled cells to show that there is only one heavy chain spot for the B4 and B15 haplotypes, and one major spot for the B12 haplotype. Fourth, we determine the peptide motifs for B4, B12, and B15 cells in detail, including pool sequences and individual peptides, and show that the motifs are consistent with the peptides binding to models of the class I molecule encoded by the abundant cDNA. Finally, having shown for three haplotypes that there is a single dominantly expressed class I molecule at the level of RNA, protein, and antigenic peptide, we show that the motifs can explain the striking MHC-determined resistance and susceptibility to Rous sarcoma virus. These results are consistent with the concept of a "minimal essential MHC" for chickens, in strong contrast to typical mammals.

Amino Acid Motifs↗

PubMed2004

Development of a SNP genotyping panel for genetic monitoring of the laboratory mouse.

We have developed a genotyping system for detecting genetic contamination in the laboratory mouse based on assaying single-nucleotide polymorphism (SNP) markers positioned on all autosomes and the X chromosome. This system provides a fast, reliable, and cost-effective way for genetic monitoring, while maintaining a very high degree of confidence. We describe the allelic distribution of 235 SNPs in 48 mouse strains, thereby creating a database of polymorphisms useful for genotyping purposes. The SNP markers used in this study were chosen from publicly available SNP databases. Four genotyping methods were evaluated, and dynamic two-tube allele-specific PCR assays were developed for each marker and tested on a set of 48 inbred mouse strains. The minimal number of assays sufficient to distinguish groups consisting of different numbers of mouse strains was estimated, and a panel of 28 SNPs sufficient to distinguish virtually all of the inbred strains tested was selected. Amplifluor SNP detection assays were developed for these markers and tested on an extended list of 96 strains. This panel was used as a genetic quality control approach to monitor the genotypes of nearly 300 inbred, wild-derived, congenic, consomic, and recombinant inbred strains maintained at The Jackson Laboratory. We have concluded that this marker panel is sufficient for genetic contamination monitoring in colonies containing a large number of genetically diverse mouse strains and that reduced versions of the panel could be implemented in facilities housing a lower number of strains.

Alleles↗

PubMed2004

An efficient SNP system for mouse genome scanning and elucidating strain relationships.

A set of 1638 informative SNP markers easily assayed by the Amplifluor genotyping system were tested in 102 mouse strains, including the majority of the common and wild-derived inbred strains available from The Jackson Laboratory. Selected from publicly available databases, the markers are on average approximately 1.5 Mb apart and, whenever possible, represent the rare allele in at least two strains. Amplifluor assays were developed for each marker and performed on two independent DNA samples from each strain. The mean number of polymorphisms between strains was 608+/-136 SD. Several tests indicate that the markers provide an effective system for performing genome scans and quantitative trait loci analyses in all but the most closely related strains. Additionally, the markers revealed several subtle differences between closely related mouse strains, including the groups of several 129, BALB, C3H, C57, and DBA strains, and a group of wild-derived inbred strains representing several Mus musculus subspecies. Applying a neighbor-joining method to the data, we constructed a mouse strain family tree, which in most cases confirmed existing genealogies.

Alleles↗

PubMed2003

The properties of the single chicken MHC classical class II alpha chain ( B-LA) gene indicate an ancient origin for the DR/E-like isotype of class II molecules.

In mammals, there are MHC class II molecules with distinctive sequence features, such as the classical isotypes DR, DQ and DP. These particular isotypes have not been reported in non-mammalian vertebrates. We have isolated the class II (B-L) alpha chain from outbred chickens as the basis for the cloning and sequencing of the cDNA. We found only one class II alpha chain transcript, which bears the major features of a classical class II alpha sequence, including the critical peptide-binding residues. The chicken sequence is more similar to human DR than to the DQ, DP, DO or DM isotypes, most significantly in the peptide-binding alpha(1) domain. The cDNA and genomic DNA sequences from chickens of diverse origins show few alleles, which differ in only four nucleotides and one amino acid. In contrast, significant restriction fragment length polymorphism is detected by Southern blot analysis of genomic DNA, suggesting considerable diversity around the gene. Analysis of a large back-cross family indicates that the class II alpha chain locus ( B-LA) is located roughly 5.6 cM from the MHC locus, which encodes the classical class II beta chains. Thus the chicken class II alpha chain gene is like the mammalian DR and E isotypes in three properties: the presence of the critical peptide-binding residues, the low level of polymorphism and sequence diversity, and the recombinational separation from the class II beta chain genes. These results indicate that the sequence features of this lineage are both functionally important and at least 300 million years old.

Alleles↗