Methods in Alcohol-Related Neuroscience Research PDF

INTRODUCTION

Complex traits, such as the human disease alcoholism, are influenced by multiple genetic and environmental factors. Animals can be used to model certain components of this disease. However, even the apparently simplified traits modeled in animals are found to be under the influence of multiple genes, each of relatively small effect. Thus, the distributions of these traits in genetically heterogeneous populations tend to be continuous, rather than displaying simple Mendelian inheritance patterns. The genomic loci that influence the quantitative variability in these traits are termed quantitative trait loci (QTL).* The identification of the genes influencing such quantitative traits can be approached using QTL mapping methods that were originally developed by plant geneticists (see Lynch and Walsh, 1998). Initial efforts may be directed at localizing an influential gene (a QTL) to a chromosome or a chromosomal segment. Additional experiments may then be performed, using stringent statistical criteria, to confirm the presence of the QTL. Finer mapping methods are then used to narrow the chromosomal region in which the QTL resides, with the ultimate goal of identifying the causative gene or regulatory element. Single gene mutants, gene expression analyses and sequence information can be used at each stage to provide evidence that a candidate gene is the QTL.

QTL mapping depends on the availability of genetic markers with known positions on particular chromosomes. When many such markers are polymorphic between strains, it is possible to determine which chromosome segments were inherited from which ancestors. Multiple markers are required for each chromosome because of crossing-over, a process that intermixes chromosome segments during meiosis. By examining many markers on each chromosome, the relationship between variability in the phenotype of interest and variability in genetic material can be determined — the essence of QTL mapping. Genetic markers are not selected based on any functional significance, but rather because their differences between strains are easily identifiable. In general, these markers correspond to non-coding regions of the genome, meaning that they are not actual genes. The most commonly used markers are regions whose PCR products are of different lengths for different mouse strains. If these concepts are unfamiliar, more thorough discussions are available (Grisel and Crabbe, 1995; Falconer and Mackay, 1996; Lynch and Walsh, 1998).

QTL mapping approaches require numerous comparisons between phenotype and genotype. When so many comparisons are made, the criteria for statistical significance must be stringent to avoid false positive errors. These stringent criteria diminish the power to detect real effects, increasing the probability of false negative errors. Thus, every QTL study grapples with the need to balance these two diametrically opposed considerations. The result is that many hundreds or even a few thousand animals are required to accurately map the location of one or more QTL for a particular trait. Frequently, multiple experiments must be performed, first to provisionally map QTL, and subsequently to confirm and more accurately map their location.

The goal of the current chapter is to serve as an introductory practical guide to the most relevant methods, technologies and approaches to QTL mapping studies with specific reference to alcohol-related basic research. Whenever possible, citations will be given to original theoretical reports, review articles, World Wide Web links and relevant findings based on these methodologies.