Applications of molecular markers to forest genetics: Genetic diversity, genetic linkage mapping, and gene expression.

摘要

This thesis is comprised of four studies utilizing molecular genetic markers with coniferous trees. PCR and direct sequencing, RAPD markers, and RFLP markers were used in studies of gene expression (mRNA editing), genetic linkage mapping, and genetic diversity. In the first study, direct sequencing of PCR products was used to demonstrate the existence of RNA editing in the mitochondria of western red cedar (Thuja plicata Donn ex D. Don). This was the first demonstration of this phenomenon in a gymnosperm. The next two studies involved the use of RAPD markers for genetic linkage mapping in conifers. In the first of these, RAPD markers were shown to segregate in a Mendelian fashion among F1 progeny of Douglas-fir (Pseudotsuga menziesii (Mirb) Franco), and hence to be useful as genetic markers in conifers. A testcross mapping strategy was proposed to allow the efficient construction of genetic linkage maps in diploid organisms such as trees using these dominant markers. The feasibility of this strategy was demonstrated in Douglas-fir. This was the first study using RAPD markers in a conifer. An alternative mapping strategy, taking advantage of the availability of haploid megagametophyte tissue in conifers, was explored in the next study. A partial genetic linkage map was constructed for a single white spruce (Picea glauca (Moench) Voss) tree by following the segregation of RAPD markers among a set of megagametophytes obtained from seeds of that tree. It was shown that the disadvantage of the dominance of RAPD markers could be overcome by using this haploid tissue as a DNA source. The approach to genetic linkage mapping first taken in this study is now commonly used by conifer geneticists. The final study was on DNA-level genetic diversity in western red cedar. Previous results with isozyme markers and terpenes had suggested that genetic diversity is relatively low in this species. Single or low copy number nuclear RFLP markers were developed to further explore this question. The resulting estimate of species-wide genetic diversity (expected heterozygosity) closely agreed with those obtained from isozyme studies. This was the first large scale population genetics study in a conifer using single or low copy number nuclear RFLP markers.