Genetic properties of RAPD markers and RNA editing in gymnosperms

This thesis is based on results presented in the six accompanying papers (designated I to VI). The first studies (Papers I and II), on RAPD variation in haploid and diploid tissues from individuals, controlled crosses, and populations of Pinus sylvestris (L.), showed that RAPD fragments have a high level of reproducibility. Most of RAPD fragments amplified were inherited as dominant Mendelian traits. Furthermore, the data showed that when complete genotype information is available. RAPD markers give estimates of genetic parameters similar to those obtained from allozyme data. However, the dominant character of RAPDs limits their usefulness for population genetic analysis of diploid material because gene frequencies must be derived indirectly assuming Hardy-Weinberg equilibrium, which may lead to biased estimates of population differentiation. Analysis described in Paper III involving re-amplification of RAPD bands and further Southern analysis of RAPD profiles revealed that individual RAPD fragments contain additional sequences, but the amount of these is usually too small to produce distortions in the expected segregation proportions. RAPD fragments amplified from P. sylvestris did not appear to be generated from repetitive sequences, and the absence of RAPD bands was not due to the absence of the corresponding sequences in the genome but due to lack of their amplification. Studies discussed in Papers IV to VI concentrated on evaluation of inheritance, evolution and variation of organellar DNA, as summarised in the following paragraph. The inheritance of mitochondrial (mt) DNA in Pinus was confirmed to be strictly maternal. However, the presence of chloroplast (cp) DNA in macrogametophytes and mtDNA in pollen suggests that in Pinus, the elimination of alternative mt- and cpDNA occurs after fertilization. RNA editing of the coxI gene is known to be generally more extensive in gymnosperms than in angiosperms, but the data showed that the coxI sequences of Larix sibirica and Ginkgo biloba were not edited, suggesting that these sequences originated from edited mitochondrial coxI transcripts. Furthermore, in contrast to earlier suggestions based on the analysis of angiosperm mtDNA, the results indicated that there was a high rate of nucleotide of the mt coxI in gymnosperms. Thus, by allowing for free, random accumulation of T - C substitutions, RNA editing can contribute to accelerated sequence divergence. The differences in substitution rate among different lineages may cause bias in phytogenetic analysis. Finally, because of the low number of informative characters, cDNAs may not be suitable for the phylogenetic analysis of closely related species.