Characterization of T‐DNA insertions in putative mitochondrial‐targeted DNA polymerase, SSB, and topoisomerase II genes in Arabidopsis thaliana

Although the mitochondrial genome of Arabidopsis thaliana has been sequenced in full, many questions remain regarding gene expression and genome replication. RecA and SSB gene homologues have been identified and shown to be targeted specifically to mitochondria. One question currently being investigated is the role of other proteins in mitochondrial DNA recombination, including the products of the DNA polymerase, SSB, and topoisomerase II genes. These proteins play important roles in DNA recombination in other organisms and homologous genes with putative targeting to mitochondria have been identified in Arabidopsis thaliana. Seeds with T‐DNA insertions in the putative mitochondrial‐targeted DNA polymerase, SSB, and topoisomerase II genes have been obtained. These seeds are allowed to germinate and grow, and are then compared phenotypically to plants grown from wild‐type seeds. Such comparisons are made with plants grown on a normal medium and on a medium containing the DNA damaging chemical MMS. Once variations in the phenotypes are observed it is necessary to utilize PCR to confirm the location of the T‐DNA insert within the gene. Other experiments include the use of electron microscopy to observe any changes in mitochondrial structure and morphology and analysis of the total amount of mitochondrial DNA by quantitative PCR. Analysis of the differences in the plants will allow for several conclusions to be drawn about the importance of each of these proteins to recombination of the mitochondrial DNA of Arabidopsis thaliana.