Dispersed repetitive sequences in the chloroplast genome of Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/3f4628805

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  • Chloroplast genomes of conifers are unusual in a number of respects, including the possession of much repetitive DNA. By studying the distribution and sequences of dispersed repetitive DNA, I hoped to gain insight into the mechanisms of chloroplast genome evolution. I used restriction mapping and DNA sequencing to characterize dispersed repetitive DNA in the chloroplast genome of Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco). A chloroplast DNA clone bank was constructed in a pUC19 plasmid vector. High stringency hybridization among these recombinant plasmids, and to restricted chloroplast DNA, was used to map repeat families. Dispersed repeats were grouped into three classes: long repeat families (around 600 bp), medium repeat families, and short repeat families (10-20 bp). The long and medium repeats were classified into a total of six families and placed on a restriction site map. Sequence analysis of one repeat family shared among three restriction fragments revealed the presence a 633 by inverted repeat. The repeat contains a complete transfer RNA-Serine (GCU) gene and an open reading frame (ORF36). This open reading frame is highly homologous with chloroplast sequences in tobacco and liverwort. Both ends of the repeat possess transposon-like structures. Two copies of the dispersed repeats are located near the endpoints of a large inversion that distinguishes Douglas-fir and radiata pine. The third copy of the repeat is also located near the endpoint of an inversion that occurred during conifer evolution, and is associated with a length mutation hotspot, and short inverted repeats; the latter appear to be capable of forming numerous stem-loop structures. Sequence comparisons with tobacco cpDNA revealed that one of the Douglas-fir chloroplast DNA clones is homologous to three tobacco chloroplast DNA segments. These regions in tobacco appear to have been juxtaposed in Douglas-fir by a number of inversions and a small deletion. An 8 by (CATCTTTT) direct repeat in tobacco is sandwiched between two inverted sections in Douglas-fir. This repeat may be a target sequence for homologous recombination.
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