| Summary: | 博士 === 國立臺灣大學 === 生態學與演化生物學研究所 === 101 === Non-photosynthetic plants only retain remnant plastids and their plastome is highly reduced. Mitrastemon kanehirai, a root holoparasite, is endemic to Taiwan and considered an endangered species. Mitrastemon kanehirai has only one plastid sequence has been reported, and the sequence, pt16S rDNA, shows increased substitution rate.
In this dissertation, the performance of six DNA extraction procedures for two non-photosynthetic plants, Balanophora japonica and M. kanehirai, were compared. All six procedures yielded DNA of sufficient quality for PCR, and the method described by Barnwell et al. (1998) performed well in isolating DNA from both species for restriction enzyme digestion. Meanwhile, enrichment of M. kanehirai plastid DNA content was achieved by using the ‘high salt’ methods based on protocol presented by Milligan (1989).
High rate of nucleotide substitution in three subcellular SSU rDNAs have been reported in heterotrophic plants, and the rate heterogeneity among these sequences are presented in this dissertation. Mt19S, pt16S and nr18S rDNA sequences from nine heterotrophic plants, including one hemiparasitic, five holoparasitic and three mycoheterotrophic plants, were examined. Rate heterogeneity among various rDNA sequences was evaluated by relative rate tests and phylogenetic analysis. Both pt16S and nr18S rDNA sequences of non-photosynthetic species show significant increases of substitution rate, but the phenomenon was not found in mt19S rDNA. The extreme divergent pt16S and nr18S rDNA sequences were found in B. japonica and M. kanehirai, and accompanied by a decrease in GC content of the sequences.
Mitrastemon kanehieai plastome was sequenced by using next generation sequencing technology. The genome is smallest plastome that have been described with size of 25,740 bp. Only 26 genes were retained in the plastome, which include 4 rRNAs, 4 tRNAs and 18 protein-coding genes. These retained genes are mostly involved in translation machinery. All photosynthesis-related genes were lost, and the inverted repeat region is absent. Despite the enormous reduction, the M. kanehirai plastome is a functional gene expression system. DNA transfer from plastid to nucleus and horizontal transfer from the host to the parasite were also observed in M. kanehirai.
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