| Summary: | 碩士 === 國立成功大學 === 生物科技研究所碩博士班 === 99 === Mitochondria are endosymbiotic from α-proteobacteria during evolution. Mammalian mitochondrial genomes are compact with approximately 16 kb in length; in contrast, plant mitochondria genomes are diverse and range from 200 to 2,400 kb in size. Up to date, according to NCBI database, only 25 species of land plants, their mitochondrial genome have been completely determined. Phalaenopsis orchid is a fascinating flower and an economically important floral crop in Taiwan. There are two native Phalaenopsis orchid species in Taiwan, P. aphrodite subsp. formosana and P. equestris, which they have been commonly used in commercial breeding program. However, the mitochondria and their genome were less studied. In this study, we have screened the high density filters which contained 41,472 Bacterial Artificial Chromosome (BAC) clones of P. equestris with 19 probes of heterologous mitochondrial DNA and 9 probes of BAC end fragments. One hundred seventy seven BAC clones which potentially contain the mitochondrial DNA were selected. Thirty nine mitochondrial DNA containing BAC clones were further identified with 7 individual probes by Dot-blots. The insert sizes of BAC clones were determined to be 80 to 100 kb in average by PFGE (pulsed field gel electrophoresis). RFLP (restriction fragment length polymorphism) analyses and cross hybridization were used to cluster 39 BAC clones into four major overlapping groups. Four selected BAC clones were further sequenced by Solexa system. BAC-25 (82 kb) which contained 6 mitochondrial genes and 1 pseudogene was identified. The rps10 is present in BAC-25, but not found in the sequenced mitochondrial genomes of monocots so far. In addition, BAC-25 had two identical 2.7 kb regions which each consist of 6 chloroplast genes of P. equestris. Furthermore, BAC-25 contained very abundant of small repetitive sequences (19~20 nt) and 2 pairs of large directed repeats (＞12 kb). Therefore, it is very likely that BAC-25 is part of mitochondrial genome of P. equestris.
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