Ki Joong Kim

A DNA barcode for land plants

DNA barcoding involves sequencing a standard region of DNA as a tool for species identification. However, there has been no agreement on which region(s) should be used for barcoding land plants. To provide a community recommendation on a standard plant barcode, we have compared the performance of 7 leading candidate plastid DNA regions (atpF–atpH spacer, matK gene, rbcL gene, rpoB gene, rpoC1 gene, psbK–psbI spacer, and trnH–psbA spacer). Based on assessments of recoverability, sequence quality, and levels of species discrimination, we recommend the 2-locus combination of rbcL+matK as the plant barcode. This core 2-locus barcode will provide a universal framework for the routine use of DNA sequence data to identify specimens and contribute toward the discovery of overlooked species of land plants.

Phylogenetic implications of rbcL sequence variation in the Asteraceae

Complete nucleotide sequences of the rbcL gene were obtained for 25 species of Asteraceae representing 15 of the currently recognized tribes and three outgroup families. A total of 345 variable nucleotide positions was identified, 170 of which were phylogenetically informative. Phylogenetic analyses of the rbcL data generated eight equally parsimonious trees with a consistency index of 0.47. Three major monophyletic clades that correspond to the subfamilies Barnadesioideae, Cichorioideae, and Asteroideae were identified in the most parsimonious cladograms; however, support for these groups was not strong (...)

Complete chloroplast genome sequences of important oilseed crop Sesamum indicum L.

Sesamum indicum is an important crop plant species for yielding oil. The complete chloroplast (cp) genome of S. indicum (GenBank acc no. JN637766) is 153,324 bp in length, and has a pair of inverted repeat (IR) regions consisting of 25,141 bp each. The lengths of the large single copy (LSC) and the small single copy (SSC) regions are 85,170 bp and 17,872 bp, respectively. Comparative cp DNA sequence analyses of S. indicum with other cp genomes reveal that the genome structure, gene order, gene and intron contents, AT contents, codon usage, and transcription units are similar to the typical angiosperm cp genomes. Nucleotide diversity of the IR region between Sesamum and three other cp genomes is much lower than that of the LSC and SSC regions in both the coding region and noncoding region. As a summary, the regional constraints strongly affect the sequence evolution of the cp genomes, while the functional constraints weakly affect the sequence evolution of cp genomes. Five short inversions associated with short palindromic sequences that form step-loop structures were observed in the chloroplast genome of S. indicum. Twenty-eight different simple sequence repeat loci have been detected in the chloroplast genome of S. indicum. Almost all of the SSR loci were composed of A or T, so this may also contribute to the A-T richness of the cp genome of S. indicum. Seven large repeated loci in the chloroplast genome of S. indicum were also identified and these loci are useful to developing S. indicum-specific cp genome vectors. The complete cp DNA sequences of S. indicum reported in this paper are prerequisite to modifying this important oilseed crop by cp genetic engineering techniques.

Complete chloroplast DNA sequence from a Korean endemic genus, Megaleranthis saniculifolia , and its evolutionary implications

The chloroplast DNA sequences of Megaleranthis saniculifolia, an endemic and monotypic endangered plant species, were completed in this study (GenBank FJ597983). The genome is 159,924 bp in length. It harbors a pair of IR regions consisting of 26,608 bp each. The lengths of the LSC and SSC regions are 88,326 bp and 18,382 bp, respectively. The structural organizations, gene and intron contents, gene orders, AT contents, codon usages, and transcription units of the Megaleranthis chloroplast genome are similar to those of typical land plant cp DNAs. However, the detailed features of Megaleranthis chloroplast genomes are substantially different from that of Ranunculus, which belongs to the same family, the Ranunculaceae. First, the Megaleranthis cp DNA was 4,797 bp longer than that of Ranunculus due to an expanded IR region into the SSC region and duplicated sequence elements in several spacer regions of the Megaleranthis cp genome. Second, the chloroplast genomes of Megaleranthis and Ranunculus evidence 5.6% sequence divergence in the coding regions, 8.9% sequence divergence in the intron regions, and 18.7% sequence divergence in the intergenic spacer regions, respectively. In both the coding and noncoding regions, average nucleotide substitution rates differed markedly, depending on the genome position. Our data strongly implicate the positional effects of the evolutionary modes of chloroplast genes. The genes evidencing higher levels of base substitutions also have higher incidences of indel mutations and low Ka/Ks ratios. A total of 54 simple sequence repeat loci were identified from the Megaleranthis cp genome. The existence of rich cp SSR loci in the Megaleranthis cp genome provides a rare opportunity to study the population genetic structures of this endangered species. Our phylogenetic trees based on the two independent markers, the nuclear ITS and chloroplast matK sequences, strongly support the inclusion of the Megaleranthis to the Trollius. Therefore, our molecular trees support Ohwi’s original treatment of Megaleranthis saniculiforia to Trollius chosenensis Ohwi.

Autocatalytic Processing of HtrA2/Omi Is Essential for Induction of Caspase-dependent Cell Death through Antagonizing XIAP

A mature form of nuclear-encoded mitochondrial serine protease HtrA2/Omi is pivotal in regulating apoptotic cell death; however, the underlying mechanism of the processing event of HtrA2/Omi and its relevant biological function remain to be clarified. Here, we describe that HtrA2/Omi is autocatalytically processed to the 36-kDa protein fragment, which is required for the cytochrome c-dependent caspase activation along with neutralizing XIAP-mediated inhibition of caspases through interaction with XIAP, eventually promoting apoptotic cell death. We have shown that the autocatalytic processing of HtrA2/Omi occurs via an intermolecular event, demonstrated by incubating an in vitro translated HtrA2/Omi (S306A) mutant with the enzymatically active glutathione S-transferase-HtrA2/Omi protein. Using N-terminal amino acid sequencing and mutational analysis, we identified that the autocatalytic cleavage site is the carboxyl side of alanine 133 of HtrA2/Omi, resulting in exposure of an inhibitor of apoptosis protein binding motif in its N terminus. Our study provides evidence that the autocatalytic processing of HtrA2/Omi is crucial for regulating HtrA2/Omi-mediated apoptotic cell death.

Genetic Diversity in the Common Terrestrial Orchid Oreorchis patens and Its Rare Congener Oreorchis coreana: Inference of Species Evolutionary History and Implications for Conservation

We hypothesized that the main Korean mountain ranges provided many refugia for boreal plant species, where they likely found relatively stable habitats and maintained large population sizes. Under this scenario, high levels of genetic variation and low degree of differentiation among populations within these species were anticipated. To test this hypothesis, we examined levels of allozyme diversity (17 loci) in 12 populations of the common terrestrial montane orchid Oreorchis patens from the main ranges in Korea and 4 populations of its rare congener O. coreana, which is restricted to the Korean island of Jeju. As expected, O. patens harbored high levels of genetic variation within populations (%P = 62.8, A = 1.96, H (o) = 0.211, and H (e) = 0.237). Allele frequency differences among populations were low (F (ST) = 0.075), and the species also displayed a significant correlation between pairwise genetic differentiation and geographical distance. All these results suggest that extant populations were founded by multiple genetically diverse individuals and that most of this initial diversity would have been maintained in the stable mountainous conditions during Quaternary climatic oscillations. In contrast, we were unable to detect any genetic diversity in O. coreana, suggesting that contemporary populations likely originated from a single ancestral source population that had lost all genetic variability. From a long-term conservation genetics perspective, extreme rarity and small population sizes, coupled with its apparent genetic uniformity, place O. coreana at a high risk of extinction. Thus, both in situ and ex situ conservation efforts should be of particular importance for this species.

Chloroplast Genome Evolution in Early Diverged Leptosporangiate Ferns

In this study, the chloroplast (cp) genome sequences from three early diverged leptosporangiate ferns were completed and analyzed in order to understand the evolution of the genome of the fern lineages. The complete cp genome sequence of Osmunda cinnamomea (Osmundales) was 142,812 base pairs (bp). The cp genome structure was similar to that of eusporangiate ferns. The gene/intron losses that frequently occurred in the cp genome of leptosporangiate ferns were not found in the cp genome of O. cinnamomea. In addition, putative RNA editing sites in the cp genome were rare in O. cinnamomea, even though the sites were frequently predicted to be present in leptosporangiate ferns. The complete cp genome sequence of Diplopterygium glaucum (Gleicheniales) was 151,007 bp and has a 9.7 kb inversion between the trnL-CAA and trnV-GCA genes when compared to O. cinnamomea. Several repeated sequences were detected around the inversion break points. The complete cp genome sequence of Lygodium japonicum (Schizaeales) was 157,142 bp and a deletion of the rpoC1 intron was detected. This intron loss was shared by all of the studied species of the genus Lygodium. The GC contents and the effective numbers of co-dons (ENCs) in ferns varied significantly when compared to seed plants. The ENC values of the early diverged leptosporangiate ferns showed intermediate levels between eusporangiate and core leptosporangiate ferns. However, our phylogenetic tree based on all of the cp gene sequences clearly indicated that the cp genome similarity between O. cinnamomea (Osmundales) and eusporangiate ferns are symplesiomorphies, rather than synapomorphies. Therefore, our data is in agreement with the view that Osmundales is a distinct early diverged lineage in the leptosporangiate ferns.

The complete chloroplast DNA sequence of Eleutherococcus senticosus (Araliaceae); comparative evolutionary analyses with other three asterids.

This study reports the complete chloroplast (cp) DNA sequence of Eleutherococcus senticosus (GenBank: JN 637765), an endangered endemic species. The genome is 156,768 bp in length, and contains a pair of inverted repeat (IR) regions of 25,930 bp each, a large single copy (LSC) region of 86,755 bp and a small single copy (SSC) region of 18,153 bp. The structural organization, gene and intron contents, gene order, AT content, codon usage, and transcription units of the E. senticosus chloroplast genome are similar to that of typical land plant cp DNA. We aligned and analyzed the sequences of 86 coding genes, 19 introns and 113 intergenic spacers (IGS) in three different taxonomic hierarchies; Eleutherococcus vs. Panax, Eleutherococcus vs. Daucus, and Eleutherococcus vs. Nicotiana. The distribution of indels, the number of polymorphic sites and nucleotide diversity indicate that positional constraint is more important than functional constraint for the evolution of cp genome sequences in Asterids. For example, the intron sequences in the LSC region exhibited base substitution rates 5–11-times higher than that of the IR regions, while the intron sequences in the SSC region evolved 7–14-times faster than those in the IR region. Furthermore, the Ka/Ks ratio of the gene coding sequences supports a stronger evolutionary constraint in the IR region than in the LSC or SSC regions. Therefore, our data suggest that selective sweeps by base collection mechanisms more frequently eliminate polymorphisms in the IR region than in other regions. Chloroplast genome regions that have high levels of base substitutions also show higher incidences of indels. Thirty-five simple sequence repeat (SSR) loci were identified in the Eleutherococcus chloroplast genome. Of these, 27 are homopolymers, while six are di-polymers and two are tri-polymers. In addition to the SSR loci, we also identified 18 medium size repeat units ranging from 22 to 79 bp, 11 of which are distributed in the IGS or intron regions. These medium size repeats may contribute to developing a cp genome-specific gene introduction vector because the region may use for specific recombination sites.

Phylogenetic and Evolutionary Implications of Interspecific Chloroplast DNA Variation in Krigia (Asteraceae-Lactuceae)

Chloroplast DNA restriction site and length variation was examined among 12 taxa of Krigia, using 34 restriction endonucleases, filter hybridization experiments, and comparative mapping procedures. A total of 252 restriction site mutations was detected, 159 of which were phylogenetically informative. The resulting phylogenetic trees correlated well with cytological and morphological data. The two sects. Krigia and Cymbia differed by at least 73 restriction site mutations. Most species within each section were characterized by many restriction site changes. Interspecific sequence divergence in Krigia was much higher than in any other previously studied genus of the Asteraceae and was comparable to the high levels of divergence found in genera of other angiosperm families. Relative rates of chloroplast DNA evolution in Krigia were significantly different among the various chromosome number groups, resulting in the rejection of a molecular clock. However, a molecular clock could not be rejected within each chromosome number group. Different regions of the chloroplast genome also showed extremely different frequencies of mutations and four hotspot areas were identified. The areas occurred in long spacer regions between gene coding sequences. Furthermore, most of the homoplasy and length variation also occurred within these hotspots. Chloroplast DNA trees were consistent with the hypothesis that an ancient allopolyploid event between species with low chromosome base numbers, followed by subsequent aneuploid reduction, resulted in K. wrightii with n = 9. The results also suggest that x = 5 is the probable ancestral base number for the genus and that aneuploid reduction is the major mode of chromosome number change in Krigia and related genera.

A chloroplast DNA phylogeny of lilacs (Syringa, Oleaceae): Plastome groups show a strong correlation with crossing groups

Phylogenetic relationships and genomic compatibility were compared for 60 accessions of Syringa using chloroplast DNA (cpDNA) and nuclear ribosomal DNA (rDNA) markers. A total of 669 cpDNA variants, 653 of which were potentially phylogenetically informative, was detected using 22 restriction enzymes. Phylogenetic analyses reveal four strongly supported plastome groups that correspond to four genetically incompatible crossing groups. Relationships of the four plastome groups (I(II(III,IV))) correlate well with the infrageneric classification except for ser. Syringa and Pinnatifoliae. Group I, which includes subg. Ligustrina, forms a basal lineage within Syringa. Group II includes ser. Syringa and Pinnatifoliae and the two series have high compatibility and low sequence divergence. Group III consists of three well-defined species groups of ser. Pubescentes. Group IV comprises all members of ser. Villosae and has the lowest interspecific cpDNA sequence divergences. Comparison of cpDNA sequence divergence with crossability data indicates that hybrids have not been successfully generated between species with divergence greater than 0.7%. Hybrid barriers are strong among the four major plastome groups, which have sequence divergence estimates ranging from 1.096 to 1.962%. In contrast, fully fertile hybrids occur between species pairs with sequence divergence below 0.4%. Three regions of the plastome have length variants of greater than 100 bp, and these indels identify 12 different plastome types that correlate with phylogenetic trees produced from cpDNA restriction site data. Biparentally inherited nuclear rDNA and maternally inherited cpDNA length variants enable the identification of the specific parentage of several lilac hybrids.