Complete Chloroplast DNA Sequence Research Articles

Wasabi Gone Wild? Origin and Characterization of the Complete Plastomes of Ulleung Island Wasabi (Eutrema japonicum; Brassicaceae) and Other Cultivars in Korea.

Korean wasabi occurs naturally on the young oceanic, volcanic Ulleung Island off the east coast of the Korean Peninsula. Although the Ulleung Island wasabi is reported as Eutrema japonicum and has been suggested to be morphologically identical to cultivars in Korea, very little is known about its taxonomic identity and relationship with other cultivars. In this study, we sequenced the complete chloroplast DNA sequences of three naturally occurring Ulleung Island wasabi plants and six cultivars ('Daewang', 'Daruma', 'Micado', 'Orochi', 'Green Thumb', and 'Shogun') from continental Korea and determined the taxonomic identity of Korean wasabi on Ulleung Island. The size and organization of the complete chloroplast genomes of the nine accessions were nearly identical to those of previously reported wasabi cultivars. In addition, phylogenetic analysis based on the complete plastomes suggested that Ulleung Island wasabi most likely comprises various wasabi cultivars with three chlorotypes ('Shogun', 'Green Thumb', and a unique Chusan type). Based on the complete plastomes, we identified eight chlorotypes for the major wasabi cultivars and the Ulleung Island wasabi. Two major groups (1-'Mazuma' and 'Daruma', and 2-'Fujidaruma'/'Shimane No. 3'/Ulleung Island wasabi/five cultivars in Korea) were also identified based on mother line genealogical history. Furthermore, different types of variations (mutations, insertions/deletions (indels), mononucleotide repeats, and inversions) in plastomes were identified to distinguish different cultivar lines and five highly divergent hotspots. The nine newly obtained complete plastomes are valuable organelle genomic resources for species identification and infraspecific phylogeographic studies on wild and cultivated wasabi.

Genetic Analysis of Hexaploid Wheat (Triticum aestivum L.) Using the Complete Sequencing of Chloroplast DNA and Haplotype Analysis of the Wknox1 Gene.

The aim of the presented study is a genetic characterization of the hexaploid wheat Triticum aestivum L. Two approaches were used for the genealogical study of hexaploid wheats—the complete sequencing of chloroplast DNA and PCR-based haplotype analysis of the fourth intron of Wknox1d and of the fifth-to-sixth-exon region of Wknox1b. The complete chloroplast DNA sequences of 13 hexaploid wheat samples were determined: Free-threshing—T. aestivum subsp. aestivum, one sample; T. aestivum subsp. compactum, two samples; T. aestivum subsp. sphaerococcum, one sample; T. aestivum subsp. carthlicoides, four samples. Hulled—T. aestivum subsp. spelta, three samples; T. aestivum subsp. vavilovii jakubz., two samples. The comparative analysis of complete cpDNA sequences of 20 hexaploid wheat samples (13 samples in this article plus 7 samples sequenced in this laboratory in 2018) was carried out. PCR-based haplotype analysis of the fourth intron of Wknox1d and of the fifth-to-sixth exon region of Wknox1b of all 20 hexaploid wheat samples was carried out. The 20 hexaploid wheat samples (13 samples in this article plus 7 samples in 2018) can be divided into two groups—T. aestivum subsp. spelta, three samples and T. aestivum subsp. vavilovii collected in Armenia, and the remaining 16 samples, including T. aestivum subsp. vavilovii collected in Europe (Sweden). If we take the cpDNA of Chinese Spring as a reference, 25 SNPs can be identified. Furthermore, 13–14 SNPs can be identified in T. aestivum subsp. spelta and subsp. vavilovii (Vav1). In the other samples up to 11 SNPs were detected. 22 SNPs are found in the intergenic regions, 2 found in introns, and 10 SNPs were found in the genes, of which seven are synonymous. PCR-based haplotype analysis of the fourth intron of Wknox1d and the fifth-to-sixth-exon region of Wknox1b provides an opportunity to make an assumption that hexaploid wheats T. aestivum subsp. macha var. palaeocolchicum and var. letshckumicum differ from other macha samples by the absence of a 42 bp insertion in the fourth intron of Wknox1d. One possible explanation for this observation would be that two Aegilops tauschii Coss. (A) and (B) participated in the formation of hexaploids through the D genome: Ae. tauschii (A)—macha (1–5, 7, 8, 10–12), and Ae. tauschii (B)—macha M6, M9, T. aestivum subsp. aestivum cv. ‘Chinese Spring’ and cv. ‘Red Doly’.

The complete chloroplast genome of Sargassum fusiforme

The complete chloroplast genome sequence of Sargassum fusiforme is presented here. Circular mapping revealed that the complete chloroplast DNA sequence of S. fusiforme was 124,298 bp in length and had an overall AT content of 69.57%, including 137 protein-coding genes, 2 open reading frames, 28 transfer RNA genes, and 6 ribosomal RNA genes. The phylogenetic tree based on Bayesian shows that all kinds of Phaeophyceae were clustered into two monophyletic groups.

The complete chloroplast genome sequence of Hypolepis sparsisora (Dennstaedtiaceae)

The complete chloroplast DNA sequence of Hypolepis sparsisora was the first time to report with 150,839 bp in length. The structure of the chloroplast genome is composed in four regions, which consisted a large single-repeat region (LSC) of 84,285 bp, a small single-repeat region (SSC) of 21,390 bp, and two inverted repeat regions (IRs) of 22,582 bp, respectively. A total of 131genes including 8 rRNA genes, 35 tRNA genes, and 83 protein-coding genes were predicted. We obtained phylogenetic tree by using the complete chloroplast DNA of 15 species ferns. The ML and BI phylogenetic analysis showed the location of H. sparsisora in Dennstaedtiaceae and other families.

The complete chloroplast genome of Gentaina urnula (Gentianaceae)

Gentiana urnula, is an alpine medicinal endemic to the Qinghai-Tibetan Plateau. In this study, the complete chloroplast DNA sequence of G. urnula was assembled，and its genome is 149,064 bp in length, including a large single-copy region (LSC) of 81,158 bp and a small single-copy region (SSC) of 17,034 bp, which was separated by a pair of 25,436 bp inverted repeat regions (IRs). A total of 130 genes are detected, including 86 protein-coding genes, 37 tRNA genes, and 7 rRNA genes. The complete chloroplast genomes of G. urnula will help to study the genetic diversity and phylogenetic analysis of Gentianales.

The complete chloroplast genome of Swertia cordata

Swertia cordata, a herbaceous plant, has been used in medicines as an alternative febrifuge and anthelmintic. In this study, the complete chloroplast DNA sequence of S. cordata was assembled. It is 153,429 bp in length, including a large single-copy region of 83,613 bp and a small single-copy region of 18,038 bp separated by a pair of inverted repeat regions of 25,889 bp each. A total of 129 genes are detected, including 84 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. The complete chloroplast genome of S. cordata will help to study the genetic diversity and phylogenetic analysis of Swertia.

The complete chloroplast genome of Sargassum horneri and its phylogenetic analysis

Complete chloroplast genome of Sargassum horneri was obtained in this work. Circular mapping revealed that the complete chloroplast DNA sequences of S. horneri was 124,075 bp in length and had an overall AT content of 69.41%, including 139 protein-coding genes, 28 transfer RNA genes, and 6 ribosomal RNA genes. The phylogenetic tree shows that S. horneri and Sargassum confusum constituted a sister clade along with Fucus vesiculosus.

Next generation sequencing technologies for the development of molecular markers and the analysis of genome diversity in Capsicum spp.

The accessibility and use of natural genetic variability in crops is essential for the management of genetic resources and the establishment of innovative breeding programs. Nuclear whole-genome sequences of several cultivated and wild peppers have been recently generated in order to provide crucial information on the evolution, domestication and divergence of pepper species (Capsicum spp.). Despite next generation sequencing (NGS) technologies providing a significant advance in generation of high throughput data, the analysis of a large-scale data and of individuals is still a challenging task. An alternative strategy to Whole Genome Sequencing is to generate a reduced representation of the genome using target enrichment strategies. Among them, genotyping-by-sequencing (GBS) provides a rapid, highly informative, cost-effective tool for exploring genetic diversity on a genome-wide scale. We applied GBS for SNP marker discovery in nuclear genomes and population structure analysis in a collection of cultivated and domesticated peppers. Information generated in this study represents a first step towards future genome-wide association mapping studies and marker-assisted selection programs. In addition, results from this analysis can be used to gain a better understanding of the genetic relationships within the Capsicum genus. In order to provide a further contribution to that issue, we also determined the complete chloroplast DNA (cpDNA) sequence of 11 Capsicum accessions for a total of 8 species generally included in the C. annuum (7), C. baccatum (3) or C. pubescens/eximium complex (1). We performed comparative analysis and identified potential molecular markers able to discriminate among Capsicum species.

The complete chloroplast genome sequence of Angelica Tsinlingensis (Apioideae)

Angelica tsinlingensis is endemic in Shaanxi province (China), and is used as Angelica sinensis in folk. Owing to important medicinal value, wild A. sinensis has been over-exploited and become quite rare in China during over 2000 years. Angelica sensu lato (s.l.; Apiaceae subfamily Apioideae) is a taxonomically complex and controversial group, and A. tsinlingensis is clearly different from members of the Angelica s.s. clade with the morphological and molecular results. The complete chloroplast DNA sequence of A. tsinlingensis (GenBank accession number: MF924726) was determined in our study. The size of chloroplast genome of A. tsinlingensis is 147,104 bp, including a large single-copy (LSC) region of 93376 bp and a small single-copy (SSC) region of 17,574 bp separated by a pair of identical inverted repeat regions (IRs) of 18,077 bp each. A total of 125 genes were successfully annotated containing 83 protein-coding genes, 34 tRNA genes and 8 rRNA genes. GC content of IRs region is the highest (44.5%). The result of Phylogenomic analysis supports the difference of A. tsinlingensis from Angelica s.s. clade.

The complete chloroplast genome of Artemisia hallaisanensis Nakai (Asteraceae), an endemic medicinal herb in Korea

We determined the complete chloroplast DNA sequence of Artemisia hallaisanensis Nakai, an endemic herbal species distributed on Jeju Island, Korea. The chloroplast DNA is 151,015 bp in length and encodes 4 rRNA, 30 tRNA, and 80 protein-coding genes. Phylogenetic analysis and sequence comparison of protein-coding genes with other Artemisa chloroplast DNAs revealed that the chloroplast genome of A. hallaisanensis is closely related to that of A. capillaris. Additionally, a unique 9 bp deletion in ycf1 gene is specific to A. hallaisanensis.

The complete chloroplast genome of Gentiana macrophylla

Gentiana macrophylla is a perennial medicinal plant in the family Gentianaceae. Natural sources for G. macrophylla plants are being exhausted due to over-collection and need urgent conservation. The complete chloroplast DNA sequence of G. macrophylla (GenBank accession number: KY856959) was determined in our study. The size of chloroplast genome of G. macrophylla is 149,916 bp, including a large single-copy (LSC) region of 82,911 bp and a small single-copy (SSC) region of 17,095 bp separated by a pair of identical inverted repeat regions (IRs) of 24,955 bp each. A total of 130 genes were successfully annotated containing 86 protein-coding genes, 36 tRNA genes and 8 rRNA genes. The GC content is 37.7%. 33 chloroplast simple sequence repeats (cpSSRs) of Gentiana macrophylla were detected, in which 32 of them are mononucleotide repeats. Phylogenomic analysis strongly supported the close relationship of G. macrophylla and other 5 species in Gentiana.

The Investigation of Organelle Genomes of Extra Nuclear Sunflower Mutants with Variegated Phenotype

The comparative analysis of complete chloroplast and mitochondrial DNA sequences of the original inbred line 3629 and three extra nuclear mutant lines variegated-1, variegated-15, variegated-17, obtained by the N-nitroso-N-methylurea induced mutagenesis, was done. The studied mutant lines were presented two different phenotypes with diverse chlorophyll content: Pale/green mutant line variegated-17 with 1.8% relative (to 3629 line) chlorophyll content in mutant leaf tissue and yellow/green lines variegated-1, variegated-15 with 6.3% and 4.3% relative chlorophyll content, respectively. Each line had unique variation sites in chloroplast DNA. Among chloroplast SNP there were determined 7 nonsynonymous substitutions, which were located in psaA (variegated-1), petD, rpl36, ccsA (variegated-15), rps4, rpoA, rpoC1 (variegated-17). The SNP of variegated-17 line has resulted in the frameshift (rpoC1) and premature stop codon formations (rpoA, rpoC1). The whole genome sequencing of mitochondrial DNA mutant lines revealed no differences as compared with original line 3629. Summarizing current data and our previous research of variegated mutants we assume that pale/green variegated phenotype is associated with significant disturbance of RNA polymerase subunits genes, and the lesions in photosynthetic genes lead to yellow/green mutant phenotype.

A Phylogenetic Analysis of Chloroplast Genomes Elucidates the Relationships of the Six Economically Important Brassica Species Comprising the Triangle of U.

The Brassica genus comprises many economically important worldwide cultivated crops. The well-established model of the Brassica genus, U’s triangle, consists of three basic diploid plant species (Brassica rapa, Brassica oleracea, and Brassica nigra) and three amphidiploid species (Brassica napus, Brassica juncea, and Brassica carinata) that arose through interspecific hybridizations. Despite being extensively studied because of its commercial relevance, several aspects of the origin of the Brassica species and the relationships within and among these six species still remain open questions. Here, we successfully de novo assembled 60 complete chloroplast genomes of Brassica genotypes of all six species. A complete map of the single nucleotide variants and insertions and deletions in the chloroplast genomes of different Brassica species was produced. The chloroplast genome consists of a Large and a Small Single Copy (LSC and SSC) region between two inverted repeats, and while these regions of chloroplast genomes have very different molecular evolutionary rates, phylogenetic analyses of different regions yielded no contradicting topologies and separated the Brassica genus into four clades. B. carinata and B. juncea share their chloroplast genome with one of their hybridization donors B. nigra and B. rapa, respectively, which fits the U model. B. rapa, surprisingly, shows evidence of two types of chloroplast genomes, with one type specific to some Italian broccoletto accessions. B. napus clearly has evidence for two independent hybridization events, as it contains either B. rapa chloroplast genomes. The divergence estimation suggests that B. nigra and B. carinata diverged from the main Brassica clade 13.7 million years ago (Mya), while B. rapa and B. oleracea diverged at 2.18 Mya. The use of the complete chloroplast DNA sequence not only provides insights into comparative genome analysis but also paves the way for a better understanding of the phylogenetic relationships within the Brassica genus.

Genome Sequences of Populus tremula Chloroplast and Mitochondrion: Implications for Holistic Poplar Breeding

Complete Populus genome sequences are available for the nucleus (P. trichocarpa; section Tacamahaca) and for chloroplasts (seven species), but not for mitochondria. Here, we provide the complete genome sequences of the chloroplast and the mitochondrion for the clones P. tremula W52 and P. tremula x P. alba 717-1B4 (section Populus). The organization of the chloroplast genomes of both Populus clones is described. A phylogenetic tree constructed from all available complete chloroplast DNA sequences of Populus was not congruent with the assignment of the related species to different Populus sections. In total, 3,024 variable nucleotide positions were identified among all compared Populus chloroplast DNA sequences. The 5-prime part of the LSC from trnH to atpA showed the highest frequency of variations. The variable positions included 163 positions with SNPs allowing for differentiating the two clones with P. tremula chloroplast genomes (W52, 717-1B4) from the other seven Populus individuals. These potential P. tremula-specific SNPs were displayed as a whole-plastome barcode on the P. tremula W52 chloroplast DNA sequence. Three of these SNPs and one InDel in the trnH-psbA linker were successfully validated by Sanger sequencing in an extended set of Populus individuals. The complete mitochondrial genome sequence of P. tremula is the first in the family of Salicaceae. The mitochondrial genomes of the two clones are 783,442 bp (W52) and 783,513 bp (717-1B4) in size, structurally very similar and organized as single circles. DNA sequence regions with high similarity to the W52 chloroplast sequence account for about 2% of the W52 mitochondrial genome. The mean SNP frequency was found to be nearly six fold higher in the chloroplast than in the mitochondrial genome when comparing 717-1B4 with W52. The availability of the genomic information of all three DNA-containing cell organelles will allow a holistic approach in poplar molecular breeding in the future.

The complete chloroplast genome of Wakame (Undaria pinnatifida), an important economic macroalga of the family Alariaceae

We decoded the complete chloroplast DNA (cpDNA) sequence of the Wakame (Undaria pinnatifida), an important economic macroalga of the family Alariaceae, by using next-generation sequencing technology. The genome consists of 130 336 bp containing a pair of inverted repeats (IRs) of 4790 bp, which was separated by a large single-copy region and a small single-copy region of 77 821 and 42 934 bp, respectively. The genic regions account for 77.7% of whole cpDNA, and the GC content of the cpDNA was 30.6%. The U. pinnatifida cpDNA encodes 153 unigenes (129 protein-coding genes, 3 rRNA genes and 21 tRNA genes). There are 1 PCG (rpl33) and 1 tRNA genes (trnL) containing an intron. A phylogenetic analysis of the four complete cpDNA from Phaeophyceae showed that U. pinnatifida is closely related to Saccharina japonica with high bootstrap value supported. The complete cpDNA of U. pinnatifida provides essential and important DNA molecular data for further phylogenetic and evolutionary analysis for brown algae.

The complete chloroplast genome of Tianshan Snow Lotus (Saussurea involucrata), a famous traditional Chinese medicinal plant of the family Asteraceae

abtractWe decoded the complete chloroplast DNA (cpDNA) sequence of the Tianshan Snow Lotus (Saussurea involucrata), a famous traditional Chinese medicinal plant of the family Asteraceae, by using next-generation sequencing technology. The genome consists of 152 490 bp containing a pair of inverted repeats (IRs) of 25 202 bp, which was separated by a large single-copy region and a small single-copy region of 83 446 bp and 18 639 bp, respectively. The genic regions account for 57.7% of whole cpDNA, and the GC content of the cpDNA was 37.7%. The S. involucrata cpDNA encodes 114 unigenes (82 protein-coding genes, 4 rRNA genes, and 28 tRNA genes). There are eight protein-coding genes (atpF, ndhA, ndhB, rpl2, rpoC1, rps16, clpP, and ycf3) and five tRNA genes (trnA-UGC, trnI-GAU, trnK-UUU, trnL-UAA, and trnV-UAC) containing introns. A phylogenetic analysis of the 11 complete cpDNA from Asteracease showed that S. involucrata is closely related to Centaurea diffusa (Diffuse Knapweed). The complete cpDNA of S. involucrata provides essential and important DNA molecular data for further phylogenetic and evolutionary analysis for Asteraceae.

The complete chloroplast genome of Aconitum chiisanense Nakai (Ranunculaceae)

We determined the complete chloroplast DNA sequence of Aconitum chiisanense Nakai, a rare Aconitum species endemic to Korea. The chloroplast genome is 155 934 bp in length and contains 4 rRNA, 30 tRNA, and 78 protein-coding genes. Phylogenetic analysis revealed that the chloroplast genome of A. chiisanense is closely related to that of A. barbatum var. puberulum. Sequence comparison with other Ranunculaceae chloroplasts identified a unique deletion in the rps16 gene of A. chiisanense chloroplast DNA that can serve as a molecular marker for species identification.

The complete chloroplast genome of Antarctic pearlwort, Colobanthus quitensis (Kunth) Bartl. (Caryophyllaceae)

We determined the complete chloroplast DNA sequence of an extremophile plant, Colobanthus quitensis (Antarctic pearlwort), by de novo assembly based on the sequencing results from Illumina MiSeq platform (Illumina Inc., San Diego, CA). The chloroplast genome of C. quitensis (NCBI accession no. KT737383) is a sequence of 151 276 bp long with a typical quadripartite structure composed of a large single copy region, a small single copy region and a pair of inverted repeats. The overall GC content of C. quitensis genome is 36.7% and it has 66 simple sequence repeats. It contains a total 112 genes including 78 protein coding genes, 30 tRNA genes, and four rRNA genes.

Complete chloroplast DNA sequences of Zanduri wheat (Triticum spp.)

Georgia plays an important role in wheat formation. In the past, the Zanduri population of Georgia was a set of diploid—Triticum monococcum var. hornemanii (2n = 14) (Gvatsa Zanduri), tetraploid Triticum timopheevii (2n = 28) (Chelta Zanduri) and hexaploid Triticum zhukovskyi Men. et Er. (2n = 42). It is a Zanduri puzzle that wild T. araraticum was not found in Georgia, though cultivated T. timopheevii was only detected here. Next-generation sequencing technologies, which have been developed in recent years, enable the determination of complete nucleotide sequences of both chloroplast and mitochondrial DNA of many higher plants, including wheat. The genetic structure of Zanduri wheat is more accurately inferred by the complete sequences of chloroplast DNA. In the present investigation, the complete sequences of three Zanduri wheats (T. timopheevii, T. zhukovskyi, and T. monococcum var. hornemanii) and wild T. araraticum are presented. Sequencing of chloroplast DNA was performed on an Illumina MiSeq platform. Chloroplast DNA molecules were assembled using the SOAPdenovo computer program. In comparison to T. araraticum, there are 12 SNPs, a 25 bp inversion in the ccsA-ndhD intergenic sequence, and a 38-bp inversion in the intergenic sequence rbcL-rpl23 pseudogene identified in T. timopheevii and T. zhukovskyi. In addition, a 24 bp repeat of trnG-trnI intergenic sequence is present as a double copy in T. araraticum, whereas in T. timopheevii and T. zhukovskyi, it is present as a triple copy. Unlike T. araraticum, T. timopheevii and T. zhukovskyi have a 6 bp repeat in the gene ndhH, which results in a dipeptide duplication in the corresponding protein. Gvatsa Zanduri (T. monococcum var. hornemanii) chloroplast DNA slightly differs from other einkorn chloroplast DNA. In comparison to T. monococcum, four SNPs can be identified in T. monococcum (Gvatsa Zanduri), two in gene matK and one in gene ndhD. The sequenced chloroplast DNA molecules were compared to other Triticum and Aegilops species, and a phylogenetic tree was constructed. T. araraticum, T. timopheevii and T. zhukovskyi chloroplast DNA showed the closest phylogenetic relationship with the chloroplast DNA of Ae. speltoides. The most significant difference was in the 114-bp deletion within the gene ndhH in the Timopheevi species.

Chloroplast genome differences between Asian and American Equisetum arvense (Equisetaceae) and the origin of the hypervariable trnY-trnE intergenic spacer.

Comparative analyses of complete chloroplast (cp) DNA sequences within a species may provide clues to understand the population dynamics and colonization histories of plant species. Equisetum arvense (Equisetaceae) is a widely distributed fern species in northeastern Asia, Europe, and North America. The complete cp DNA sequences from Asian and American E. arvense individuals were compared in this study. The Asian E. arvense cp genome was 583 bp shorter than that of the American E. arvense. In total, 159 indels were observed between two individuals, most of which were concentrated on the hypervariable trnY-trnE intergenic spacer (IGS) in the large single-copy (LSC) region of the cp genome. This IGS region held a series of 19 bp repeating units. The numbers of the 19 bp repeat unit were responsible for 78% of the total length difference between the two cp genomes. Furthermore, only other closely related species of Equisetum also show the hypervariable nature of the trnY-trnE IGS. By contrast, only a single indel was observed in the gene coding regions: the ycf1 gene showed 24 bp differences between the two continental individuals due to a single tandem-repeat indel. A total of 165 single-nucleotide polymorphisms (SNPs) were recorded between the two cp genomes. Of these, 52 SNPs (31.5%) were distributed in coding regions, 13 SNPs (7.9%) were in introns, and 100 SNPs (60.6%) were in intergenic spacers (IGS). The overall difference between the Asian and American E. arvense cp genomes was 0.12%. Despite the relatively high genetic diversity between Asian and American E. arvense, the two populations are recognized as a single species based on their high morphological similarity. This indicated that the two regional populations have been in morphological stasis.