Inverted Repeat Research Articles

Decoding the Chloroplast Genome of Korean endemic plant Acer okamotoanum: Comparative Genomics, Phylogenetic Insights, and Potential for Marker Development.

Acer okamotoanum, a medicinally significant endemic plant of Korea, has seen limited genomic research. To address this gap, we conducted a comprehensive sequencing and analysis of its chloroplast genome. The assembled genome is 156,242bp in length, with typical quadripartite structure, consisting of a large single-copy region, a small single-copy region, and two inverted repeat regions. It contains 130 genes, including 85 protein-coding, 37 tRNA, and 8 rRNA genes. Sixteen genes have a single intron, while clpP and ycf3 possess two introns each. Additionally, 17 genes are duplicated within the inverted repeat regions. The genome analysis revealed 92 Simple Sequence Repeats (SSRs), predominantly located in intergenic regions, with a bias toward A/T-rich codons. Comparative analysis with five closely related Acer species highlighted a highly conserved genomic structure, but also revealed differences in SSRs and repeat sequences. Hypervariable regions, such as rpl32-trnL and ycf1, were identified as potential molecular markers for phylogenetic and population studies. Phylogenetic analysis involving 37 chloroplast genomes confirmed the monophyly of the Acer genus and placed A. okamotoanum within the Platanoidea section, closely related to A. truncatum. This study improves the understanding of A. okamotoanum's genomic structure, offering insights for phylogenetic analysis, marker development, and conservation efforts.

Chloroplast genome characteristics and phylogenetic analysis of Macropanax rosthornii (Harms) C.Y. Wu ex G. Hoo (Araliaceae)

Macropanax rosthornii (Harms) C.Y.Wu ex G.Hoo 1900 is a small tree or evergreen shrub of the family Araliaceae, mainly distributed in southern China and Vietnam with a small distribution area. M. rosthornii is a kind of high-quality wild ornamental plant resource and is also known for its effective medicinal treatment for fracture, rheumatoid arthritis, traumatic injury, infant malnutrition, etc. In this study, the first complete chloroplast genome of M. rosthornii was reported and phylogenetic analysis was conducted with other 10 species from Araliaceae. The chloroplast genome was 152,831 bp with 39% overall GC content and includes a large single-copy (LSC) region (93,751 bp), a small single-copy (SSC) region (18,930 bp), and a pair of inverted repeats (IR) regions (20,075 bp). There are 109 genes in the chloroplast genome of M. rosthornii, including 78 protein-coding genes, 21 tRNA genes, and 10 rRNA genes. The phylogenetic tree showed that M. rosthornii is closely related to Hydrocotyle vulgaris and Hydrocotyle sibthorpioides. The chloroplast genome reported here will be beneficial for its interspecific identification and evolutionary studies of Macropanax.

The complete chloroplast genome sequence of Cathetus clarkei Hook.f.1890 R.W.Bouman, 2022 (Phyllanthaceae)

Cathetus clarkei, a significant folk medicinal plant, is utilized to treat a variety of ailments. In this study, we reported the complete chloroplast genome sequence of this species. The length of the complete chloroplast genome was 155,810 bp, included a pair of inverted repeat (IR) regions (26,340 bp), a large single-copy region (LSC, 84,853 bp), and a small single-copy region (SSC, 18,277 bp). It comprised 128 genes, including 83 protein-coding genes, 37 tRNA genes, and eight rRNA genes. The total GC content was 36.8%. The phylogenetic tree revealed that C. clarkei had a close relationship with P. cochinchinensis, followed by P. reticulatus. This study provides a reference for important medicinal plants within the Phyllanthaceae, and we can gain a deeper understanding of how the species adapts to changes in the environment, helping us to identify and protect it.

The complete chloroplast genome of Heracleum hemsleyanum Diels (Apioideae), a traditional medicinal herb in China

Heracleum hemsleyanum Diels is a traditional medicinal herb in China. We reported its first complete chloroplast genome. The chloroplast genome was 146,775 bp in length with 37.53% GC content, containing a large single copy region (LSC, 93,309 bp), a small single copy region (SSC, 17,502 bp), and a pair of inverted repeat regions (IRs, 17,982 bp). Moreover, the chloroplast genome encoded 130 genes, including 86 protein-coding genes (PCGs), 36 transfer RNA genes (tRNAs), and eight ribosomal RNA genes (rRNAs). Phylogenetic analysis indicated that H. hemsleyanum was closely related to Heracleum moellendorffii and Heracleum yungningense. This assembled chloroplast genome will provide vital information on the genetic resources, phylogenetic relationships, and the species identification of the genus Heracleum.

Comparative and phylogenetic analyses of the chloroplast genomes of mangrove plants

Mangrove ecosystems play a crucial role in coastal environment by stabilizing shorelines, reducing erosion, and providing habitat for diverse wildlife. However, there are fewer studies on the genetic management of mangrove plants. In this study, the chloroplast genome of Hibiscus tiliaceus L. was obtained through whole genome resequencing. The complete chloroplast genome of this species was 161,764 bp in length and comprised 84 protein-coding genes, eight rRNA (ribosomal RNA) genes, 37 tRNA (transfer RNA) genes. We conducted a comparative analysis of the chloroplast genomes of 26 mangrove species and found the conservation of genome structure among these mangrove plants, including the number of protein-coding genes, rRNA genes, and tRNA genes. However, there were noticeable differences in the number of SSRs (simple sequence repeats), the number of repeat sequences, and the borders of the IR (inverted repeat) regions, indicating genetic differentiation among mangrove plants. Besides, we found special differences in the distribution of certain genes in chloroplast genomes of certain species. Moreover, phylogenetic analysis based on chloroplast genome data clearly revealed that neither true mangrove plants nor semi-mangrove plants were monophyletic. These results revealed that chloroplast genomes will be useful to help understand the genetic differentiation and phylogenetic relationships of mangrove plants, which could assist in the genetic management, conservation, and restoration of mangrove ecosystems.

The complete chloroplast genome sequences of monotypic genus Pseudogalium, and comparative analyses with its relative genera

BackgroundPseudogalium is a new monotypic genus with two subspecies in China and one in Japan, which holds a distinctive phylogenetic position and ecological significance within the tribe Rubieae. Chloroplast genomes contain abundant information for resolving phylogenetic relationships. To investigate the phylogenetics of P. paradoxum and its related genera, we first sequenced, assembled, and annotated the chloroplast genome of two subspecies of P. paradoxum in China and reconstructed the phylogenetic trees. Due to the lack of samples of P. paradoxum subsp. franchetianum from Japan, this study only analyzed and discussed P. paradoxum subsp. paradoxum and P. paradoxum subsp. duthiei.ResultsThis study had shown that the complete chloroplast genomes of Pseudogalium ranged from 153,093 bp to 153,333 bp in length with 130 genes in total, all of which had typical circular structures consisting of a large single-copy region, a small single-copy region and a pair of inverted repeat regions. The comparative analysis showed that the chloroplast genome of P. paradoxum was conserved in the inverted repeat regions. Additionally, we identified 60 dispersed repeat sequences, 61–63 simple sequence repeats, and 30 codons within the 82 protein-coding genes that exhibited RSCU values greater than one. Furthermore, we detected highly divergent regions that hold potential as new DNA barcodes for species identification. Compared with Pseudogalium, the gene number, gene length, and GC content in the chloroplast genomes of Galium and Rubia exhibited differential characteristics, and the dispersed repeat sequences and SSRs in Galium and Rubia were significantly different. Phylogenetic analysis based on the whole chloroplast genomes showed that Pseudogalium can be treated as a new genus, with P. paradoxum subsp. paradoxum and P. paradoxum subsp. duthiei considered as two distinct subspecies of P. paradoxum.ConclusionsThe complete chloroplast genomes of P. paradoxum were first reported in this study, which provided a new insight into phylogeny and taxonomy of this genus. Phylogenetic analyses strongly supported the following proposals: (1) P. paradoxum can be isolated as a genus closely related to Galium; (2) P. paradoxum subsp. paradoxum and P. paradoxum subsp. duthiei form distinct clades, both of which can be considered as subspecies of P. paradoxum.

The complete chloroplast genome of Barringtonia asiatica (L.) Kurz (Lecythidaceae)

Genomic resources for Barringtonia asiatica (L.) Kurz, a pantropical beach forest species native to the Philippines, have not been published to date, despite its medicinal importance and potential for biomedical applications. In this study, we assembled and characterized the first complete chloroplast genome of B. asiatica using Illumina paired-end sequencing technology. It displayed a typical quadripartite structure with a sequence length of 158,794 bp, comprised of a large single copy of 88,196 bp, small single-copy of 18,448 bp, and a pair of inverted repeat regions of 26,075 bp each. The cp genome contained 35 tRNA genes, eight rRNA genes, and 86 protein-coding genes with an overall GC content of 36.8%. Phylogenetic analysis revealed a close evolutionary relationship between B. racemosa and B. fusicarpa, forming a monophyletic group with B. asiatica with high bootstrap support, thereby enhancing our understanding of the phylogeny, systematics, and genetics of Asian Lecythidaceae species.

The complete chloroplast genome and phylogenetic analysis of Persicaria jucunda (Meisn.) Migo (Polygonaceae)

Persicaria jucunda is a plant distributed at meadow or wetland. Our study reports the complete chloroplast genome. The chloroplast genome of P. jucunda is a typical tetrameric structure with a total length of 159,843 bp, containing a large single-copy (LSC) region of 84,350 bp, a small single-copy (SSC) region of 13,151 bp, and two inverted repeat regions (IRs) of 31,171 bp. The total GC content was 38.2%, 36.5% for the LSC region, 33.2% for the SSC region, and 41.5% for the IR region. The P. jucunda chloroplast genome contains 128 genes, including 83 protein-coding genes, 37 tRNA genes, and eight rRNA genes. Phylogenetic analysis based on 36 chloroplast genomes showed similarity to P. foliosa and P. kawagoeana among the species analyzed. The chloroplast genome provides a valuable genetic resource for phylogenetic studies.

AAV vector transduction restriction and attenuated toxicity in hESCs via a rationally designed inverted terminal repeat.

Adeno-associated virus (AAV) inverted terminal repeats (ITRs) induce p53-dependent apoptosis in human embryonic stem cells (hESCs). To interrogate this phenomenon, a synthetic ITR (SynITR), harboring substitutions in putative p53 binding sites was generated and evaluated for vector production and gene delivery. Replication of SynITR flanked transgenic genome was similar compared to wildtype (wt)ITR, with a modest increase in vector titers. Packaged in the AAV2 capsid, wtITR and SynITR vectors demonstrated similar transduction efficiencies of human cells without toxicity. Following AAV2-wtITR vector infection of hESCs, rapid apoptosis was observed as reported. In contrast, infection by AAV2 vectors packaged with SynITRs attenuated the wtITR-induced hESC toxicity. While hESC particle entry and the abundance of double stranded circular episomes was similar for the ITR contexts, reporter expression was inhibited from transduced SynITR genomes. Mechanistically, infection of hESCs induced γH2AX in an ITR-independent manner, however, canonical activation of p53α was uncoupled using AAV-SynITR. Further investigations in hESCs revealed additional novel findings: (i) p53β is uniquely and constitutively activeand (ii) AAV vector infection, independent of the ITR sequence, induces activation of p53ψ. The data herein reveal an ITR-dependent AAV vector transduction restriction specific to hESCs and manipulation of the DNA damage response via ITR engineering.

Characteristics and phylogenetic analysis of the complete chloroplast genome of Rubus swinhoei Hance 1866 from the family Rosaceae

Rubus swinhoei Hance is an important plant owing to its medicinal root and edible fruit, and extensively distributed in China. In this study, we reported the complete chloroplast genome of R. swinhoei. The chloroplast genome was 156,335 bp in size with the overall GC content of 37.15%, having a circular and quadripartite structure, which contained a large single-copy (LSC) and a small single-copy (SSC) regions of 85,897 bp and 18,858 bp separated by a pair of 25,790 bp inverted repeat (IR) regions. The complete chloroplast genome comprised 131 unique genes of which 86 and 37 were protein-coding genes and tRNA genes, respectively, and also eight rRNA genes. The phylogenetic analysis revealed that R. swinhoei was closely related to R. kawakamii. The genome information reported in this paper will be beneficial for further investigation on the evolution of this species.

Expanding the Diversity of Actinobacterial Tectiviridae: A Novel Genus from Microbacterium.

Six novel Microbacterium phages belonging to the Tectiviridae family were isolated using Microbacterium testaceum as a host. Phages MuffinTheCat, Badulia, DesireeRose, Bee17, SCoupsA, and LuzDeMundo were purified from environmental samples by students participating in the Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) program at Alliance University, New York. The phages have linear dsDNA genomes 15,438-15,636 bp with 112-120 bp inverted terminal repeats. Transmission electron microscopy (TEM) imaging analysis revealed that the six novel phages have six-sided icosahedral double-layered capsids with an internal lipid membrane that occasionally forms protruding nanotubules. Annotation analysis determined that the novel Microbacterium phages all have 32-34 protein-coding genes and no tRNAs. Like other Tectiviridae, the phage genomes are arranged into two segments and include three highly conserved family genes that encode a DNA polymerase, double jelly-roll major capsid protein, and packaging ATPase. Although the novel bacteriophages have 91.6 to 97.5% nucleotide sequence similarity to each other, they are at most 58% similar to previously characterized Tectiviridae genera. Consequently, these novel Microbacterium phages expand the diversity of the Tectiviridae family, and we propose they form the sixth genus, Zetatectivirus.

The complete chloroplast genome of Solanum melongena ‘Yunqie 9’

Solanum melongena ‘Yunqie 9’ was selected by the Horticultural Research Institute of Yunnan Academy of Agricultural Sciences based on the local environment of Yunnan Province. It is excellent in fruit quality and yield, but it is relatively weak in disease resistance. No information on complete chloroplast genome and position in the phylogeny of Solanum to restrict its genetic improvement. In this study, the chloroplast genome of ‘Yunqie 9’ was sequenced using Illumina high-throughput sequencing technology. The size of the complete chloroplast genome was 155,579 bp in length with an GC content of 37.70%, composed of a large single-copy region (86,189 bp), a small single-copy region (18,504 bp), and a pair of inverted repeat regions (25,443 bp). A total of 134 coding genes were annotated in the entire chloroplast genome, including 89 protein-coding genes, 37 transfer RNA genes, and 8 ribosomal RNA genes. Phylogenetic analysis based on chloroplast genome sequences of 20 species in the Solanaceae family indicated that except for the cultivated Solanum, ’Yunqie 9’ was closely related to Solanum melongena×Solanum torvum and Solanum insanum.

A recent large-scale intraspecific IR expansion and evolutionary dynamics of the plastome of Peucedanum japonicum

Peucedanum japonicum (PJ), a member of the Apiaceae family, is widely distributed and cultivated in East Asian countries for edible and functional foods. In this study, we compared the plastid genomes (plastomes) and 45S nuclear ribosomal DNA (45S nrDNA) simultaneously from 10 PJ collections. Plastome-based phylogenetic analysis showed that the PJ accessions were monophyletic within the genus Peucedanum. However, ten plastomes were classified into two different groups according to their length of inverted repeat (IR) block, the short-type (S-type) plastome group containing the 18.6 kbp of the original IR and the long-type (L-type) plastome group containing the 35.7 kbp of expanded IR by duplication of the 17.1 kbp of the large single copy region. A total of nine single nucleotide polymorphisms and eight insertions or deletions were identified among the five L-type plastomes, whereas large variations were identified among the five S-type plastomes. Calculation of synonymous substitution rates and divergence time estimation suggested that the 17 kbp IR expansion occurred recently. Molecular markers were developed and validated to classify the 55 PJ germplasm according to their plastome types. Our study would be useful for unraveling the dynamic evolution of plastomes in the Apiaceae family and for the molecular breeding of PJ.

Characterization of the complete plastid genome of Clivia mirabilis (Amaryllidaceae)

Clivia mirabilis Rourke 2002 is an evergreen herbaceous flower with high ornamental value. In this study, we sequenced the complete chloroplast (cp) genome of C. mirabilis and reported it for the first time. The cp genome was 158,914 base pairs (bp) in total length, including two inverted repeats (IRs, 27,052 bp), separated by a large single-copy region (LSC, 86,519 bp) and a small single-copy region (SSC, 18,291 bp). There are 133 different genes in the cp genome of Clivia mirabilis, including 87 protein-coding genes, 38 transfer RNA genes, and eight ribosomal RNA genes. The overall GC content of the cp genome was 37.9%. Our phylogenetic analysis showed that C. mirabilis formed a monophyletic clade with the other sampled species of Clivia, falling into the Amaryllidoideae clade. Our findings could be used to identify and analyze the genetic diversity of C. mirabilis and provide new data for taxonomic and phylogenetic studies of Clivia.

The complete chloroplast genome of Hemisteptia lyrata (Bunge) Fisch. & C. A. Mey. 1836 (Asteraceae) and its phylogenetic analysis

Hemisteptia lyrata, widely distributed in China, has a 152,467 bp chloroplast genome with a large single-copy (LSC) region of 83,473 bp, a small single-copy (SSC) region of 18,594 bp, a pair of inverted repeat regions (IRs) of 25,194 bp in length. The GC content is 36.46%. A total of 133 genes were encoded, of which 88 were protein coding genes, eight were rRNA genes, and 37 were tRNAs. The phylogenetic analysis using the maximum-likelihood method showed that H. lyrata is closely related to the species in Saussurea. This study provides genomic resources for phylogenetic genetics and resource exploitation of H. lyrata.

Asynchronous evolution of centromeric sequences across chromosomes in Pyricularia oryzae

Centromeres are essential for chromosome segregation, yet they are among the most rapidly evolving regions of the genome. The mechanisms driving this rapid evolution of centromeric sequences are still not well understood. In this study, we identified the centromeric sequences of a wheat-infecting Pyricularia oryzae strain (Br48) using CENP-A chromatin immunoprecipitation followed by high-throughput sequencing. The Br48 centromeres range from 71 kb to 101 kb in length and are highly AT-rich (72.1-75.5%) and repeat-rich (63.4-85.0%). These regions are also enriched in H3K9me3 and 5-methylcytosine but depleted of H3K4me2 and H3K27me3. During the analysis of repetitive sequences in the Br48 centromere, we identified a stretch of approximately 530 bp that is tightly associated with centromeres in the Pyricularia fungus. We designated this element as the centromere-associated IR element (CenIR), as it often forms inverted repeat structures with two elements adjacent in reverse orientation. A comparison of putative centromere sequences across phylogenetically distinct P. oryzae strains suggests that changes in centromeric sequences are non-uniform across chromosomes and do not always align with the fungal phylogenetic relationships. Repeat-induced point mutation (RIP)-like CG to TA transitions likely accelerate base substitutions in the centromeres of Pyricularia fungi.

The Complete Chloroplast Genome of Erythrina variegata L. (Papilionoideae, Fabaceae).

Erythrina variegata L. 1754, a thorny deciduous tree of Fabaceae, contains various chemical compounds such as alkaloids, flavonoids, and triterpenoids and exhibits anti-depressant, anti-inflammatory, and antidiabetic activities. However, genomic data of E. variegata are limited. In this study, the complete chloroplast genome of E. variegata was sequenced and characterized using Illumina sequencing platform. The chloroplast genome of E. variegata was 152,351 bp in length and consisted of a large single copy (82,907 bp), a small single copy (26,309 bp), and two inverted repeat regions (16,826 bp). There were 79 protein-coding genes, 30 transfer RNA genes, and four ribosomal RNA genes. Comparative analysis revealed high conservation of chloroplast genomes among Erythrina species regarding genome size, structure, and gene content. The phylogenetic study also indicated a close relationship between E. variagata and E. sanwicensis. This study provides initial plastome data for further genomic studies examining E. variegata and related species in Fabaceae.

Complete chloroplast genome of Eleutherococcus divaricatus var. chiisanensis (Araliaceae), an endemic Korean species

Eleutherococcus Maxim. belongs to the family Araliaceae, with approximately 40 species distributed in Eastern Asia and the Himalayan regions. Among these, Eleutherococcus divaricatus var. chiisanensis is distributed only in the central Korean Peninsula. The complete chloroplast (cp) genome sequence of Eleutherococcus divaricatus var. chiisanensis (Araliaceae) was determined to be 156,765 bp in length, consisting of large (86,592 bp) and small (18,239 bp) single-copy regions and a pair of identical inverted repeats (25,967 bp). The overall GC content of the chloroplast genome was 37.9%, and in the large singlecopy sequence, the small single-copy sequence, and each of inverted repeat sequences, the contents were 36.2%, 32%, and 43%, respectively. The genome consists of 132 genes, made up of 87 protein-coding genes, 37 tRNA genes, and eight rRNA genes. A cp genome phylogenetic analysis of the 12 taxa inferred from the cp genome revealed a close relationship between Eleutherococcus sessiliflorus and E. divaricatus var. chiisanensis<. The complete cp genome sequence of E. divaricatus var. chiisanensis provides important information for future phylogenetic and evolutionary studies of the genus Eleutherococcus.

Complete chloroplast genome of Syzygium glomeratum (Myrtaceae) and phylogenetic analysis

Syzygium glomeratum (Lam.) DC., known as “Tram Tron” in Vietnam, is an evergreen tree known for its medicinal properties. To understand the genomic and evolutionary basis of this plant, we sequenced and assembled the complete chloroplast genome of S. glomeratum for the first time, using the Illumina platform. The complete chloroplast of S. glomeratum was 158,469 bp in length and contained a large single-copy region (87,962 bp) and a small single-copy region (18,391 bp) separated by inverted repeat regions (26,058 bp). The genome encoded 85 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. A phylogenetic analysis of 42 Syzygium species revealed significant insight into their evolutionary relationships, indicating a closer relationship between S. glomeratum and its sister group (S. cinereum and S. claviflorum). In conclusion, our results provide genomic information pertaining to the chloroplast genome in S. glomeratum, which is also genetic information useful for further study of biodiversity, conservation, and evolutionary biology.

Complete chloroplast genome sequence of a novel Withania somnifera (L.) Dunal: Comparative genomics and phylogenetic insights

This study introduces a novel variety of the highly esteemed medicinal plant Withania somnifera (L.) Dunal from the family Solanaceae. The new variety, Withania somnifera var. abhaica Nadia, A. Ali & M.S. Alwahibi, var. nov., is distributed at high altitudes in the Abha hills of Saudi Arabia. The distinct characteristics of the novel variety of W. somnifera include elliptic-elongated leaves that are thick and semi-succulent in nature, and a fruiting calyx with a bifurcated tip measuring approximately 0.5 mm in length, with each bifurcated tip being botuliform in shape. Using next-generation sequencing (NGS) techniques, we investigated the chloroplast genome of this variety. The complete chloroplast genome of W. somnifera var. abhaica from the Abha region, Saudi Arabia, spans 153,621 bp, with a GC content of 37.7%. It includes a large single-copy (LSC) region of 84,972 bp (GC 35.8%), a small single-copy (SSC) region of 18,400 bp (GC 31.7%), and two inverted repeats (IRs) of 50,249 bp (GC 43.2%). Annotation of the chloroplast genome identified 131 genes, comprising 86 protein-encoding genes (PCGs), eight ribosomal RNA genes, and 37 transfer RNA genes. Repeat analysis identified 38 simple sequence repeats (SSRs) and 50 longer repeat sequences in the plastome. A total of 66 RNA-editing sites were detected across 24 PCGs of the plastome. Comparative genomic studies including synteny analysis, supported and validated the assembled plastome. Nucleotide diversity analysis revealed psbJ, psbA, ndhF, and ycf1 as the most hypervariable barcodes. Phylogenetic analyses suggested the monophyly of the genus Withania. Moreover, the newly sequenced chloroplast genome of W. somnifera var. abhaica was found to be distinct from the typical W. somnifera. Bangladesh J. Plant Taxon. 31(2): 205-223, 2024 (December)