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40 years ago, organelle genomes were assumed to be streamlined and, perhaps, unexciting remnants of their prokaryotic past. However, the field of organelle genomics has exposed an unparallel diversity in genome architecture (i.e. genome size, structure, and content). The transcription of these eccentric genomes can be just as elaborate - organelle genomes are pervasively transcribed into a plethora of RNA types. However, while organelle protein-coding genes are known to produce polycistronic transcripts that undergo heavy posttranscriptional processing, the nature of organelle noncoding transcriptomes is still poorly resolved. Here, we review how wet-lab experiments and second-generation sequencing data (i.e. short reads) have been useful to determine certain types of organelle RNAs, particularly noncoding RNAs. We then explain how third-generation (long-read) RNA-Seq data represent the new frontier in organelle transcriptomics. We show that public repositories (e.g. NCBI SRA) already contain enough data for inter-phyla comparative studies and argue that organelle biologists can benefit from such data. We discuss the prospects of using publicly available sequencing data for organelle-focused studies and examine the challenges of such an approach. We highlight that the lack of a comprehensive database dedicated to organelle genomics/transcriptomics is a major impediment to the development of a field with implications in basic and applied science., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: Pepper (Capsicum pubescens), one of five domesticated pepper species, has unique characteristics, such as numerous hairs on the epidermis of its leaves and stems, black seeds, and vibrant purple flowers. To date, no studies have reported on the complete assembly of the mitochondrial genome (mitogenome) of C. pubescens. Understanding the mitogenome is crucial for further research on C. pubescens., Results: In our study, we successfully assembled the first mitogenome of C. pubescens, which was assigned the GenBank accession number OP957066. This mitogenome has a length of 454,165 bp and exhibits the typical circular structure observed in most mitogenomes. We annotated a total of 70 genes, including 35 protein-coding genes (PCGs), 30 tRNA genes, 3 rRNA genes, and 2 pseudogenes. Compared to the other three pepper mitogenomes (KJ865409, KJ865410, and MN196478), C. pubescens OP957066 exhibited four unique PCGs (atp4, atp8, mttB, and rps1), while two PCGs (rpl10 and rps3) were absent. Notably, each of the three pepper mitogenomes from C. annuum (KJ865409, KJ865410, and MN196478) experienced the loss of four PCGs (atp4, atp8, mttB, and rps1). To further explore the evolutionary relationships, we reconstructed a phylogenetic tree using the mitogenomes of C. pubescens and fourteen other species. Structural comparison and synteny analysis of the above four pepper mitogenomes revealed that C. pubescens shares high sequence similarity with KJ865409 and that C. pubescens has rearranged with the other three pepper mitogenomes. Interestingly, we observed 72 similar sequences between the mitochondrial and chloroplast genomes, which accounted for 12.60% of the mitogenome, with a total length of 57,207 bp. These sequences encompassed 12 tRNA genes and the rRNA gene (rrn18). Remarkably, selective pressure analysis suggested that the nad5 gene underwent obvious positive selection. Furthermore, a single-base mutation in three genes (nad1, nad2, and nad4) resulted in an amino acid change., Conclusion: This study provides a high-quality mitogenome of pepper, providing valuable molecular data for future investigations into the exchange of genetic information between pepper organelle genomes., Competing Interests: Declarations Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Organelle genomes serve as crucial datasets for investigating the genetics and evolution of plants and animals, genome diversity, and species identification. To enhance the collection, analysis, and visualisation of such data, we have developed a novel open-source software tool named Organelle Genome Utilities (OGU). The software encompasses three modules designed to streamline the handling of organelle genome data. The data collection module is dedicated to retrieving, validating and organising sequence information. The evaluation module assesses sequence variance using a range of methods, including novel metrics termed stem and terminal phylogenetic diversity. The primer module designs universal primers for downstream applications. Finally, a visualisation pipeline has been developed to present comprehensive insights into organelle genomes across different lineages rather than focusing solely on individual species. The performance, compatibility and stability of OGU have been rigorously evaluated through benchmarking with four datasets, including one million mixed GenBank records, plastid genomic data from the Lamiaceae family, mitochondrial data from rodents, and 308 plastid genomes sourced from various angiosperm families. Based on software capabilities, we identified 30 plastid intergenic spacers. These spacers exhibit a moderate evolutionary rate and offer practical utility comparable to coding regions, highlighting the potential applications of intergenic spacers in organelle genomes. We anticipate that OGU will substantially enhance the efficient utilisation of organelle genomic data and broaden the prospects for related research endeavours., (© 2024 The Author(s). Molecular Ecology Resources published by John Wiley & Sons Ltd.)

Background: The genomes within organelles are crucial for physiological functions such as respiration and photosynthesis and may also contribute to environmental adaptation. However, the limited availability of genetic resources, particularly mitochondrial genomes, poses significant challenges for in-depth investigations., Results: Here, we explored various assembly methodologies and successfully reconstructed the complex organelle genomes of two Rhododendron species: Rhododendron nivale subsp. boreale and Rhododendron vialii. The mitogenomes of these species exhibit various conformations, as evidenced by long-reads mapping. Notably, only the mitogenome of R. vialii can be depicted as a singular circular molecule. The plastomes of both species conform to the typical quadripartite structure but exhibit elongated inverted repeat (IR) regions. Compared to the high similarity between plastomes, the mitogenomes display more obvious differences in structure, repeat sequences, and codon usage. Based on the analysis of 58 organelle genomes from angiosperms inhabiting various altitudes, we inferred the genetic adaptations associated with high-altitude environments. Phylogenetic analysis revealed partial inconsistencies between plastome- and mitogenome-derived phylogenies. Additionally, evolutionary lineage was determined to exert a greater influence on codon usage than altitude. Importantly, genes such as atp4, atp9, mttB, and clpP exhibited signs of positive selection in several high-altitude species, suggesting a potential link to alpine adaptation., Conclusions: We tested the effectiveness of different organelle assembly methods for dealing with complex genomes, while also providing and validating high-quality organelle genomes of two Rhododendron species. Additionally, we hypothesized potential strategies for high-altitude adaptation of organelles. These findings offer a reference for the assembly of complex organelle genomes, while also providing new insights and valuable resources for understanding their adaptive evolution patterns., (© 2024. The Author(s).)

The use of complete organelle genomes, including chloroplast and mitochondrial genomes, is a powerful molecular method for studying biological evolution and gene transfer. However, in the case of Polygonaceae, an important family with numerous edible, medicinal, and ornamental species, the mitochondrial genomes of only three species have been sequenced and analyzed. In this study, we present the mitochondrial and chloroplast genomes of two important Tibetan medicinal plants, Bistorta viviparum and B. macrophyllum. All the organelle genomes are assembled into a single circular structure and contain a common set of 32 protein-coding genes (PCGs). Some genes such as rps2 and ndhF were found to have high nucleotide polymorphism (Pi) in the chloroplast genomes, while cox1, mttB and rps12 showed pronounced Pi values in the mitochondrial genomes. Furthermore, our analysis revealed that most chloroplast genes and mitochondrial PCGs in Polygonaceae plants are under purifying selection. However, a few genes, including the chloroplast gene psaJ and the mitochondrial genes ccmFc, atp8 and nad4, showed positive selection in certain Polygonaceae plants, as indicated by a Ka/Ks ratio greater than one. Structural variation analysis revealed a wealth of differences between the mitochondrial genomes of five Polygonaceae species, with a particularly notable large-scale inversion observed between Reynoutria japonica and Fallopia aubertii. Furthermore, an analysis of the homologous sequences in the chloroplast and mitochondrial genomes revealed that the rps7 has been transferred from the chloroplast to the mitochondrial genome in all five Polygonaceae species. Finally, ecological niche models were constructed for B. viviparum and B. macrophyllum, indicating that mean annual temperature and altitude are the main climatic factors influencing the distribution of both species. Although the current distribution of B. viviparum is significantly wider than that of B. macrophyllum, projections suggest that the optimal growth ranges of both species will expand in the future, with B. macrophyllum potentially exceeding B. viviparum. This study not only contributes to the plastid genome database for Polygonaceae plants, but also provides theoretical insights into the adaptive evolution of these plants., (© 2024. The Author(s).)

Taxus plants are the exclusive source of paclitaxel, an anticancer drug with significant medicinal and economic value. Interspecies hybridization and gene introgression during evolution have obscured distinctions among Taxus species, complicating their phylogenetic classification. While the chloroplast genome of Taxus wallichiana, a widely distributed species in China, has been sequenced, its mitochondrial genome (mitogenome) remains uncharacterized.We sequenced and assembled the T. wallichiana mitogenome using BGI short reads and Nanopore long reads, facilitating comparisons with other gymnosperm mitogenomes. The T. wallichiana mitogenome spanning 469,949 bp, predominantly forms a circular configuration with a GC content of 50.51%, supplemented by 3 minor configurations mediated by one pair of LRs and two pairs of IntRs. It includes 32 protein-coding genes, 7 tRNA genes, and 3 rRNA genes, several of which exist in multiple copies.We detailed the mitogenome's structure, codon usage, RNA editing, and sequence migration between organelles, constructing a phylogenetic tree to elucidate evolutionary relationships. Unlike typical gymnosperm mitochondria, T. wallichiana shows no evidence of mitochondrial-plastid DNA transfer (MTPT), highlighting its unique genomic architecture. Synteny analysis indicated extensive genomic rearrangements in T. wallichiana, likely driven by recombination among abundant repetitive sequences. This study offers a high-quality T. wallichiana mitogenome, enhancing our understanding of gymnosperm mitochondrial evolution and supporting further cultivation and utilization of Taxus species., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The genomes of peridinin-containing dinoflagellate chloroplasts have a very unusual organisation. These genomes are highly fragmented and greatly reduced, with most of the usual complement of chloroplast genes relocated to the nucleus. Dinoflagellate chloroplasts highlight evolutionary changes that are found to varying extents in a number of other organelle genomes. These include the chloroplast genome of the green alga Boodlea and other Cladophorales, and the mitochondrial genomes of blood-sucking and chewing lice, the parasitic plant Rhopalocnemis phalloides, the red alga Rhodosorus marinus and other members of the Stylonematophyceae, diplonemid flagellates, and some Cnidaria. Consideration of the coding content of the remnant chloroplast genomes indicates that organelles may preferentially retain genes for proteins important in initiating assembly of complexes, and the same is largely true for mitochondria. We propose a new principle, of CO-location for COntrol of Assembly (COCOA), indicating the importance of retaining these genes in the organelle. This adds to, but does not invalidate, the existing hypotheses of the multisubunit completion principle, CO-location for Redox Regulation (CORR) and Control by Epistasy of Synthesis (CES)., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Cephaleuros species are well-known as plant pathogens that cause red rust or algae spot diseases in many economically cultivated plants that grow in shady and humid environments. Despite their prevalence, the adaptive evolution of these pathogens remains poorly understood. We sequenced and characterized three Cephaleuros ( Cephaleuros lagerheimii , Cephaleuros diffusus , and Cephaleuros virescens ) chloroplast genomes, and compared them with seven previously reported chloroplast genomes. The chloroplast sequences of C. lagerheimii , C. diffusus , and C. virescens were 480,613 bp, 383,846 bp, and 472,444 bp in length, respectively. These chloroplast genomes encoded 94 genes, including 27 tRNA genes, 3 rRNA genes, and 64 protein-coding genes. Comparative analysis uncovered that the variation in genome size was principally due to the length of intergenic spacer sequences, followed by introns. Furthermore, several highly variable regions ( trnY-GTA , trnL-TAG , petA , psbT , trnD-GTC , trnL-TAA , ccsA , petG , psaA , psaB , rps11 , rps2 , and rps14 ) were identified. Codon bias analysis revealed that the codon usage pattern of Cephaleuros is predominantly shaped by natural selection. Additionally, six chloroplast protein-coding genes ( atpF , chlN , psaA , psaB , psbA , and rbcL ) were determined to be under positive selection, suggesting they may play a vital roles in the adaptation of Cephaleuros to low-light intensity habitats.

Background: Primulina hunanensis, a troglobitic plant within the Primulina genus of Gesneriaceae family, exhibits robust resilience to arid conditions and holds great horticultural potential as an ornamental plant. The work of chloroplast genome (cpDNA) has been recently accomplished, however, the mitochondrial genome (mtDNA) that is crucial for plant evolution has not been reported., Results: In this study, we sequenced and assembled the P. hunanensis complete mtDNA, and elucidated its evolutionary and phylogenetic relationships. The assembled mtDNA spans 575,242 bp with 43.54% GC content, encompassing 60 genes, including 37 protein-coding genes (PCGs), 20 tRNA genes, and 3 rRNA genes. Notably, high number of repetitive sequences in the mtDNA and substantial sequence translocation from chloroplasts to mitochondria were observed. To determine the evolutionary and taxonomic positioning of P. hunanensis, a phylogenetic tree was constructed using mitochondrial PCGs from P. hunanensis and 32 other taxa. Furthermore, an exploration of PCGs relative synonymous codon usage, identification of RNA editing events, and an investigation of collinearity with closely related species were conducted., Conclusions: This study reports the initial assembly and annotation of P. hunanensis mtDNA, contributing to the limited mtDNA repository for Gesneriaceae plants and advancing our understanding of their evolution for improved utilization and conservation., (© 2024. The Author(s).)

We report the complete organellar genome sequences of an ultrasmall green alga, Medakamo hakoo strain M-hakoo 311, which has the smallest known nuclear genome in freshwater green algae. Medakamo hakoo has 90.8-kb chloroplast and 36.5-kb mitochondrial genomes containing 80 and 33 putative protein-coding genes, respectively. The mitochondrial genome is the smallest in the Trebouxiophyceae algae studied so far. The GC content of the nuclear genome is 73%, but those of chloroplast and mitochondrial genomes are 41% and 35%, respectively. Codon usages in the organellar genomes have a different tendency from that in the nuclear genome. The organellar genomes have unique characteristics, such as the biased encoding of mitochondrial genes on a single strand and the absence of operon structures in chloroplast ribosomal genes. Medakamo hakoo will be helpful for understanding the evolution of the organellar genome and the regulation of gene expression in chloroplasts and mitochondria.

Rehmannia chingii is an important medicinal plant with immense value in scientific research. However, its mitochondrial genome (mitogenome) has not yet been characterized. Herein, based on whole-genome Illumina short reads and PacBio HiFi reads, we obtained the complete mitogenome of R. chingii through a de novo assembly strategy. We carried out comparative genomic analyses and found that, in comparison with the plastid genome (plastome) showing a high degree of structural conservation, the R. chingii mitogenome structure is relatively complex, showing an intricate ring structure with 16 connections, owing to five repetitive sequences. The R. chingii mitogenome was 783,161 bp with a GC content of 44.8% and contained 77 genes, comprising 47 protein-coding genes (CDS), 27 tRNA genes, and 3 rRNA genes. We counted 579 RNA editing events in 47 CDS and 12,828 codons in all CDSs of the R. chingii mitogenome. Furthermore, 24 unique sequence transfer fragments were found between the mitogenome and plastome, comprising 8 mitogenome CDS genes and 16 plastome CDS genes, corresponding to 2.39% of the R. chingii mitogenome. Mitogenomes had shorter but more collinear regions, evidenced by a comparison of the organelles of non-parasitic R. chingii , hemiparasitic Pedicularis chinensis , and holoparasitic Aeginetia indica in the Orobanchaceae family. Moreover, from non-parasitic to holoparasitic species, the genome size in the mitogenomes of Orobanchaceae species did not decrease gradually. Instead, the smallest mitogenome was found in the hemiparasitic species P. chinensis, with a size of 225,612 bp. The findings fill the gap in the mitogenome research of the medicinal plant R. chingii , promote the progress of the organelle genome research of the Orobanchaceae family, and provide clues for molecular breeding.

Chrysanthemum indicum , a species native to Eastern Asia is well known as one of the progenitor species of the cultivated Chrysanthemum which is grown for its ornamental and medicinal value. Previous genomic studies on Chrysanthemum have largely ignored the dynamics of plastid genome (plastome) and mitochondria genome (mitogenome) evolution when analyzing this plant lineage. In this study, we sequenced and assembled the plastomes and mitogenomes of diploid and tetraploid C. indicum as well as the morphologically divergent variety C. indicum var. aromaticum . We used published data from 27 species with both plastome and mitogenome complete sequences to explore differences in sequence evolution between the organellar genomes. The size and structure of organellar genome between diploid and tetraploid C. indicum were generally similar but the tetraploid C. indicum and C. indicum var. aromaticum were found to contain unique sequences in the mitogenomes which also contained previously undescribed open reading frames (ORFs). Across Chrysanthemum mitogenome structure varied greatly but sequences transferred from plastomes in to the mitogenomes were conserved. Finally, differences observed between mitogenome and plastome gene trees may be the result of the difference in the rate of sequence evolution between genes in these two genomes. In total the findings presented here greatly expand the resources for studying Chrysanthemum organellar genome evolution with possible applications to conservation, breeding, and gene banking in the future., Competing Interests: Authors HL, LX, QM, and MW were employed by the company China Resources Sanjiu Medical & Pharmaceutical Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Liu, Hou, Xu, Ma, Wei, Tembrock, Zhang and Wu.)

Sphaeropleales have the characteristics of rapid growth, high oil content, and efficient removal rates of nitrogen and phosphorus in sewage waters, and is potentially valuable in biodiesel production and environmental remediation. In this study, we isolated a strain of Sphaeropleales, Chlorolobion braunii strain ITBB-AG6 from an azolla community in a sewage pond. Its mitochondrial genome contains 110,124 bp and harbors at least 40 genes, including 15 protein-coding genes, 20 tRNA genes, and three rRNA genes. The protein-coding genes include two for ATP synthases, seven for NAD(P)H-quinone oxidoreductases (nad), three for cytochrome c oxidase subunits (coxs), and one for cytochrome b (cob). Transfer RNA genes for 18 amino acids were identified, in which the tRNA genes for leucine and serine are doubled, but the tRNA genes for threonine and valine are not annotated. Phylogenetic analysis using the mitochondrial genomes of seven families of Sphaeropleales indicated that ITBB-AG6 is closely related to Monoraphidium neglectum , and falls in the family Selenastraceae with 100% bootstrap support. Two species in the family Neochloridaceae are separated by a species in Hydrodictyaceae, indicating a polyphyletic nature. These findings revealed the complicated phylogenetic relationships of the Sphaeropleales and the necessity of genome sequences in the taxonomy of microalgae., Competing Interests: The authors report there are no competing interests to declare., (© 2023 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.)

Main Conclusion: The first complete mitochondrial genome of Carex (C. breviculmis) was sequenced and assembled, and its genomic signature was analyzed and the possible conformations of its mitochondrial genome were validated. Carex breviculmis is a very adaptable grass that is highly resistant to environmental stresses such as drought and low light. It is also admired as a landscape plant with high development prospects and scientific research value. In this study, the mitochondrial genome of C. breviculmis was assembled using Pacbio and Illumina sequencing data. We detected 267 pairs of repeats and found that three pairs of repeats could mediate the recombination of its mitochondrial genome and formed four possible conformations, of which we verified the two conformations mediated by the shortest pair of repeats using PCR amplification and Sanger sequencing. The major conformation of the C. breviculmis mitochondrial genome is a 1,414,795 bp long circular molecule with 33 annotated protein-coding genes, 15 tRNA genes, and three rRNA genes. We detected a total of 25 homologous sequences between the chloroplast and mitochondrial genomes, corresponding to 0.40% of the mitochondrial genome. Combined with the low GC content (41.24%), we conclude that the reduction in RNA editing sites in the C. breviculmis mitochondrial genome may be due to an accumulation of point mutations in C-to-T or retroprocessing events within the genome. The relatively low number of RNA editing sites in its mitochondrial genome could serve as important material for subsequent studies on the selection pressure of RNA editing in angiosperms. A maximum likelihood analysis based on 23 conserved mitochondrial genes from 28 species reflects an accurate evolutionary and taxonomic position of C. breviculmis. This research provided us with a comprehensive understanding of the mitochondrial genome of Carex and also provided important information for its molecular breeding., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The Cyanidiophyceae, an extremophilic red algal class, is distributed worldwide in extreme environments. Species grow either in acidic hot environments or in dim light conditions (e.g., "cave Cyanidium"). The taxonomy and classification systems are currently based on morphological, eco-physiological, and molecular phylogenetic characters; however, previous phylogenetic results showed hidden diversity of the Cyanidiophyceae and suggested a revision of the classification system. To clarify phylogenetic relationships within this red algal class, we employ a phylogenomic approach based on 15 plastomes (10 new) and 15 mitogenomes (seven new). Our phylogenies show consistent relationships among four lineages (Galdieria, "cave Cyanidium", Cyanidium, and Cyanidioschyzon lineages). Each lineage is distinguished by organellar genome characteristics. The "cave Cyanidium" lineage is a distinct clade that diverged after the Galdieria clade but within a larger monophyletic clade that included the Cyanidium and Cyanidioschyzon lineages. Because the "cave Cyanidium" lineage is a mesophilic lineage that differs substantially from the other three thermoacidophilic lineages, we describe it as a new order (Cavernulicolales). Based on this evidence, we reclassified the Cyanidiophyceae into four orders: Cyanidiales, Cyanidioschyzonales, Cavernulicolales ord. nov., and Galdieriales ord. nov. The genetic distance among these four orders is comparable to, or greater than, the distances found between other red algal orders and subclasses. Three new genera (Cavernulicola, Gronococcus, Sciadococcus), five new species (Galdieria javensis, Galdieria phlegrea, Galdieria yellowstonensis, Gronococcus sybilensis, Sciadococcus taiwanensis), and a new nomenclatural combination (Cavernulicola chilensis) are proposed., (© 2023 Phycological Society of America.)

Background: Plant organelle genomes are a valuable resource for evolutionary biology research, yet their genome architectures, evolutionary patterns and environmental adaptations are poorly understood in many lineages. Rhodiola species is a type of flora mainly distributed in highland habitats, with high medicinal value. Here, we assembled the organelle genomes of three Rhodiola species (R. wallichiana, R. crenulata and R. sacra) collected from the Qinghai-Tibet plateau (QTP), and compared their genome structure, gene content, structural rearrangements, sequence transfer and sequence evolution rates., Results: The results demonstrated the contrasting evolutionary pattern between plastomes and mitogenomes in three Rhodiola species, with the former possessing more conserved genome structure but faster evolutionary rates of sequence, while the latter exhibiting structural diversity but slower rates of sequence evolution. Some lineage-specific features were observed in Rhodiola mitogenomes, including chromosome fission, gene loss and structural rearrangement. Repeat element analysis shows that the repeats occurring between the two chromosomes may mediate the formation of multichromosomal structure in the mitogenomes of Rhodiola, and this multichromosomal structure may have recently formed. The identification of homologous sequences between plastomes and mitogenomes reveals several unidirectional protein-coding gene transfer events from chloroplasts to mitochondria. Moreover, we found that their organelle genomes contained multiple fragments of nuclear transposable elements (TEs) and exhibited different preferences for TEs insertion type. Genome-wide scans of positive selection identified one gene matR from the mitogenome. Since the matR is crucial for plant growth and development, as well as for respiration and stress responses, our findings suggest that matR may participate in the adaptive response of Rhodiola species to environmental stress of QTP., Conclusion: The study analyzed the organelle genomes of three Rhodiola species and demonstrated the contrasting evolutionary pattern between plastomes and mitogenomes. Signals of positive selection were detected in the matR gene of Rhodiola mitogenomes, suggesting the potential role of this gene in Rhodiola adaptation to QTP. Together, the study is expected to enrich the genomic resources and provide valuable insights into the structural dynamics and sequence divergences of Rhodiola species., (© 2023. The Author(s).)

Plant genomes are comprised of nuclear, plastid and mitochondrial components characterized by different patterns of inheritance and evolution. Genetic markers from the three genomes provide complementary tools for investigations of inheritance, genetic relationships and phenotypic contributions. Plant mitochondrial genomes are challenging for universal marker development because they are highly variable in terms of size, gene order and intergenic sequences and highly conserved with respect to protein-coding sequences. PCR amplification of introns with primers that anneal to conserved, flanking exons is effective for the development of polymorphic nuclear genome markers. The potential for plant mitochondrial intron polymorphisms to distinguish between congeneric species or intraspecific varieties has not been systematically investigated and is possibly constrained by requirements for intron secondary structure and interactions with co-evolved organelle intron splicing factors. To explore the potential for broadly applicable plant mitochondrial intron markers, PCR primer sets based upon conserved sequences flanking 11 introns common to seven angiosperm species were tested across a range of plant orders. PCR-amplified introns were screened for indel polymorphisms among a group of cross-compatible Citrus species and relatives; two Raphanus sativus mitotypes; representatives of the two Phaseolus vulgaris gene pools; and congeneric pairs of Cynodon , Cenchrus , Solanum , and Vaccinium species. All introns were successfully amplified from each plant entry. Length polymorphisms distinguishable by gel electrophoresis were common among genera but infrequent within genera. Sequencing of three introns amplified from 16 entries identified additional short indel polymorphisms and nucleotide substitutions that separated Citrus , Cynodon , Cenchrus and Vaccinium congeners, but failed to distinguish Solanum congeners or representatives of the Phaseolus vulgaris major gene pools. The ability of primer sets to amplify a wider range of plant species' introns and the presence of intron polymorphisms that distinguish congeners was confirmed by in silico analysis. While mitochondrial intron variation is limited in comparison to nuclear introns, these exon-based primer sets provide robust tools for the amplification of mitochondrial introns across a wide range of plant species wherein useful polymorphisms can be identified., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Grosser, Sites, Murata, Lopez, Chamusco, Love Harriage, Grosser, Graham, Gmitter and Chase.)

Clematis is one of the large worldwide genera of the Ranunculaceae Juss. Family, with high ornamental and medicinal value. China is the modern distribution centre of Clematis with abundant natural populations. Due to the complexity and high morphological diversity of Clematis , the genus is difficult to classify systematically, and in particular, the phylogenetic position of the endangered Clematis acerifolia is highly controversial. The use of the mitochondrial complete genome is a powerful molecular method that is frequently used for inferring plants phylogenies. However, studies on Clematis mitogenome are rare, thus limiting our full understanding of its phylogeny and genome evolution. Here, we sequenced and annotated the C. acerifolia mt genome using Illumina short- and Nanopore long-reads, characterized the species first complete mitogenome, and performed a comparative phylogenetic analysis with its close relatives. The total length of the C. acerifolia mitogenome is 698,247 bp and the main structure is multi-branched (linear molecule 1 and circular molecule 2). We annotated 55 genes, including 35 protein-coding, 17 tRNA, and 3 rRNA genes. The C. acerifolia mitogenome has extremely unconserved structurally, with extensive sequence transfer between the chloroplast and mitochondrial organelles, sequence repeats, and RNA editing. The phylogenetic position of C. acerifolia was determined by constructing the species mitogenome with 24 angiosperms. Further, our C. acerifolia mitogenome characteristics investigation included GC contents, codon usage, repeats and synteny analysis. Overall, our results are expected to provide fundamental information for C. acerifolia mitogenome evolution and confirm the validity of mitochondrial analysis in determining the phylogenetic positioning of Clematis plants., Competing Interests: YY was employed by Wuhan Benagen Technology Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Liu, Qu, Yuan, Zhao, Chen, Han, Li, El-Kassaby, Yin, Xie, Tong and Liu.)

Pectinodesmus pectinatus is a green alga of commercial interest in sewage purification. Clarification of its organelle genomes is helpful for genetic manipulation, taxonomic revisions and evolutionary research. Here, de novo sequencing was used to determine chloroplast genome and mitochondrial genome of P. pectinatus strain F34. The chloroplast genome was composed of a large single-copy (LSC) region of 99,156 bp, a small single-copy (SSC) region of 70,665 bp, and a pair of inverted repeats (IRs) with a length of 13,494 bp each separated by LSC and SSC. The chloroplast genome contained 69 protein-coding genes, 25 transfer-RNA (tRNA) genes, 3 ribosomal RNA (rRNA) genes. The mitochondrial genome was 32,195 bp in length and consisted of 46 unique genes, including 16 protein-coding genes, 27 tRNA genes and 3 rRNA genes. The predominant mutations in organelle genomes were T/A to G/C transitions. Phylogenic analysis indicated P. pectinatus was a sister species to Tetradesmus obliquus and Hariotina sp. within the Pectinodesmus genus. In analysis with CGView Comparison Tool, P. pectinatus organelle genomes displayed the highest sequence similarity with that of T. obliquus . These findings advanced research on the taxonomy and phylogeny of Chlorophyceae algae and particularly revealed the role of P. pectinatus in microalgae evolution.

Background: Okra (Abelmoschus esculentus L. Moench) is an economically important crop and is known for its slimy juice, which has significant scientific research value. The A. esculentus chloroplast genome has been reported; however, the sequence of its mitochondrial genome is still lacking., Results: We sequenced the plastid and mitochondrial genomes of okra based on Illumina short reads and Nanopore long reads and conducted a comparative study between the two organelle genomes. The plastid genome of okra is highly structurally conserved, but the mitochondrial genome of okra has been confirmed to have abundant subgenomic configurations. The assembly results showed that okra's mitochondrial genome existed mainly in the form of two independent molecules, which could be divided into four independent molecules through two pairs of long repeats. In addition, we found that four pairs of short repeats could mediate the integration of the two independent molecules into one complete molecule at a low frequency. Subsequently, we also found extensive sequence transfer between the two organelles of okra, where three plastid-derived genes (psaA, rps7 and psbJ) remained intact in the mitochondrial genome. Furthermore, psbJ, psbF, psbE and psbL were integrated into the mitochondrial genome as a conserved gene cluster and underwent pseudogenization as nonfunctional genes. Only psbJ retained a relatively complete sequence, but its expression was not detected in the transcriptome data, and we speculate that it is still nonfunctional. Finally, we characterized the RNA editing events of protein-coding genes located in the organelle genomes of okra., Conclusions: In the current study, our results not only provide high-quality organelle genomes for okra but also advance our understanding of the gene dialogue between organelle genomes and provide information to breed okra cultivars efficiently., (© 2022. The Author(s).)

Modification of plant mitochondrial genomes is still a difficult task, especially in multicellular plants. Transcription activator-like effector nucleases with a mitochondrial localization signal (mitoTALENs) can cut out a desired sequence from the mitochondrial genome in plants. Although vector construction of mitoTALENs is complicated, the modification efficiency is high enough to achieve homoplasmy of multicopy mitochondrial genomes. Here I describe how to design mitoTALENs to select a target, construct a vector, and select the mitochondrial transformants., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

The complete chloroplast and mitochondrial genome sequences of Scopelophila cataractae (Pottiaceae, Bryophyta) are determined. The chloroplast genome is 122,290 bp with 118 genes and the mitochondrial genome is 105,607 bp with 67 genes, both genomes are circular. This study showed the S. cataractae plastome contains the smallest genome size, and a functional trnP GGG gene, relative to other pottiaceous species. Phylogenetic inferences support the sister relationship of S. cataractae to all other pottiaceous accessions., Competing Interests: No potential conflict of interest was reported by the author(s)., (© 2021 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.)

B. oleracea Ogura CMS is an alloplasmic male-sterile line introduced from radish by interspecific hybridization and protoplast fusion. The introduction of alien cytoplasm resulted in many undesirable traits, which affected the yield of hybrids. Therefore, it is necessary to identify the composition and reduce the content of alien cytoplasm in B. oleracea Ogura CMS. In the present study, we sequenced, assembled, and compared the organelle genomes of Ogura CMS cabbage and its maintainer line. The chloroplast genome of Ogura-type cabbage was completely derived from normal-type cabbage, whereas the mitochondrial genome was recombined from normal-type cabbage and Ogura-type radish. Nine unique regions derived from radish were identified in the mitochondrial genome of Ogura-type cabbage, and the total length of these nine regions was 35,618 bp, accounting for 13.84% of the mitochondrial genome. Using 32 alloplasmic markers designed according to the sequences of these nine regions, one novel sterile source with less alien cytoplasm was discovered among 305 materials and named Bel CMS. The size of the alien cytoplasm in Bel CMS was 21,587 bp, accounting for 8.93% of its mtDNA, which was much less than that in Ogura CMS. Most importantly, the sterility gene orf138 was replaced by orf112 , which had a 78-bp deletion, in Bel CMS. Interestingly, Bel CMS cabbage also maintained 100% sterility, although orf112 had 26 fewer amino acids than orf138 . Field phenotypic observation showed that Bel CMS was an excellent sterile source with stable 100% sterility and no withered buds at the early flowering stage, which could replace Ogura CMS in cabbage heterosis utilization.

Allopolyploidy is widely present across plant lineages. Though estimating the correct phylogenetic relationships and origin of allopolyploids may sometimes become a hard task. In the genus Stylosanthes Sw. (Leguminosae), an important legume crop, allopolyploidy is a key speciation force. This makes difficult adequate species recognition and breeding efforts on the genus. Based on comparative analysis of nine high-throughput sequencing (HTS) samples, including three allopolyploids ( S . capitata Vogel cv. "Campo Grande," S . capitata "RS024" and S . scabra Vogel) and six diploids ( S . hamata Taub, S . viscosa (L.) Sw., S . macrocephala M. B. Ferreira and Sousa Costa, S . guianensis (Aubl.) Sw., S . pilosa M. B. Ferreira and Sousa Costa and S . seabrana B. L. Maass & 't Mannetje) we provide a working pipeline to identify organelle and nuclear genome signatures that allowed us to trace the origin and parental genome recognition of allopolyploids. First, organelle genomes were de novo assembled and used to identify maternal genome donors by alignment-based phylogenies and synteny analysis. Second, nuclear-derived reads were subjected to repetitive DNA identification with RepeatExplorer2. Identified repeats were compared based on abundance and presence on diploids in relation to allopolyploids by comparative repeat analysis. Third, reads were extracted and grouped based on the following groups: chloroplast, mitochondrial, satellite DNA, ribosomal DNA, repeat clustered- and total genomic reads. These sets of reads were then subjected to alignment and assembly free phylogenetic analyses and were compared to classical alignment-based phylogenetic methods. Comparative analysis of shared and unique satellite repeats also allowed the tracing of allopolyploid origin in Stylosanthes , especially those with high abundance such as the StyloSat1 in the Scabra complex. This satellite was in situ mapped in the proximal region of the chromosomes and made it possible to identify its previously proposed parents. Hence, with simple genome skimming data we were able to provide evidence for the recognition of parental genomes and understand genome evolution of two Stylosanthes allopolyploids., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Oliveira, Nunes, Dos Santos, Ferreira Gomes, Costa, Van-Lume, Marques Da Silva, Oliveira, Simon, Lima, Gissi, Almeida, Souza and Marques.)

Besides the nuclear genome, plants possess two small extra chromosomal genomes in mitochondria and chloroplast, respectively, which contribute a small fraction of the organelles' proteome. Both mitochondrial and chloroplast DNA have originated endosymbiotically and most of their prokaryotic genes were either lost or transferred to the nuclear genome through endosymbiotic gene transfer during the course of evolution. Due to their immobile nature, plant nuclear and organellar genomes face continuous threat from diverse exogenous agents as well as some reactive by-products or intermediates released from various endogenous metabolic pathways. These factors eventually affect the overall plant growth and development and finally productivity. The detailed mechanism of DNA damage response and repair following accumulation of various forms of DNA lesions, including single and double-strand breaks (SSBs and DSBs) have been well documented for the nuclear genome and now it has been extended to the organelles also. Recently, it has been shown that both mitochondria and chloroplast possess a counterpart of most of the nuclear DNA damage repair pathways and share remarkable similarities with different damage repair proteins present in the nucleus. Among various repair pathways, homologous recombination (HR) is crucial for the repair as well as the evolution of organellar genomes. Along with the repair pathways, various other factors, such as the MSH1 and WHIRLY family proteins, WHY1, WHY2, and WHY3 are also known to be involved in maintaining low mutation rates and structural integrity of mitochondrial and chloroplast genome. SOG1, the central regulator in DNA damage response in plants, has also been found to mediate endoreduplication and cell-cycle progression through chloroplast to nucleus retrograde signaling in response to chloroplast genome instability. Various proteins associated with the maintenance of genome stability are targeted to both nuclear and organellar compartments, establishing communication between organelles as well as organelles and nucleus. Therefore, understanding the mechanism of DNA damage repair and inter compartmental crosstalk mechanism in various sub-cellular organelles following induction of DNA damage and identification of key components of such signaling cascades may eventually be translated into strategies for crop improvement under abiotic and genotoxic stress conditions. This review mainly highlights the current understanding as well as the importance of different aspects of organelle genome maintenance mechanisms in higher plants., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Mahapatra, Banerjee, De, Mitra, Roy and Roy.)

Thorea hispida exclusively inhabits freshwater environments and is characterized by a triphasic life history. In this study, the organelle genomes and transcriptomes of different life history stages of T. hispida were examined using next generation sequencing. The chloroplast and mitochondrial genomes of the chantransia stage were 175,747 and 25,411 bp in length, respectively. The chantransia stage was highly similar to the gametophyte stage based on comparisons of organelle genomes and phylogenetic reconstruction. Transcriptomic comparisons of two stages found that ribosome-related genes were the most up-regulated in the gametophyte stage of T. hispida. Seven meiosis-specific genes, including SPO11 initiator of meiotic double-stranded breaks(spo11), meiotic nuclear divisions 1(mnd1), RAD51 recombinase(rad51), mutS homolog 4(msh4), mutS homolog 5(msh5), REC8 meiotic recombination protein(rec8), and DNA helicase Mer3(mer3), were differentially regulated between the two life history stages. The organelle genomes and transcriptomes from T. hispida provided in this study will be valuable for future studies of freshwater red algae., (Copyright © 2020 Elsevier Inc. All rights reserved.)

In the present work, a barcode-DNA analysis method is described for the detection of plant oil adulteration in milk and dairy products. The method relies on the fact that plant DNA should not be present in readily detectable amounts in a dairy product unless it contains undeclared plant material. Thus, a universal plant barcode is chosen as the target to be amplified from dairy samples. Accordingly, barcode PCR-CE (PCR-capillary electrophoresis) assays are described, which do not require preliminary information on the species source of the adulterant oil type. Two PCR-CE assays, one operating on the plastid trnL (UAA) intron and the other targeting its inner P6 loop in nested format, were shown to detect corn, soybean, rapeseed and sunflower oils in clarified butter, milk and yogurt. Both barcodes are robustly amplified with extremely conserved primers. While the intron provides the species discrimination ability, the P6 loop provides superior detection sensitivity., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

GetOrganelle is a state-of-the-art toolkit to accurately assemble organelle genomes from whole genome sequencing data. It recruits organelle-associated reads using a modified "baiting and iterative mapping" approach, conducts de novo assembly, filters and disentangles the assembly graph, and produces all possible configurations of circular organelle genomes. For 50 published plant datasets, we are able to reassemble the circular plastomes from 47 datasets using GetOrganelle. GetOrganelle assemblies are more accurate than published and/or NOVOPlasty-reassembled plastomes as assessed by mapping. We also assemble complete mitochondrial genomes using GetOrganelle. GetOrganelle is freely released under a GPL-3 license ( https://github.com/Kinggerm/GetOrganelle ).

Utricularia amethystina Salzm. ex A.St.-Hil. & Girard (Lentibulariacea e ) is a highly polymorphic carnivorous plant taxonomically rearranged many times throughout history. Herein, the complete chloroplast genomes (cpDNA) of three U. amethystina morphotypes: purple-, white-, and yellow-flowered, were sequenced, compared, and putative markers for systematic, populations, and evolutionary studies were uncovered. In addition, RNA-Seq and RNA-editing analysis were employed for functional cpDNA evaluation. The cpDNA of three U. amethystina morphotypes exhibits typical quadripartite structure. Fine-grained sequence comparison revealed a high degree of intraspecific genetic variability in all morphotypes, including an exclusive inversion in the psb M and pet N genes in U. amethystina yellow. Phylogenetic analyses indicate that U. amethystina morphotypes are monophyletic. Furthermore, in contrast to the terrestrial Utricularia reniformis cpDNA, the U. amethystina morphotypes retain all the plastid NAD(P)H-dehydrogenase ( ndh ) complex genes. This observation supports the hypothesis that the ndh s in terrestrial Utricularia were independently lost and regained, also suggesting that different habitats (aquatic and terrestrial) are not related to the absence of Utricularia ndh s gene repertoire as previously assumed. Moreover, RNA-Seq analyses recovered similar patterns, including nonsynonymous RNA-editing sites (e.g., rps14 and petB ). Collectively, our results bring new insights into the chloroplast genome architecture and evolution of the photosynthesis machinery in the Lentibulariaceae.

Background: Land plant organellar genomes have significant impact on metabolism and adaptation, and as such, accurate assembly and annotation of plant organellar genomes is an important tool in understanding the evolutionary history and interactions between these genomes. Intracellular DNA transfer is ongoing between the nuclear and organellar genomes, and can lead to significant genomic variation between, and within, species that impacts downstream analysis of genomes and transcriptomes., Results: In order to facilitate further studies of cytonuclear interactions in Eucalyptus, we report an updated annotation of the E. grandis plastid genome, and the second sequenced and annotated mitochondrial genome of the Myrtales, that of E. grandis. The 478,813 bp mitochondrial genome shows the conserved protein coding regions and gene order rearrangements typical of land plants. There have been widespread insertions of organellar DNA into the E. grandis nuclear genome, which span 141 annotated nuclear genes. Further, we identify predicted editing sites to allow for the discrimination of RNA-sequencing reads between nuclear and organellar gene copies, finding that nuclear copies of organellar genes are not expressed in E. grandis., Conclusions: The implications of organellar DNA transfer to the nucleus are often ignored, despite the insight they can give into the ongoing evolution of plant genomes, and the problems they can cause in many applications of genomics. Future comparisons of the transcription and regulation of organellar genes between Eucalyptus genotypes may provide insight to the cytonuclear interactions that impact economically important traits in this widely grown lignocellulosic crop species.

A genome-wide detection of phylogenetic signals by next generation sequencing (NGS) has recently emerged as a promising genomic approach for phylogenetic analysis of non-model organisms. Here we explored the use of a multiplexed shotgun sequencing method to assess the phylogenetic relationships of 18 Linum samples representing 16 species within four botanical sections of the flax genus Linum. The whole genome DNAs of 18 Linum samples were fragmented, tagged, and sequenced using an Illumina MiSeq. Acquired sequencing reads per sample were further separated into chloroplast, mitochondrial and nuclear sequence reads. SNP calls upon genome-specific sequence data sets revealed 6143 chloroplast, 2673 mitochondrial, and 19,562 nuclear SNPs. Phylogenetic analyses based on three-genome SNP data sets with and without missing observations showed congruent three-genome phylogenetic signals for four botanical sections of the Linum genus. Specifically, two major lineages showing a separation of Linum-Dasylinum sections and Linastrum-Syllinum sections were confirmed. The Linum section displayed three major branches representing two major evolutionary stages leading to cultivated flax. Cultivated flax and its immediate progenitor were formed as its own branch, genetically more closely related to L. decumbens and L. grandiflorum with chromosome count of eight, and distantly apart from six other species with chromosome count of nine. Five species of the Linastrum and Syllinum sections were genetically more distant from cultivated flax, but they appeared to be more closely related to each other, even with variable chromosome counts. These findings not only provide the first evidence of congruent three-genome phylogenetic pathways within the Linum genus, but also demonstrate the utility of the multiplexed shotgun sequencing in acquisition of three-genome phylogenetic signals of non-model organisms., (Crown Copyright © 2016. Published by Elsevier Inc. All rights reserved.)

A lot is known about the evolution and architecture of plastid, mitochondrial, and nuclear genomes, but surprisingly little is known about their relative rates of mutation. Most available relative-rate data come from seed plants, which, with few exceptions, have a mitochondrial mutation rate that is lower than those of the plastid and nucleus. But new findings from diverse plastid-bearing lineages have shown that for some eukaryotes the mitochondrial mutation rate is an order of magnitude greater than those of the plastid and nucleus. Here, we explore for the first time relative rates of mutation within the Glaucophyta-one of three main lineages that make up the Archaeplastida (or Plantae sensu lato). Nucleotide substitution analyses from distinct isolates of the unicellular glaucophyte Cyanophora paradoxa reveal 4-5-fold lower rates of mutation in the plastid and nucleus than the mitochondrion, which is similar to the mutational pattern observed in red algae and haptophytes, but opposite to that of seed plants. These data, together with data from previous reports, suggest that for much of the known photosynthetic eukaryotic diversity, plastid DNA mutations occur less frequently than those in mitochondrial DNA., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Eighteen classical mutant strains of Neurospora crassa were subject to whole genome sequence analysis and the mitochondrial genome is analyzed. Overall, the mitochondrial genomes of the classical mutant strains are 99.45 to 99.98 % identical to the reference genome. Two-thirds of the SNPs and three-fourths of indels identified in this analysis are shared among more than one strain. Most of the limited variability in mitochondrial genome sequence is neutral with regard to protein structure. Despite the fact that the mitochondrial genome is present in multiple copies per cell, many of the polymorphisms were homozygous within each strain. Conversely, some polymorphisms, especially those associated with large scale rearrangements are only present in a fraction of the reads covering each region. The impact of this variation is unknown and further studies will be necessary to ascertain if this level of polymorphism is common among fungi and whether it reflects the impact of ageing cultures.