Synonymous Changes Research Articles

Case report: Multiple approach analysis in a case of clinically assessed myotonia congenita

Myotonia congenita, both in a dominant (Thomsen disease) and recessive form (Becker disease), is caused by molecular defects in CLCN1 that encodes the major skeletal muscle chloride channel, ClC-1. This channel is important for the normal repolarization of muscle action potentials and consequent relaxation of the muscle, and its dysfunction leads to impaired muscle relaxation after voluntary or evoked contraction and muscle stiffness. More than 300 CLCN1 pathogenic variants have been found in association with congenital myotonia, inherited as recessive or dominant traits (with complete or incomplete penetrance). In this study, we describe the case of a 44-year-old woman complaining of “leg stiffness” since the age of 20 years and presenting with transient muscle weakness, especially after sitting for several minutes, with grip myotonia and feet myotonia, cold-sensitive and warm-up. The strength was normal, but muscle hypertrophy in the lower limbs was evident. EMG myotonia was detected in all explored muscles. The patient’s father had precocious cataract correction but did not show myotonic discharges at EMG. Examination of the patient’s sons (aged 18 years and 12 years) was unremarkable. The patient started treatment with mexiletine, with improvement in grip myotonia and limb stiffness, but it was soon interrupted due to gastrointestinal disturbances. Direct sequencing of CLCN1 identified the previously described heterozygous intronic variant c.1471 + 1G > A, which resulted in the skipping of exon 13 in the CLCN1 muscle transcript. In addition, the rare heterozygous synonymous nucleotide change c.762C > T p.Cys254Cys was identified and predicted to alter physiological splicing. The detection of multiple splicing abnormalities leading to premature termination codons supported the in silico prediction. We developed a Western blot assay to assess the ClC-1 protein in muscle biopsy, and we observed that ClC-1 levels were consistently reduced in the patient’s muscle, supporting the pathogenic behavior of the variants disclosed. Overall, we report a novel case of Becker myotonia and highlight the importance of multiple levels of analysis to achieve a firm molecular diagnosis.

In depth sequencing of a serially sampled household cohort reveals the within-host dynamics of Omicron SARS-CoV-2 and rare selection of novel spike variants.

SARS-CoV-2 has undergone repeated and rapid evolution to circumvent host immunity. However, outside of prolonged infections in immunocompromised hosts, within-host positive selection has rarely been detected. The low diversity within-hosts and strong genetic linkage among genomic sites make accurately detecting positive selection difficult. Longitudinal sampling is a powerful method for detecting selection that has seldom been used for SARS-CoV-2. Here we combine longitudinal sampling with replicate sequencing to increase the accuracy of and lower the threshold for variant calling. We sequenced 577 specimens from 105 individuals from a household cohort primarily during the BA.1/BA.2 variant period. There was extremely low diversity and a low rate of divergence. Specimens had 0-12 intrahost single nucleotide variants (iSNV) at >0.5% frequency, and the majority of the iSNV were at frequencies <2%. Within-host dynamics were dominated by genetic drift and purifying selection. Positive selection was rare but highly concentrated in spike. Two individuals with BA.1 infections had S:371F, a lineage defining substitution for BA.2. A Wright Fisher Approximate Bayesian Computational model identified positive selection at 14 loci with 7 in spike, including S:448 and S:339. We also detected significant genetic hitchhiking between synonymous changes and nonsynonymous iSNV under selection. The detectable immune-mediated selection may be caused by the relatively narrow antibody repertoire in individuals during the early Omicron phase of the SARS-CoV-2 pandemic. As both the virus and population immunity evolve, understanding the corresponding shifts in SARS-CoV-2 within-host dynamics will be important.

In silico framework for genome analysis

Genomes hold the complete genetic information of an organism. Examining and analyzing genomic data plays a critical role in properly understanding an organism, particularly the main characteristics, functionalities, and evolving nature of harmful viruses. However, the rapid increase in genomic data poses new challenges and demands for extracting meaningful and valuable insights from large and complex genomic datasets. In this paper, a novel Framework for Genome Data Analysis (F4GDA), is developed that offers various methods for the analysis of viral genomic data in various forms. The framework’s methods can not only analyze the changes in genomes but also various genome contents. As a case study, the genomes of five SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) VoC (variants of concern), which are divided into three types/groups on the basis of geographical locations, are analyzed using this framework to investigate (1) the nucleotides, amino acids and synonymous codon changes in the whole genomes of VoC as well as in the Spike (S) protein, (2) whether different environments affect the rate of changes in genomes, (3) the variations in nucleotide bases, amino acids, and codon base compositions in VoC genomes, and (4) to compare VoC genomes with the reference genome sequence of SARS-CoV-2.

Whole Mitochondrial Genome Sequencing Analysis of Canine Testicular Tumours.

Currently, the molecular background based on mitochondrial DNA (mtDNA) analysis of canine testicular tumours is underestimated. The available data mostly focus on histopathological evaluations, with a few reports of nuclear genome (nDNA) studies. Tumourigenesis represents a highly complex and diverse genetic disorder, which can also encompass defects in mtDNA. The aim of this study was to identify molecular changes in whole mitochondrial genome sequences obtained from dogs affected by testicular tumours. Samples of blood, tumour, and healthy tissue were collected from each animal, and mtDNA (ultimately 45 samples) was subsequently sequenced. Thereafter, protein analyses were performed to assess the impact of the identified molecular alterations on the amino acid level. The total number of observed changes included 722 SNPs, 12 mutations, 62 indels, 5 indel mutations, and 35 heteroplasmic sites. The highest number of mtDNA variants in protein-coding genes COX1, COX3, ATP6, ND1, ND4, and ND5 was observed. Interestingly, SNPs were found in 10 out of 22 tRNA genes. Most of the identified mtDNA defects were synonymous changes at the amino acid level. Also, polymorphisms and heteroplasmy were frequently observed in the variable number of tandem repeat (VNTR) regions, especially in its fragment spanning 16,138-16,358 bp. Based on the obtained results, it was possible to select 11 polymorphisms that occurred in all the tested samples (benign, malignant) and an additional five SNPs identified only in benign neoplasms. The comprehensive analysis of malignant testicular tumours demonstrated a significant diversity in their molecular profiles, with changes ranging from 17 to 101 per sample.

Mitogenomics of a declining species, the Rio Grande silvery minnow (Hybognathus amarus), with boom–bust population dynamics

Abstract Large mitochondrial DNA fragments and nearly complete mitogenomes were obtained from nextera-tagmented reductively amplified DNA (nextRAD) data from the Rio Grande silvery minnow (Hybognathus amarus). We used these data to characterize variation across the mitogenome and to evaluate changes in mitochondrial diversity across a 20 year time series punctuated by marked species decline. We observed different evolutionary rates among protein-coding genes and inflated values of the ratio of non-synonymous change to non-synonymous sites to synonymous changes per synonymous site for several protein-coding genes, violating assumptions of selective neutrality. Analyses suggested that non-synonymous substitutions are effectively neutral, and our results were not consistent with genetic draft. Mitogenomes revealed large long-term effective population size for Rio Grande silvery minnow, and time-series data indicated recent reductions in mitochondrial diversity via genetic drift associated with population bottlenecks. The results presented in this study show that in some cases, nextRAD sequences contain informative mitochondrial DNA data.

Pandagma: a tool for identifying pan-gene sets and gene families at desired evolutionary depths and accommodating whole-genome duplications.

Identification of allelic or corresponding genes (pan-genes) within a species or genus is important for discovery of biologically significant genetic conservation and variation. Similarly, identification of orthologs (gene families) across wider evolutionary distances is important for understanding the genetic basis for similar or differing traits. Especially in plants, several complications make identification of pan-genes and gene families challenging, including whole-genome duplications, evolutionary rate differences among lineages, and varying qualities of assemblies and annotations. Here, we document and distribute a set of workflows that we have used to address these problems. Pandagma is a set of configurable workflows for identifying and comparing pan-gene sets and gene families for annotation sets from eukaryotic genomes, using a combination of homology, synteny, and expected rates of synonymous change in coding sequence. The Pandagma workflows, example configurations, implementation details, and scripts for retrieving public datasets, are available at https://github.com/legumeinfo/pandagma.

Chikungunya Virus RNA Secondary Structures Impact Defective Viral Genome Production.

Chikungunya virus (CHIKV) is a mosquito-borne RNA virus that poses an emerging threat to humans. In a manner similar to other RNA viruses, CHIKV encodes an error-prone RNA polymerase which, in addition to producing full-length genomes, gives rise to truncated, non-functional genomes, which have been coined defective viral genomes (DVGs). DVGs have been intensively studied in the context of therapy, as they can inhibit viral replication and dissemination in their hosts. In this work, we interrogate the influence of viral RNA secondary structures on the production of CHIKV DVGs. We experimentally map RNA secondary structures of the CHIKV genome using selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP), which couples chemical labelling with next-generation sequencing. We correlate the inferred secondary structure with preferred deletion sites of CHIKV DVGs. We document an increased probability of DVG generation with truncations at unpaired nucleotides within the secondary structure. We then generated a CHIKV mutant bearing synonymous changes at the nucleotide level to disrupt the existing RNA secondary structure (CHIKV-D2S). We show that CHIKV-D2S presents altered DVG generation compared to wild-type virus, correlating with the change in RNA secondary structure obtained by SHAPE-MaP. Our work thus demonstrates that RNA secondary structure impacts CHIKV DVG production during replication.

Synthetic genomes unveil the effects of synonymous recoding.

Engineering the genetic code of an organism provides the basis for (i) making any organism safely resistant to natural viruses and (ii) preventing genetic information flow into and out of genetically modified organisms while (iii) allowing the biosynthesis of genetically encoded unnatural polymers1-4. Achieving these three goals requires the reassignment of multiple of the 64 codons nature uses to encode proteins. However, synonymous codon replacement-recoding-is frequently lethal, and how recoding impacts fitness remains poorly explored. Here, we explore these effects using whole-genome synthesis, multiplexed directed evolution, and genome-transcriptome-translatome-proteome co-profiling on multiple recoded genomes. Using this information, we assemble a synthetic Escherichia coli genome in seven sections using only 57 codons to encode proteins. By discovering the rules responsible for the lethality of synonymous recoding and developing a data-driven multi-omics-based genome construction workflow that troubleshoots synthetic genomes, we overcome the lethal effects of 62,007 synonymous codon swaps and 11,108 additional genomic edits. We show that synonymous recoding induces transcriptional noise including new antisense RNAs, leading to drastic transcriptome and proteome perturbation. As the elimination of select codons from an organism's genetic code results in the widespread appearance of cryptic promoters, we show that synonymous codon choice may naturally evolve to minimize transcriptional noise. Our work provides the first genome-scale description of how synonymous codon changes influence organismal fitness and paves the way for the construction of functional genomes that provide genetic firewalls from natural ecosystems and safely produce biopolymers, drugs, and enzymes with an expanded chemistry.

FANCA c.3624C>T (p.Ser1208=) is a hypomorphic splice variant associated with delayed onset of Fanconi anemia

AbstractFanconi anemia (FA) is a hereditary, DNA repair deficiency disorder caused by pathogenic variants in any 1 of 22 known genes (FANCA-FANCW). Variants in FANCA account for nearly two-thirds of all patients with FA. Clinical presentation of FA can be heterogeneous and include congenital abnormalities, progressive bone marrow failure, and predisposition to cancer. Here, we describe a relatively mild disease manifestation among 6 individuals diagnosed with FA, each compound heterozygous for 1 established pathogenic FANCA variant and 1 FANCA exon 36 variant, c.3624C>T. These individuals had delayed onset of hematological abnormalities, increased survival, reduced incidence of cancer, and improved fertility. Although predicted to encode a synonymous change (p.Ser1208=), the c.3624C>T variant causes a splicing error resulting in a FANCA transcript missing the last 4 base pairs of exon 36. Deep sequencing and quantitative reverse transcription polymerase chain reaction analysis revealed that 6% to 10% of the FANCA transcripts included the canonical splice product, which generated wild-type FANCA protein. Consistently, functional analysis of cell lines from the studied individuals revealed presence of residual FANCD2 ubiquitination and FANCD2 foci formation, better cell survival, and decreased late S/G2 accumulation in response to DNA interstrand cross-linking agent, indicating presence of residual activity of the FA repair pathway. Thus, the c.3624C>T variant is a hypomorphic allele, which contributes to delayed manifestation of FA disease phenotypes in individuals with at least 1 c.3624C>T allele.

Influence of the mRNA initial region on protein production: a case study using recombinant detoxified pneumolysin as a model.

Recombinant proteins are of great importance in modern society, mostly as biopharmaceutical products. However, challenging and complex processes with low production yield are major drawbacks. Normally, the optimization to overcome these obstacles is focused on bioreactor and purification processes, and the biomolecular aspects are neglected, seen as less important. In this work, we present how the 5' mRNA secondary structure region can be relevant for translation and, therefore, protein production. For this, Escherichia coli BL21(DE3) clones, producing recombinant detoxified pneumolysin (PdT) with and without the N-terminal His-tag, were cultivated in 10-L bioreactors. Another version of the pdt gene (version 2) with synonymous changes in the 5'-end nucleotide sequence was also obtained. Protein production, plasmid stability, carbon sources, and acetic acid were quantified during the cultures. Furthermore, in silico mRNA analyses were performed using TIsigner and RNAfold. The results showed that the His-tag presence at the N-terminus generated a minimum 1.5-fold increase in target protein synthesis, which was explained by the in silico mRNA analyses that returned an mRNA secondary structure easier to translate and, therefore, higher protein production than without the His-tag. The pdt gene version 2 showed lower 5' mRNA opening energy than version 1, allowing higher PdT production even without a tag. This work reveals that simple mRNA analyses during heterologous gene design and production steps can help reach high-recombinant protein titers in a shorter time than using only traditional bioprocess optimization strategies.

Whole-genome comparison using complete genomes from Campylobacter fetus strains revealed single nucleotide polymorphisms on non-genomic islands for subspecies differentiation.

Bovine Genital Campylobacteriosis (BGC), caused by Campylobacter fetus subsp. venerealis, is a sexually transmitted bacterium that significantly impacts cattle reproductive performance. However, current detection methods lack consistency and reliability due to the close genetic similarity between C. fetus subsp. venerealis and C. fetus subsp. fetus. Therefore, this study aimed to utilize complete genome analysis to distinguish genetic features between C. fetus subsp. venerealis and other subspecies, thereby enhancing BGC detection for routine screening and epidemiological studies. This study reported the complete genomes of four C. fetus subsp. fetus and five C. fetus subsp. venerealis, sequenced using long-read sequencing technologies. Comparative whole-genome analyses (n = 25) were conducted, incorporating an additional 16 complete C. fetus genomes from the NCBI database, to investigate the genomic differences between these two closely related C. fetus subspecies. Pan-genomic analyses revealed a core genome consisting of 1,561 genes and an accessory pangenome of 1,064 genes between the two C. fetus subspecies. However, no unique predicted genes were identified in either subspecies. Nonetheless, whole-genome single nucleotide polymorphisms (SNPs) analysis identified 289 SNPs unique to one or the C. fetus subspecies. After the removal of SNPs located on putative genomic islands, recombination sites, and those causing synonymous amino acid changes, the remaining 184 SNPs were functionally annotated. Candidate SNPs that were annotated with the KEGG "Peptidoglycan Biosynthesis" pathway were recruited for further analysis due to their potential association with the glycine intolerance characteristic of C. fetus subsp. venerealis and its biovar variant. Verification with 58 annotated C. fetus genomes, both complete and incomplete, from RefSeq, successfully classified these seven SNPs into two groups, aligning with their phenotypic identification as CFF (Campylobacter fetus subsp. fetus) or CFV/CFVi (Campylobacter fetus subsp. venerealis and its biovar variant). Furthermore, we demonstrated the application of mraY SNPs for detecting C. fetus subspecies using a quantitative PCR assay. Our results highlighted the high genetic stability of C. fetus subspecies. Nevertheless, Campylobacter fetus subsp. venerealis and its biovar variants encoded common SNPs in genes related to glycine intolerance, which differentiates them from C. fetus subsp. fetus. This discovery highlights the potential of employing a multiple-SNP assay for the precise differentiation of C. fetus subspecies.

Characterization of the novel HLA-B*15:01:75 allele identified by next-generation sequencing in a Chinese family.

HLA-B*15:01:75 differs from HLA-B*15:01:01:01 by a single synonymous change in exon 2.

A REVIEW ON COMPUTATIONAL APPROACHES FOR IDENTIFYING NON-SYNONYMOUS MUTATION AND DNA METHYLATION IN CANCER

Cancer is a complex disease where, genetic and epigenetic changes plays a major role. Non- synonymous mutations and DNA methylation are important factors in cancer starts and development. Non- synonymous mutations are genetic changes that cause amino acid substitutions within proteins, which frequently result in defective or carcinogenic products. DNA methylation, on the other hand, is the addition of a methyl group to cytosine residues, which affects gene expression and contributes to cancer. Understanding the underlying mechanisms and designing targeted therapeutics require identifying these molecular events in cancer genomes. The traditional methods of identifying the mutations and DNA methylation has made a great challenge in the research fields. Computational approaches have shown to be important in this attempt. They cover a wide range of approaches, from mutation variant calling procedures to methylation pattern analysis tools. These methods will make use of high-throughput sequencing data to detect and characterize non- synonymous mutations and DNA methylation changes on a genome-wide scale. In this review, we will discuss about the multi omics dataset retrievals, tools used for the identification of mutated non- synonymous mutation and the algorithms and techniques used for DNA Methylation for identifying mutated regions. The critical values for the specific results also been noted in this study. Keywords: DNA Methylation, Non-synonymous genomic mutation, cancer, bioinformatics tools, algorithms

Suppression of the dwarf phenotype of an Arabidopsis mutant defective in thermospermine biosynthesis by a synonymous codon change in the SAC51 uORF.

Thermospermine plays a critical role in negatively regulating xylem development in angiosperms. A mutant of Arabidopsis thaliana that is defective in thermospermine biosynthesis, acaulis5 (acl5), exhibits a dwarf phenotype with excessive xylem formation. Mechanistically thermospermine acts in attenuating the inhibitory effect of an evolutionarily conserved upstream open reading frame (uORF) on the main ORF of SAC51, which encodes a basic helix-loop-helix protein involved in xylem repression. Here, we revealed that a semidominant suppressor of acl5, sac503, which partially restores the acl5 phenotype, has a point mutation in the conserved uORF of SAC51 with no amino acid substitution in the deduced peptide sequence. In transgenic lines carrying the β-glucuronidase (GUS) reporter gene fused with the SAC51 5' region containing the uORF, the mutant construct was shown to confer higher GUS activity than does the wild-type SAC51 construct. We confirmed that sac503 mRNA was more stable than SAC51 mRNA in acl5. These results suggest that the single-base change in sac503 positively affects the translation of its main ORF instead of thermospermine. We further found that the uORF-GUS fusion protein could be synthesized in planta from the wild-type and sac503 translational fusion constructs.

Gene recoding by synonymous mutations creates promiscuous intragenic transcription initiation in mycobacteria.

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis, one of the deadliest infectious diseases worldwide. Previous studies have established that synonymous recoding to introduce rare codon pairings can attenuate viral pathogens. We hypothesized that non-optimal codon pairing could be an effective strategy for attenuating gene expression to create a live vaccine for Mtb. We instead discovered that these synonymous changes enabled the transcription of functional mRNA that initiated in the middle of the open reading frame and from which many smaller protein products were expressed. To our knowledge, this is one of the first reports that synonymous recoding of a gene in any organism can create or induce intragenic transcription start sites.

Analysis of SARS-CoV-2 Population Genetics from Samples Associated with Huanan Market and Early Cases Identifies Substitutions Associated with Future Variants of Concern.

SARS-CoV-2 began spreading through human-to-human transmission first within China and then worldwide, with increasing sequence diversity associated with time and the further spread of the virus. The spillover events in the Huanan market were associated with two lineages of SARS-CoV-2 (lineages A and B). Infecting virus populations and those in infected individuals consist of a dominant genomic sequence and minor genomic variants; these latter populations can indicate sites on the genome that may be subject to mutational changes-either neutral or advantageous sites and those that act as a reservoir for future dominant variants-when placed under selection pressure. The earliest deposited sequences with human infections associated with the Huanan market shared very close homology with each other and were all lineage B. However, there were minor genomic variants present in each sample that encompassed synonymous and non-synonymous changes. Fusion sequences characteristic of defective RNA were identified that could potentially link transmission chains between individuals. Although all the individuals appeared to have lineage B as the dominant sequence, nucleotides associated with lineage A could be found at very low frequencies. Several substitutions (but not deletions) associated with much later variants of concern (VoCs) were already present as minor genomic variants. This suggests that low-frequency substitutions at the start of a pandemic could be a reservoir of future dominant variants and/or provide information on potential sites within the genome associated with future plasticity.

Detection of two SNPs of the LIPE gene in Holstein-Friesian cows with divergent milk production.

The LIPE gene (lipase E, hormone-sensitive type), also known as hormone-sensitive lipase, acts as a primary regulator of lipid metabolism during lactation in cows. We studied a total of two hundred Holstein-Friesian cows and performed sequencing analysis that revealed two synonymous nucleotide changes within the LIPE gene: a transition change, c.276 T > C in exon 2 (g.50631651 T > C; position 351 of GenBank: ON638900) and a transversion change, c.219C > A in exon 6 (g.50635369C > A; position 1070 of GenBank: ON638901). The observed genotypes were TC and CC for the c.276 T > C SNP and CC and CA for the c.219C > A SNP. Notably, the heterozygous TC genotype of the T351C SNP exhibited a significant association with high milk yield. Furthermore, the T351C SNP displayed significant associations with various milk parameters, including temperature, freezing point, density and the percentages of fat, protein, lactose, solids and solids-not-fat, with the homozygous CC genotype showing higher values. The c.219C > A SNP also demonstrated a significant association with milk composition, with heterozygous genotypes (CA) exhibiting higher percentages of fat, protein, and lactose compared to homozygous genotypes (CC). This effect was consistent among both high and low milk producers for fat and lactose percentages, while high milk producers exhibited a higher protein percentage than low milk producers. These findings highlight the importance of considering the detected SNPs in marker-assisted selection and breeding programs for the identification of high milk-producing Holstein-Friesian cows and potentially other breeds. Moreover, this study strongly supports the fundamental role of the LIPE gene in milk production and composition in lactating animals.

Predicting and Improving Insertions by Prime Editing

Predicting and Improving Insertions by Prime Editing

Evolutionary investigations of the biosynthetic diversity in the skin microbiome using lsaBGC.

Bacterial secondary metabolites, synthesized by enzymes encoded in biosynthetic gene clusters (BGCs), can underlie microbiome homeostasis and serve as commercialized products, which have historically been mined from a select group of taxa. While evolutionary approaches have proven beneficial for prioritizing BGCs for experimental characterization efforts to uncover new natural products, dedicated bioinformatics tools designed for comparative and evolutionary analysis of BGCs within focal taxa are limited. We thus developedlineagespecificanalysis of BGCs (lsaBGC;https://github.com/Kalan-Lab/lsaBGC) to aid exploration of microdiversity and evolutionary trends across homologous groupings of BGCs, gene cluster families (GCFs), in any bacterial taxa of interest.lsaBGC enables rapid and direct identification of GCFs in genomes, calculates evolutionary statistics and conservation for BGC genes, and builds a framework to allow for base resolution mining of novel variants through metagenomic exploration. Through application of the suite to four genera commonly found in skin microbiomes, we uncover new insights into the evolution and diversity of their BGCs. We show that the BGC of the virulence-associated carotenoid staphyloxanthin inStaphylococcus aureusis ubiquitous across the genusStaphylococcus. While one GCF encoding the biosynthesis of staphyloxanthin showcases evidence for plasmid-mediated horizontal gene transfer (HGT) between species, another GCF appears to be transmitted vertically amongst a sub-clade of skin-associatedStaphylococcus. Further, the latter GCF, which is well conserved inS. aureus, has been lost in mostStaphylococcus epidermidis, which is the most commonStaphylococcusspecies on human skin and is also regarded as a commensal. We also identify thousands of novel single-nucleotide variants (SNVs) within BGCs from theCorynebacterium tuberculostearicumsp. complex, a narrow, multi-species clade that features the most prevalentCorynebacteriumin healthy skin microbiomes. Although novel SNVs were approximately 10 times as likely to correspond to synonymous changes when located in the top five percentile of conserved sites,lsaBGC identified SNVs that defied this trend and are predicted to underlie amino acid changes within functionally key enzymatic domains. Ultimately, beyond supporting evolutionary investigations of BGCs,lsaBGC also provides important functionalities to aid efforts for the discovery or directed modification of natural products.

Mitochondrial genome of Artemisia argyi L. suggested conserved mitochondrial protein-coding genes among genera Artemisia, Tanacetum and Chrysanthemum

BackgroundArtemisia argyi L., also known as mugwort, is a perennial herb whose leaves are commonly used as a source of traditional medicines. However, the evolution and structure of the mitochondrial genome (mitogenome) in A. argyi remain unclear. In this study, the mitogenome of A. argyi was assembled and characterized for the first time. ResultsThe mitogenome of A. argyi was a circular molecule of 229,354 bp. It encodes 56 genes, including 33 protein-coding genes (PCGs), 20 tRNA genes, and three rRNA genes, and three pseudogenes. Five trans-spliced introns were observed in three PCGs namely, nad1, nad2 and nad5. Repeat analysis identified 65 SSRs, 14 tandem repeats, and 167 dispersed repeats. The A. argyi mitogenome contains 12 plastid transfer sequences from 79 bp to 2552 bp. Five conserved MTPTs were identified in all 18 Asteraceae species. Comparison of mitogenome between A. argyi and one Artemisia specie and two Chrysanthemum species showed 14 conserved gene clusters. Phylogenetic analysis with organelle genomes of A. argyi and 18 other Anthemideae plants showed inconsistent phylogenetic trees, which implied that the evolutionary rates of PCGs and rrna genes derived from mitochondrion and plastid were incongruent. The Ka/Ks ratio of the 27 shared protein-coding genes in the 18 Anthemideae species are all less than 1 indicating that these genes were under the effect of purifying selection. Lastly, a total of 568 RNA editing sites in PCGs were further identified. The average editing frequency of non-synonymous changes was significantly higher than that of synonymous changes (one-sample Student’s t-test, p-values ≤ 0.05) in three tissues (root, leaf and stem). ConclusionsIn this study, the gene content, genome size, genome comparison, mitochondrial plastid sequences, dN/dS analysis of mitochondrial protein-coding genes, and RNA-editing events in A. argyi mitogenome were determined, providing insights into the phylogenetic relationships of Asteraceae plant.