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BACKGROUND: The high-altitude-adapted frog Rana kukunoris, occurring on the Tibetan plateau, is an excellent model to study life history evolution and adaptation to harsh high-altitude environments. However, genomic resources for this species are still underdeveloped constraining attempts to investigate the underpinnings of adaptation. RESULTS: The R. kukunoris genome was assembled to a size of 4.83 Gb and the contig N50 was 1.80 Mb. The 6555 contigs were clustered and ordered into 12 pseudo-chromosomes covering ~ 93.07% of the assembled genome. In total, 32,304 genes were functionally annotated. Synteny analysis between the genomes of R. kukunoris and a low latitude species Rana temporaria showed a high degree of chromosome level synteny with one fusion event between chr11 and chr13 forming pseudo-chromosome 11 in R. kukunoris. Characterization of features of the R. kukunoris genome identified that 61.5% consisted of transposable elements and expansions of gene families related to cell nucleus structure and taste sense were identified. Ninety-five single-copy orthologous genes were identified as being under positive selection and had functions associated with the positive regulation of proteins in the catabolic process and negative regulation of developmental growth. These gene family expansions and positively selected genes indicate regions for further interrogation to understand adaptation to high altitude. CONCLUSIONS: Here, we reported a high-quality chromosome-level genome assembly of a high-altitude amphibian species using a combination of Illumina, PacBio and Hi-C sequencing technologies. This genome assembly provides a valuable resource for subsequent research on R. kukunoris genomics and amphibian genome evolution in general.
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The underground sedimentary bauxite ore body in Shanxi province has a shallow burial depth; the valley terrain caused stress concentration on a pillar which affected the pillar's safety and goaf stability. This paper proposed a pillar safety coefficient calculation method affected by the goaf structural parameters and the valley terrain, which was based on a pillar mechanics analysis under the valley terrain. The results show that the overlying valley terrain will cause stress concentration on the pillar, reducing the adequate bearing capacity and the pillar stability. The increase of the goaf span b and the height of the pillar h is extensively detrimental to pillar stability. Meanwhile, increasing the pillar burial depth would cause the pillar to weaken, but can effectively decrease the influence of the valley terrain. Furthermore, when the angle between the goaf strike and the valley strike ß < 50°, ß has a more significant impact on the stress concentration and safety coefficient. The stability of an underground sedimentary bauxite pillar is calculated by the method, the result complied with the actual situation.
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Classification of the genus Rhacophorus has been problematic. In particular there has been considerable controversy surrounding the phylogenetic relationships among Rhacophorus rhodopus, R. bipunctatus, and R. reinwardtii. To examine the relationship among these Rhacophorus species, we assembled the complete mitochondrial genome sequence of R. rhodopus. The R. rhodopus genome is 15,789 bp in length with 12 protein-coding genes (PCGs) (losing ND5), two ribosomal genes, 22 transfer RNA genes, and a control region (D-loop). Base composition of the overall sequence was 60.86% for A + T content and 39.14% for C + G content. Most of the PCGs used ATG as a start codon, except for the COX I gene, which used the ATA start codon. COX I and ND6 used AGG and ATP8 stop codons respectively, while ND3 and ND4L used the TAA stop codon. For the remaining seven genes, the stop codons was incomplete. In addition, both 5' and 3' of the control areas had distinct repeating regions. Based on three datasets and two methods (Bayesian inference (BI) and maximum likelihood (ML)), we reconstructed three phylogenetic trees to explore the taxonomic status of the species and the phylogenetic relationship among R. rhodopus, R. bipunctatus and R. reinwardtii. Our results indicated that these three species are non-monophyletic; thus, the phylogenetic relationship among them is complex and difficult to determine. Further, R. rhodopus is divided into three lineages from different parts of China. The two Rhacophorus samples showed very close phylogenetic relationship with R. rhodopus. Our results add to the mitochondrial genome database of amphibians and will help to disentangle the phylogenetic relationships within the Rhacophoridae.
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Genoma Mitocondrial , Animais , Anuros/genética , Teorema de Bayes , Códon de Iniciação/genética , Códon de Terminação , Genoma Mitocondrial/genética , Fases de Leitura Aberta/genética , Filogenia , RNA de Transferência/genética , Análise de Sequência de DNARESUMO
CRISPR-based genome-editing tools have emerged as an efficient tool for functional genomics studies. Online tools and databases have been developed to facilitate the design and selection of CRISPR single guide RNA (sgRNA) for gene modifications. However, to the best of our knowledge, none of these tools or database are designated to cell surface proteins. In a previous study, we described the development and application of surfaceome CRISPR libraries targeting to cell surface proteins on human cells. Here, we present the design and construction of an online tool and database (https://crispr-surfaceome.siais.shanghaitech.edu.cn/home), named CRISPR-Surfaceome, for the design of highly efficient sgRNA targeting to the surface proteins on human cells. To show case and validate the efficiencies of sgRNAs designed by this online tool, we chose ICAM-1 gene for knockout studies and found that all the 10 designed ICAM-1 sgRNAs could efficiently generate knockout cells, with more than 80% gene disruption rates. These ICAM-1 knockout cells were found to be resistant to the infection of rhinovirus (RV), which utilizes ICAM-1 as the receptor. Therefore, CRISPR-Surfaceome can serve the research community for the functional genomics studies on cell surface proteins, such as identification of pathogen receptors and discovery of drug targets.
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BACKGROUND: Previous studies have indicated that the programmed death molecule 1 (PD-1) signaling pathway may play a key role in rheumatoid arthritis (RA). However, the pathogenesis of rheumatoid arthritis-related interstitial lung disease (RA-ILD) is not clear. We examined the serum levels of soluble PD-1 in patients with RA and its relationship with RA-ILD. METHODS: Blood samples were obtained from 87 patients with RA (58 with ILD and 29 without ILD) and 45 healthy controls. Serum sPD-1 was measured by Enzyme-Linked Immunosorbent Assay. The pulmonary interstitial disease score was completed by a pulmonary physician and a radiologist through chest high-resolution computed tomography. Patients with RA-ILD were tested for lung function [e.g., forced vital capacity (FVC%), diffusing capacity of lungs for carbon monoxide (DLCO%)]. Associations between ILD and various markers, including sPD-1 and confounding factors, were investigated by logistic regression analysis. Diagnostic values of sPD-1 for the presence of ILD were investigated using receiver operating characteristic curve analysis. RESULTS: Serum sPD-1 levels were higher in RA patients with ILD than in RA patients without ILD and healthy controls (185.1 ± 109.0 pg/ml vs. 119.1 ± 77.5 pg/ml vs. 52.1 ± 21.7 pg/ml, P < 0.05). Serum sPD-1 levels were positively correlated with RF titer (P = 0.02, r = 0.249), anti-cyclic citrullinated peptide antibody status (P = 0.02, r = 0.243), and serum IgG levels (P < 0.001, r = 0.368), negatively associated with FVC% (P = 0.02, r = - 0.344), forced expiratory volume (FEV1%) (P = 0.01, r = - 0.354), total lung capacity (TLC%) (P = 0.046, r = - 0.302), and was independently associated with the presence of ILD in RA patients by multivariate logistic regression analysis. The sensitivity and specificity of sPD-1 levels for the detection of ILD in RA patients were 58.6% and 75.9%, respectively. The area under the curve was 0.689. CONCLUSION: Serum sPD-1 levels were increased in RA patients with ILD. Increased sPD-1 may be a valuable biomarker to predict the presence of ILD in patients with RA.
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Artrite Reumatoide/diagnóstico , Biomarcadores/sangue , Doenças Pulmonares Intersticiais/diagnóstico , Pulmão/patologia , Receptor de Morte Celular Programada 1/sangue , Idoso , Artrite Reumatoide/imunologia , Feminino , Humanos , Doenças Pulmonares Intersticiais/imunologia , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Prognóstico , Testes de Função Respiratória , Regulação para CimaAssuntos
Anticorpos Neutralizantes/imunologia , Vacinas contra COVID-19/imunologia , COVID-19/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Antivirais/imunologia , COVID-19/prevenção & controle , COVID-19/virologia , Humanos , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Vacinas de Produtos Inativados/imunologiaRESUMO
Nonalcoholic steatohepatitis (NASH) is an aggressive liver disease threatening human health, yet no medicine is developed to treat this disease. In this study, we first discovered that Leptin mutant rats (LepΔI14/ΔI14) exhibit characteristic NASH phenotypes including steatosis, lymphocyte infiltration, and ballooning after postnatal week 16. We then examined NASH progression by performing an integrated analysis of hepatic transcriptome in Leptin-deficient rats from postnatal 4 to 48 weeks. Initially, simple steatosis in LepΔI14/ΔI14 rats were observed with increased expression of the genes encoding for rate-limiting enzymes in lipid metabolism such as acetyl-CoA carboxylase and fatty acid synthase. When NASH phenotypes became well developed at postnatal week 16, we found gene expression changes in insulin resistance, inflammation, reactive oxygen species and endoplasmic reticulum stress. As NASH phenotypes further progressed with age, we observed elevated expression of cytokines and chemokines including C-C motif chemokine ligand 2, tumor necrosis factor É, interleukin-6, and interleukin-1ß together with activation of the c-Jun N-terminal kinase and nuclear factor-κB pathways. Histologically, livers in LepΔI14/ΔI14 rats exhibited increased cell infiltration of MPO+ neutrophils, CD8+ T cells, CD68+ hepatic macrophages, and CCR2+ inflammatory monocyte-derived macrophages associated with macrophage polarization from M2 to M1. Subsequent cross-species comparison of transcriptomes in human, rat, and mouse NASH models indicated that Leptin-deficient rats bear more similarities to human NASH patients than previously established mouse NASH models. Taken together, our study suggests that LepΔI14/ΔI14 rats are a valuable pre-clinical rodent model to evaluate NASH drug safety and efficacy.
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Progressão da Doença , Perfilação da Expressão Gênica , Leptina/deficiência , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/patologia , Animais , Regulação da Expressão Gênica , Inflamação/patologia , Leptina/metabolismo , Fígado/metabolismo , Fígado/patologia , Camundongos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Fenótipo , Ratos , Fatores de Tempo , Transcriptoma/genéticaRESUMO
The Omei wood frog (Rana omeimontis), endemic to central China, belongs to the family Ranidae. In this study, we achieved detail knowledge about the mitogenome of the species. The length of the genome is 20,120 bp, including 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a noncoding control region. Similar to other amphibians, we found that only nine genes (ND6 and eight tRNA genes) are encoded on the light strand (L) and other genes on the heavy strand (H). Totally, The base composition of the mitochondrial genome included 27.29% A, 28.85% T, 28.87% C, and 15.00% G, respectively. The control regions among the Rana species were found to exhibit rich genetic variability and A + T content. R. omeimontis was clustered together with R. chaochiaoensis in phylogenetic tree. Compared to R. amurensis and R. kunyuensi, it was more closely related to R. chaochiaoensis, and a new way of gene rearrangement (ND6-trnE-Cytb-D-loop-trnL2 (CUN)-ND5-D-loop) was also found in the mitogenome of R. amurensis and R. kunyuensi. Our results about the mitochondrial genome of R. omeimontis will contribute to the future studies on phylogenetic relationship and the taxonomic status of Rana and related Ranidae species.
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The Günther's frog (Hylarana guentheri) belongs to a member of the family Ranidae. We provide a complete mitogenome of H. guentheri and examine its phylogenetic position with other related species. Its mitogenome is a closed circular molecule 18,698 bp in length including 13 protein coding genes, 22 tRNA coding genes, two rRNA-coding genes, and a control region (CR) that are conserved in most Ranidae mitogenomes. The overall base composition of the H. guentheri mitogenome is 29.27% A, 30.45% T, 26.14% C, and 14.15% G, which is typical for Amphibious animals' mitochondrial genomes. The alignment of the Ranidae species control regions showed high levels of genetic variation and abundant AT content. Seven tandem repeats were found in the control region. Phylogenetic analysis with Bayesian inference and maximum likelihood based on 13 protein-coding genes indicated that H. guentheri is more closely related to Nidirana okinavana than to Babina subaspera and B. holsti. The complete mitogenome of H. guentheri provides a potentially useful resource for further exploration of the taxonomic status and phylogenetic relationships of Hylarana and related species.
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In this study, the complete mitochondrial genome of big-eyed mountain keelback Pseudoxenodon macrops was sequenced adopting Illumina high-throughput sequencing technology. The complete mitogenome of the species was 19,444 bp in length, including 13 protein-coding genes, 22 transfer RNA (tRNA) genes, two ribosomal RNA genes, and two non-coding control regions (CR). The overall base composition of mitogenome was 32.0% A, 25.5% T, 28.2% C, and 14.3% G. Most mitochondrial genes are encoded on the heavy strand, only ND6 and eight tRNA genes are on the light strand. We expect that the presented mitogenome can provide important data for future studies on phylogenetic relationship and population genetics of this species.
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In this study, the complete mitochondrial genome of Duttaphrynus himalayanus was sequenced adopting Illumina high-throughput sequencing method. The complete mitogenome of the species was 17,172 bp in length, including 13 protein-coding genes, 22 transfer RNA (tRNA) genes, two ribosomal RNA genes, and a non-coding control region (CR). The overall base composition of mitogenome was 29.7% A, 29.6% T, 26.0% C, and 14.7% G. Most mitochondrial genes are encoded on the heavy strand, only ND6 and eight tRNA genes on the light strand. The complete mitogenome of D. himalayanus can provide an important data for future studies on phylogenetic relationship and population genetics of this species.
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In this study, we obtained the complete mitochondrial genome sequence of Nanorana chayuensis. The mitogenome length is 17,882 bp, including 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNA), 2 ribosomal RNA genes (rRNA), and 1 non-coding control region (CR). Present data will contribute to further analysis of phylogenetic relationship and population genetics of this species.
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Large-scale estimation of forest biomass has received much attention. Constructing a stand-level biomass model is a method for estimating tree layer biomass. In this study, we constructed stand biomass models of Korean pine plantations based on aggregation method 1, aggregation method 2, adjustment method, and disaggregation method. The prediction precision of four additive methods was compared and analyzed to provide theoretical basis for biomass prediction of Korean pine plantations in Heilongjiang Province. Weighted functions were used to eliminate the heteroscedasticity of each model, with the leave-one-out cross validation (LOOCV) as the validation method. The results showed that the overall prediction ability of the adjustment method was slightly better than other methods. The specific prediction precision was ranked as adjustment method > aggregation method 1 > aggregation method 2 > disaggregation method. The prediction precision of four additive methods was not consistent when considering their prediction ability of different stand basal areas. When the stand basal area of Korean pine plantations was distributed in the interval of 0-10 or 50-60 m2·hm-2, the parameter estimation values of disaggregation method performed better. When the stand basal area was distributed in other intervals, the parameter estimation values of adjustment method was better.
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Pinus , Biomassa , China , Florestas , ÁrvoresRESUMO
To compare clinical characteristics and identify long-term outcomes of Chinese patients with systemic sclerosis (SSc) with and without muscle involvement.We retrospectively investigated the medical records, laboratory results, and computed tomography images of 204 consecutive SSc patients. Kaplan-Meier analysis was performed to determine survival rates. Patients were allocated into groups with and without myopathy.The prevalence of myopathy was 21.6%. The myopathy group was more likely to develop diffuse cutaneous involvement (90.9% vs 56%, Pâ=â.006), interstitial lung disease (90% vs 56%, Pâ<â.001), digestive system involvement (56.7% vs 29.3%, Pâ=â.001), pulmonary hypertension (29.5% vs 10.5%, Pâ=â.004), and pericardial effusion (25% vs. 10%, Pâ=â.019). Patients with myopathy had lower single-breath diffusing capacity of the lung for carbon oxide (46.5â±â11.1 vs 57.1â±â13.4, Pâ<â.001).Further, the myopathy group has similar results in interstitial lung disease associated higher resolution computed tomography score (186.8â±â64.5 vs 152.3â±â45.5, Pâ=â.037), Valentini score for disease activity (3.4â±â0.9 vs 2.0â±â0.9, Pâ<â.001) and modified Rodnan total skin score (19.4â±â6.1 vs 15.1â±â7.7, Pâ=â.002), compared with non-myopathy group. Kaplan-Meier survival analysis revealed decreased overall survival rate of the myopathy group (Pâ=â.028).SSc Patients with myopathy had more severe clinical manifestations and higher disease activity compared with those without, which affected survival rates and indicated worse prognosis.
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Doenças Musculares/mortalidade , Escleroderma Sistêmico/mortalidade , Adulto , Idoso , China/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Doenças Musculares/etiologia , Estudos Retrospectivos , Escleroderma Sistêmico/complicaçõesRESUMO
Magnolia officinalis bark is a traditional Chinese medicine for gastrointestinal tract disorders. In this study, we explored the effects of M. officinalis extraction on intestinal flora to reveal its mechanism. Thirty SPF mice were divided into five groups: C (control), M (M. officinalis), A (antibiotics: cefradine and gentamicin sulfate), A&M (antibiotics + M. officinalis) and A&N (antibiotics + natural recovery). Faecal samples of all groups were collected and the taxonomic composition and diversity of bacteria was characterized using the 16S rRNA gene (16S). Alpha diversity showed gut bacteria diversity significantly decreased in the A group of mice but increased markedly after administration of M. officinalis extract. Beta diversity indicated that C, M and A&M shared similar bacterial community structure while A and A&N exhibited a different bacterial community. Furthermore, RDA combined with spearman correlation heatmap suggested the five physiological indicators (weight, fur, activity and feces) were highly correlated with bacterial community structure and diversity. Finally, functional categorization of the assigned OTUs was performed using the PICRUSt tool. The changes in PICRUSt inferred that function profile and metabolic pathways were observed in A and A&M, therefore the M. officinalis extract improved the intestinal flora of A&M and normalized its metabolic pathways gradually, improving mouse weight, fur quality, activity and feces qualities.
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Microbioma Gastrointestinal , Magnolia , Animais , Antibacterianos , Disbiose , Camundongos , RNA Ribossômico 16S/genéticaRESUMO
The Hongyuan breed Yak (Bos grunniens) belongs to a member of t the subfamily Bovinae. We provide a complete mitogenome of B. grunniens and analyze its phylogenetic relationship with other related species. Its mitogenome is a circular molecule with 16,322 bp in size, including 13 protein coding genes, 22 tRNA genes, 2 rRNA genes, and a non-coding control region (D-loop, CR) that are conserved in most Bovidae mitogenomes. The total base composition of the B. grunniens mitogenome is 33.67% A, 27.29% T, 25.84% C, and 13.20% G. The gene composition, structure and the arrangement for B. grunniens are similar to those of most other Bovidae species. Phylogenetic analysis of mitochondrial genomes of 30 close species with Bayesian inference and maximum likelihood based on 13 protein-coding genes indicated that B. grunniens breed Hongyuan is more closely related to B. grunniens breed Qinghai Plateau than to B. grunniens breed Xuedong and B. grunniens breed Maiwa. The complete mitogenome of B. grunniens breed Hongyuan provides a potentially useful resource for further exploration of the taxonomic status and phylogenetic relationships of Bovinae and related species.
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This is the first study to describe the mitochondrial genome of the Himalayan Griffon, Gyps himalayensis, which is an Old World vulture belonging to the family Accipitridae and occurring along the Himalayas and the adjoining Tibetan Plateau. Its mitogenome is a closed circular molecule 17,381 bp in size containing 13 protein-coding genes, 22 tRNA coding genes, two rRNA-coding genes, a control region (CR), and an extra pseudo-control region (CCR) that are conserved in most Accipitridae mitogenomes. The overall base composition of the G. himalayensis mitogenome is 24.55% A, 29.49% T, 31.59% C, and 14.37% G, which is typical for bird mitochondrial genomes. The alignment of the Accipitridae species control regions showed high levels of genetic variation and abundant AT content. At the 5' end of the domain I region, a long continuous poly-C sequence was found. Two tandem repeats were found in the pseudo-control regions. Phylogenetic analysis with Bayesian inference and maximum likelihood based on 13 protein-coding genes indicated that the relationships at the family level were (Falconidae + (Cathartidae + (Sagittariidae + (Accipitridae + Pandionidae))). In the Accipitridae clade, G. himalayensis is more closely related to Aegypius monachus than to Spilornis cheela. The complete mitogenome of G. himalayensis provides a potentially useful resource for further exploration of the taxonomic status and phylogenetic history of Gyps species.
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OBJECTIVE: To determine the serum levels of Dickkopf-1 (DKK-1) and sclerostin, as well as their correlations with the structural damage assessed by modified stoke ankylosing spondylitis spine score (mSASSS) and the disease activity evaluated by ankylosing spondylitis disease activity score (ASDAS) in patients with ankylosing spondylitis (AS). METHODS: Eighty-eight AS patients, 26 rheumatoid arthritis (RA) patients, and 26 age- and gender-matched healthy controls (HC) were collected from rheumatic clinic of the Second Affiliated Hospital of Zhejiang University, School of Medicine, between March 2015 and July 2015. Demographic data, parameters of ASDAS, and image evaluations of spine (i.e., mSASSS) were collected. The serum levels of DKK-1 and sclerostin were measured using commercially available ELISA kits. RESULTS: Both DKK-1 and sclerostin were significantly higher in the AS patients than in the controls (1855 ± 84.58 vs. 1406 ± 99.76 pg/ml and 106 ± 6.75 vs. 62.78 ± 6.39 pmol/l, respectively, P < 0.05). The correlation analysis suggested a negative correlation between serum sclerostin and mSASSS (P = 0.019, r2 = 0.062). DKK-1 had a trend of positive correlation with mSASSS, but was not statistically significant (P > 0.05). There was no association between the serum levels of DKK-1 or sclerostin and disease activity assessed by ASDAS (P > 0.05). DKK-1 and sclerostin had a negative correlation (P = 0.013, r2 = 0.07). CONCLUSION: In the present study, the expressions of serum DKK-1 and sclerostin were independent of disease activity. Sclerostin was negatively correlated with the mSASSS, which suggests that sclerostin may be a potential marker indicating the spine ossification process in AS. The specific mechanism remains to be investigated.
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Proteínas Morfogenéticas Ósseas/sangue , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Coluna Vertebral/diagnóstico por imagem , Espondilite Anquilosante/sangue , Proteínas Adaptadoras de Transdução de Sinal , Adulto , Artrite Reumatoide/sangue , Artrite Reumatoide/diagnóstico por imagem , Feminino , Marcadores Genéticos , Humanos , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Espondilite Anquilosante/diagnóstico por imagem , Adulto JovemRESUMO
Insulin resistance is a pathophysiological hallmark of type 2 diabetes and nonalcoholic fatty liver disease. Under the condition of fat accumulation in the liver, suppression of hepatic glucose production by insulin is diminished. In order to gain deeper understanding of dysregulation of glucose production in metabolic diseases, in the present study, we performed an unbiased phenotypic screening in primary human hepatocytes to discover novel mechanisms that regulate gluconeogenesis in the presence of insulin. To optimize phenotypic screening process, we used a chemical genetic screening approach by building a small-molecule library with prior knowledge of activity-based protein profiling. The "positive hits" result from the screen will be small molecules with known protein targets. This makes downstream deconvolution process (i.e., determining the relevant biological targets) less time-consuming. To unbiasedly decipher the molecular targets, we developed a novel statistical method and discovered a set of genes, including DDR3 and CACNA1E that suppressed gluconeogenesis in human hepatocytes. Further investigation, including transcriptional profiling and gene network analysis, was performed to understand the molecular functions of DRD3 and CACNA1E in human hepatocytes.
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Members of the Nanorana genus (family Dicroglossidae) are often referred to as excellent model species with which to study amphibian adaptations to extreme environments and also as excellent keystone taxa for providing insights into the evolution of the Dicroglossidae. However, a complete mitochondrial genome is currently only available for Nanorana pleskei. Thus, we analyzed the complete mitochondrial genomes of Nanorana parkeri and Nanorana ventripunctata to investigate their evolutionary relationships within Nanorana and their phylogenetic position in the family Dicroglossidae. Our results showed that the genomes of N. parkeri (17,837 bp) and N. ventripunctata (18,373 bp) encode 13 protein-coding genes (PCGs), two ribosomal RNA genes, 23 transfer RNA (tRNA) genes, and a noncoding control region. Overall sequences and genome structure of the two species showed high degree of similarity with N. pleskei, although the motif structures and repeat sequences of the putative control region showed clear differences among these three Nanorana species. In addition, a tandem repeat of the tRNA-Met gene was found located between the tRNA-Gln and ND2 genes. On both the 5' and 3'-sides, the control region possessed distinct repeat regions; however, the CSB-2 motif was not found in N. pleskei. Based on the nucleotide sequences of 13 PCGs, our phylogenetic analyses, using Bayesian inference and maximum-likelihood methods, illustrate the taxonomic status of Nanorana with robust support showing that N. ventripunctata and N. pleskei are more closely related than they are to N. parkeri. In conclusion, our analyses provide a more robust and reliable perspective on the evolutionary history of Dicroglossidae than earlier analyses, which used only a single species (N. pleskei).