Two mitochondrial genes under episodic positive selection in subterranean octodontoid rodents

Tuco-tucos (Ctenomys) and related coruros (Spalacopus) are South American subterranean rodents. An energetically demanding lifestyle within the hypoxic, underground atmosphere may change the selective regime on oxidative phosphorylation. We examined whether weak and/or episodic positive directional selection affected the evolution of two mitochondrial genes (COX2, CytB), in a background of purifying selection in these lineages. We estimated rates of synonymous (dS) and non-synonymous (dN) substitutions and found: 1) significantly higher dN/dS ratio in subterranean groups relative to non-subterranean related species, and 2) two codons in each gene under episodic selection: 94 and 277 of COX2 and 269 and 307 of CytB.     

The complete mitochondrial genome of the leafminer Liriomyza sativae (Diptera: Agromyzidae): great difference in the A+T-rich region compared to Liriomyza trifolii

The complete mitochondrial genome sequence of Liriomyza sativae Blanchard (15,551 bp) was determined and analyzed in this study. The circular genome contained 37 genes including 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and an A + T-rich region. The initiation codons of COI and ND1 were ‘ATCA’ and ‘GTG’, respectively. ND2 gene used the truncated termination codon ‘T’. All the tRNA genes had the typical cloverleaf secondary structures except for tRNA^Ser(AGN) gene, which was found with the absence of a DHU arm. In addition, a tRNA-like secondary structure (tRNA^Met) was found in the A + T-rich region. The great difference was that the length of L. sativae A + T-rich region was 597 bp shorter than that of Liriomyza trifolii (Burgess). Meanwhile, some minor differences such as ‘TATA’ block were also observed in L. sativae in contrast to ‘TACA’ block in L. trifolii. There were also some essential structure elements such as ‘TATA’ block, ‘G(A)_nT’ block, poly-T stretch and stem-and-loop structure in the A + T-rich region of L. sativae mitochondrial genome.     

Eight new mtDNA sequences of glass sponges reveal an extensive usage of + 1 frameshifting in mitochondrial translation

Three previously studied mitochondrial genomes of glass sponges (phylum Porifera, class Hexactinellida) contained single nucleotide insertions in protein coding genes inferred as sites of + 1 translational frameshifting. To investigate the distribution and evolution of these sites and to help elucidate the mechanism of frameshifting, we determined eight new complete or nearly complete mtDNA sequences from glass sponges and examined individual mitochondrial genes from three others. We found nine new instances of single nucleotide insertions in these sequences and analyzed them both comparatively and phylogenetically. The base insertions appear to have been gained and lost repeatedly in hexactinellid mt protein genes, suggesting no functional significance for the frameshifting sites. A high degree of sequence conservation, the presence of unusual tRNAs, and a distinct pattern of codon usage suggest the “out-of-frame pairing” model of translational frameshifting. Additionally, we provide evidence that relaxed selection pressure on glass sponge mtDNA – possibly a result of their low growth rates and deep-water lifestyle – has allowed frameshift insertions to be tolerated for hundreds of millions of years. Our study provides the first example of a phylogenetically diverse and extensive usage of translational frameshifting in animal mitochondrial coding sequences.     

基于ND4和ND5基因序列分析的鳅超科鱼类系统发育关系

ND4和ND5是线粒体基因组中编码NADH脱氢酶亚基4和亚基5的两个蛋白质编码基因.该研究以鳅超科鱼类为研究对象,新测定了10个物种的ND4和ND5基因全序列以及中间的3个tRNA基因共212 bp的序列,结合从GenBank 下载的15个物种的15条序列进行序列比较和系统发育关系分析.结果显示:鳅超科鱼类ND4基因全长1380～1387 bp,以ATG为起始密码子,终止密码子为不完全终止信号;ND5基因全长1821～1839bp,同样起始密码子为ATG,终止密码子为TAA或TAG;ND4和ND5基因之间插入了3个tRNA基因,分别编码携带组氨酸、丝氨酸、亮氨酸的tRNA.ND4和ND5基因(包含3个tRNA基因)中A、T、G、C的平均含量分别为30.4%、27.3%、14.2%、28.1%,A+T(57.7%)的含量高于G+C(42.3%)的含量.转换与颠换比(Ti/Tv)平均值为1.586.选取斑马鱼和鲤鱼作为外类群,采用最大简约法(MP)、最大似然法(ML)和贝叶斯推断法(BI)进行系统发育树的重建.三种方法的系统发育分析结果都显示:花鳅亚科、条鳅亚科、沙鳅亚科、平鳍鳅科及Vaillantellidae分别构成单系;它们的系统发育关系为:(Vaillantellidae+(沙鳅亚科+(花鳅亚科+(条鳅亚科+平鳍鳅科).这与线粒体全基因组和某些核基因(如RAG1基因)的研究结果类似,且支持率较高,表明ND4和ND5基因用于鳅超科鱼类的系统发育分析是可行的;但是该研究的结果有别于其他线粒体基因的分析结果,如基于cytb和D-loop基因进行的系统发育分析表明,条鳅亚科和花鳅亚科聚为姐妹群,再和平鳍鳅科聚在一起.这种差异可能是由于使用的基因长度差异造成的,长度越长,信息量越大,所反映的系统发育结果可能更加接近真实情况.     

Molecular evolution of two duplicated CDPK genes CPK7 and CPK12 in grass species: a case study in wheat (Triticum aestivum L.)

Gene duplication contributes to the expansion of gene families and subsequent functional diversification. Calcium-dependent protein kinases (CDPKs) are members of an important calcium sensor family involved in abiotic and biotic stress signaling in plants. We report here the molecular evolution and expression analysis of a pair of duplicated CDPK genes CPK7 and CPK12 that arose in the common ancestor of grass species. With higher nonsynonymous/synonymous ratios (dN/dS, or ω), CPK12 genes appear to diverge more rapidly than CPK7s, suggesting relaxed selection constraints on CPK12s. Sliding window analysis revealed increased dN and ω values at N-terminal regions and the calcium-binding EF hand loops. Likelihood analyses using various models in PAML 4.0 showed purifying selection on both CPK7 and CPK12 lineages. In addition to the divergence in cis-element combinations on their promoters, functional divergence of CPK7 and CPK12 genes was also observed in wheat where TaCPK7 was found to respond to drought (PEG), salt (NaCl), cold, and hydrogen peroxide (H(2)O(2)) while TaCPK12 responded only to the treatment of ABA, a feature that may complement or expand TaCPK7-mediated stress signaling networks of wheat. The contrasting expression patterns of CPK7 and CPK12 genes under stress conditions were also observed in rice, suggesting conservative functional evolution of these genes. Since no positive selection was detected between the two lineages, the divergence of CPK7 and CPK12 genes should be ascribed to subfunctionalization, rather than neofunctionalization. Thus, our work demonstrates another case of evolutionary employment of duplicated genes via subfunctionalization for better adaptation.     

Phylogenetic utility of two existing and four novel nuclear gene loci in reconstructing Tree of Life of ray-finned fishes: The order Cypriniformes (Ostariophysi) as a case study

After the completion of several entire genome projects and a remarkable increase in public genetic databases in the recent years the results of post-genomic analyses can facilitate a better understanding of the genomic evolution underlying the diversity of organisms and the complexity of gene function. This influx of genomic information and resources is also beneficial to the discipline of systematic biology. In this paper, we describe a set of 6 previous and 22 new PCR/sequencing primers for RAG1, Rhodopsin and four novel nuclear markers from IRBP, EGR1, EGR2B and EGR3 that we developed through an approach making use of public genetic/genomic data mining for one of the ongoing tree of life projects aimed at understanding the evolutionary relationships of the planet's largest clade of freshwater fishes — the Cypriniformes. The primers and laboratory protocols presented here were successfully tested in 33 species comprising all cypriniform family and subfamily groups. Phylogenetic performance of each gene, as well as their implications in the investigation of the evolution of cypriniform fishes were assessed and discussed.     

Mitochondrial DNA as effective molecular markers for the genetic variation and phylogeny of the family Osteoglossidae

The present study examined the genetic variation of the family Osteoglossidae from different geographical locations based on the mitochondrial NADH dehydrogenase subunit 2 (ND2) and ATPase subunit 6 (ATPase6) genes; we then re-constructed the phylogenetic relationships using the two sequences in combination. The results showed that the partial sequences of mitochondrial ND2 and ATPase6 of the family Osteoglossidae were 813 bp and 669 bp, respectively. A total of 42 species-specific nucleotide positions of the family Osteoglossidae were found to be useful for molecular identification. The sequence variation showed greater differences (8.3% ~ 28.1% for the combined sequences, 8.3% ~ 26.7% for the ND2 gene, and 9.3% ~ 28.7% for the ATPase6 gene) among the different species of Osteoglossidae, and there was a significant association between the genetic difference and geographical location. Phylogenetic analyses using neighbor-joining, Bayesian inference, and maximum parsimony (MP) methods based on the combined sequences of the two genes were able to distinguish the different species and were in agreement with the existing taxonomy based on morphological characters and in association with the geographical distribution among seven species of the family Osteoglossidae.     

The complete mitochondrial genome of Macrobrachium nipponense

The complete mitochondrial (mt) genome sequence plays an important role in the accurate determination of phylogenetic relationships among metazoans. Herein, we determined the complete mt genome sequence, structure and organization of Macrobrachium nipponense (M. nipponense) (GenBank ID: NC_015073.1) and compared it to that of Macrobrachium lanchesteri (M. lanchesteri) and Macrobrachium rosenbergii (M. rosenbergii). The 15,806 base pair (bp) M. nipponense mt genome, which is comprised of 37 genes, including 13 protein-coding genes (PCGs), 22 transfer RNAs (tRNAs) and 2 ribosomal RNAs (rRNAs), is slightly larger than that of M. lanchesteri (15,694 bp, GenBank ID: NC_012217.1) and M. rosenbergii (15,772 bp, GenBank ID: NC_006880.1). The M. nipponense genome contains a high AT content (66.0%), which is a common feature among metazoan mt genomes. Compared with M. lanchesteri and M. rosenbergii, we found a peculiar non-coding region of 950 bp with a microsatellite-like (TA)₆ element and many hairpin structures. The 13 PCGs are comprised of a total of 3707 codons, excluding incomplete termination codons, and the most frequently used amino acid is Leu (16.0%). The predicted start codons in the M. nipponense mt genome include ATG, ATC and ATA. Seven PCGs use TAA as a stop codon, whereas two use TAG, three use T and only one uses TA. Twenty-three of the genes are encoded on the L strand, and ND1, ND4, ND5, ND4L, 12S rRNA, 16S rRNA, tRNA^His, tRNA^Pro, tRNA^Phe, tRNA^Val, tRNA^Gln, tRNA^Cys, tRNA^Tyr and a tRNA^Leu are encoded on the H strand. The two rRNAs of M. nipponense and M. rosenbergii are encoded on the H strand, whereas the M. lanchesteri rRNAs are encoded on the L stand.     

Characterization of the complete mitochondrial genome of Tryporyza incertulas, in comparison with seven other Pyraloidea moths

The complete 15,223-bp mitochondrial genome (mitogenome) of Tryporyza incertulas (Walker) (Lepidoptera: Pyraloidea: Crambidae) was determined, characterized and compared with seven other species of superfamily Pyraloidea. The order of 37 genes was typical of insect mitochondrial DNA sequences described to date. Compared with other moths of Pyraloidea, the A + T biased (77.0%) of T. incertulas was the lowest. Eleven protein-coding genes (PCGs) utilized the standard ATN, but cox1 used CGA and nad4 used AAT as the initiation codons. Ten protein-coding genes had the common stop codon TAA, except nad3 having TAG as the stop codon, and cox2, nad4 using T, TA as the incomplete stop codons, respectively. All of the tRNA genes had typical cloverleaf secondary structures except trnS1(AGN), in which the dihydrouridine (DHU) arm did not form a stable stem-loop structure. There was a spacer between trnQ and nad2, which was common in Lepidoptera moths. A 6-bp motif ‘ATACTA’ between trnS2(UCN) and nad1, a 7-bp motif “AGC(T)CTTA” between trnW and trnC and a 6-bp motif “ATGATA” of overlapping region between atp8 and atp6 were found in Pyraloidea moths. The A + T-rich region contained an ‘ATAGT(A)’-like motif followed by a poly-T stretch. In addition, two potential stem-loop structures, a duplicated 19-bp repeat element, and two microsatellites ‘(TA)₁₂’ and ‘(TA)₉’ were observed in the A + T-rich region of T. incertulas mitogenome. Finally, the phylogenetic relationships of Pyraloidea species were constructed based on amino acid sequences of 13 PCGs of mitogenomes using Bayesian inference (BI) and maximum likelihood (ML) methods. These molecular-based phylogenies supported the morphological classification on relationships within Pyraloidea species.     

The complete mitochondrial genome of the Chinese hook snout carp Opsariichthys bidens (Actinopterygii: Cypriniformes) and an alternative pattern of mitogenomic evolution in vertebrate

The complete mitochondrial genome sequence of the Chinese hook snout carp, Opsariichthys bidens, was newly determined using the long and accurate polymerase chain reaction method. The 16,611-nucleotide mitogenome contains 13 protein-coding genes, two rRNA genes (12S, 16S), 22 tRNA genes, and a noncoding control region. We use these data and homologous sequence data from multiple other ostariophysan fishes in a phylogenetic evaluation to test hypothesis pertaining to codon usage pattern of O. bidens mitochondrial protein genes as well as to re-examine the ostariophysan phylogeny. The mitochondrial genome of O. bidens reveals an alternative pattern of vertebrate mitochondrial evolution. For the mitochondrial protein genes of O. bidens, the most frequently used codon generally ends with either A or C, with C preferred over A for most fourfold degenerate codon families; the relative synonymous codon usage of G-ending codons is greatly elevated in all categories. The codon usage pattern of O. bidens mitochondrial protein genes is remarkably different from the general pattern found previously in the relatively closely related zebrafish and most other vertebrate mitochondria. Nucleotide bias at third codon positions is the main cause of codon bias in the mitochondrial protein genes of O. bidens, as it is biased particularly in favor of C over A. Bayesian analysis of 12 concatenated mitochondrial protein sequences for O. bidens and 46 other teleostean taxa supports the monophyly of Cypriniformes and Otophysi and results in a robust estimate of the otophysan phylogeny.     

Structure and expression of the overlapping ND4L and ND5 genes of Neurospora crassa mitochondria

Summary Genes homologous to the mammalian mitochondrial NADH dehydrogenase subunit genes ND4L and ND5 were identified in the mitochondrial genome of the filamentous fungus Neurospora crassa, and the structure and expression of these genes was examined. The ND4L gene (interrupted by one intervening sequence) potentially encodes an 89 residue long hydrophobic protein that shares about 26% homology (or 41% homology if conservative amino acid substitutions are allowed) with the analogous human mitochondrial protein. The ND5 gene (which contains two introns) encodes a 715 residue polypeptide that shares 23% homology with the human analogue; a 300 amino acid long region is highly conserved (50% homology) in the two ND5 proteins. The stop codon of the ND4L gene overlaps the initiation codon of the downstream ND5 gene, and the two genes are contranscribed and probably cotranslated. A presumed mature dicistronic (ND4L plus ND5) RNA was detected. The postulated mRNA (about 3.2 kb) contains 5 and 3 non-coding regions of about 86 and 730 nucleotides, respectively; this species is generated from very large precursor RNAs by a complex processing pathway. The ND4L and ND5 introns are all stable after their excision from the precursor species.Abbreviations bp base pairs - rRNA ribosomal RNA - ND NADH dehydrogenase - URF unidentified reading frame - kDal kilodaltons; a.a., amino acid     

Salmonid Opsin Sequences Undergo Positive Selection and Indicate an Alternate Evolutionary Relationship in Oncorhynchus

Positive selection can be demonstrated by statistical analysis when non-synonymous nucleotide substitutions occur more frequently than synonymous substitutions (dN>dS). This pattern of sequence evolution has been observed in the rhodopsin gene of cichlids. Mutations in opsin genes resulting in amino acid (AA) replacement appear to be associated with the evolution of specific color patterns and the evolution of courtship behaviors. Within fish, AA replacements in opsin proteins have improved vision at great depths and have occurred in deep-sea species. Salmonids experience diverse photic environments during their life history. Furthermore, sexual selection has resulted in species-specific male and female coloration during spawning. To look for evidence of positive selection in salmonid opsins, we sequenced the RH1, RH2, LWS, SWS1, and SWS2 genes from six Pacific salmon species as well as the Atlantic salmon. These salmonids include landlocked and migratory species and species that vary in their coloration during spawning. In each opsin gene comparison from all species sampled, traditional dN:dS analysis did not indicate positive selection. However, the more sensitive Creevey–McInerney statistical analysis indicates that RH1 and RH2 experienced positive selection early in the evolution and speciation of salmonids.     

The complete mitochondrial genome sequence and gene organization of the mud crab (Scylla paramamosain) with phylogenetic consideration

The complete mitochondrial genome is of great importance for better understanding the genome-level characteristics and phylogenetic relationships among related species. In the present study, we determined the complete mitochondrial genome DNA sequence of the mud crab (Scylla paramamosain) by 454 deep sequencing and Sanger sequencing approaches. The complete genome DNA was 15,824 bp in length and contained a typical set of 13 protein-coding genes, 22 transfer RNA (tRNA) genes, two ribosomal RNA (rRNA) genes and a putative control region (CR). Of 37 genes, twenty-three were encoded by the heavy strand (H-strand), while the other ones were encoded by light strand (L-strand). The gene order in the mitochondrial genome was largely identical to those obtained in most arthropods, although the relative position of gene tRNA^His differed from other arthropods. Among 13 protein-coding genes, three (ATPase subunit 6 (ATP6), NADH dehydrogenase subunits 1 (ND1) and ND3) started with a rare start codon ATT, whereas, one gene cytochrome c oxidase subunit I (COI) ended with the incomplete stop codon TA. All 22 tRNAs could fold into a typical clover-leaf secondary structure, with the gene sizes ranging from 63 to 73 bp. The phylogenetic analysis based on 12 concatenated protein-coding genes showed that the molecular genetic relationship of 19 species of 11 genera was identical to the traditional taxonomy.     

A phylogenetic study of the malagasy couas with insights into cuckoo relationships

The avian family Cuculidae (cuckoos) is a diverse group of birds that vary considerably in behaviors of interest to behavioral ecologists, e.g., obligate brood parasitism and cooperative breeding. The taxonomy of this group has historically been relatively stable but has not been extensively evaluated using molecular methods. The goal of this study was to evaluate phylogenetic relationships within the ecologically diverse genus Coua and the placement of Coua among major cuckoo lineages. We sequenced 429 bp of cytochrome b (cyt b) and 522 bp of ND2, both mitochondrial genes, for 26 species of cuckoos spanning 13 genera. We also included the enigmatic hoatzin (Opisthocomus hoazin) and used two Tauraco species as outgroups. ND2 exhibited higher rates of DNA sequence and amino acid substitution than cyt b; however, this did not greatly affect the overall levels of phylogenetic resolution and support provided by these two genes. Combined analyses produced two alternative phylogenies, depending on weighting scheme, both of which were fully resolved and were characterized by high bootstrap support. These phylogenies recovered monophyly for all of the traditional cuckoo subfamilies and indicated, with strong support, that the hoatzin is outside of Cuculidae. Within Coua, an arboreal and a terrestrial clade were identified. In contrast, habitat choice of Coua species did not greatly reflect the phylogeny.     

Upregulation of human mitochondrial NADH dehydrogenase subunit 5 in intestinal epithelial cells is modulated by Vibrio cholerae pathogenesis

Cholera still remains an important global predicament especially in India and other developing countries. Vibrio cholerae, the etiologic agent of cholera, colonizes the small intestine and produces an enterotoxin that is largely responsible for the watery diarrheal symptoms of the disease. Using RNA arbitrarily primed PCR, ND5 a mitochondria encoded subunit of complex I of the mitochondrial respiratory chain was found to be upregulated in the human intestinal epithelial cell line Int407 following exposure to V. cholerae. The upregulation of ND5 was not observed when Int407 was infected with Escherichia coli strains. Incubation with heat-killed V. cholerae or cholera toxin or culture supernatant also showed no such upregulation indicating the involvement of live bacteria in the process. Infection of the monolayer with aflagellate non-motile mutant of V. cholerae O395 showed a very significant (59-fold) downregulation of ND5. In contrast, a remarkable upregulation of ND5 expression (200-fold) was observed in a hyperadherent icmF insertion mutant with reduced motility. V. cholerae cheY4 null mutant defective in adherence and motility also resulted in significantly reduced levels of ND5 expression while mutant with the cheY4 gene duplicated showing increased adherence and motility resulted in increased expression of ND5. These results clearly indicate that both motility and adherence to intestinal epithelial cells are possible triggering factors contributing to ND5 mRNA expression by V. cholerae. Interestingly infection with insertion mutant in the gene coding for ToxR, the master regulator of virulence in V. cholerae resulted in significant downregulation of ND5 expression. However, infection with ctxA or toxT insertion mutants did not show any significant changes in ND5 expression compared to wild-type. Almost no expression of ND5 was observed in case of mutation in the gene coding for OmpU, a ToxR activated protein. Thus, infection of Int407 with virulence mutant strains of V. cholerae revealed that the ND5 expression is modulated by the virulence of V. cholerae in a ToxT independent manner. Although no difference in the mitochondrial copy number could be detected between infected and uninfected cells, the modulation of the expression of other mitochondrial genes were also observed. Incidentally, upon V. cholerae infection, complex I activity was found to increase about 3-folds after 6 h. This is the first report of alteration in mitochondrial gene expression upon infection of a non-invasive enteric bacterium like V. cholerae showing its modulation with adherence, motility and virulence of the organism.     

Variation in Coding (NADH Dehydrogenase Subunits 2, 3, and 6) and Noncoding Intergenic Spacer Regions of the Mitochondrial Genome in Octocorallia (Cnidaria: Anthozoa)

Low rates of evolution in cnidarian mitochondrial genes such as COI and 16S rDNA have hindered molecular systematic studies in this important invertebrate group. We sequenced fragments of 3 mitochondrial protein-coding genes (NADH dehydrogenase subunits ND2, ND3 and ND6) as well as the COI-COII intergenic spacer, the longest noncoding region found in the octocoral mitochondrial genome, to determine if any of these regions contain levels of variation sufficient for reconstruction of phylogenetic relationships among genera of the anthozoan subclass Octocorallia. Within and between the soft coral families Alcyoniidae and Xeniidae, sequence divergence in the genes ND2 (539 bp), ND3 (102 bp), and ND6 (444 bp) ranged from 0.5% to 12%, with the greatest pairwise distances between the 2 families. The COI-COII intergenic spacer varied in length from 106 to 122 bp, and pairwise sequence divergence values ranged from 0% to 20.4%. Phylogenetic trees constructed using each region separately were poorly resolved. Better phylogenetic resolution was obtained in a combined analysis using all 3 protein-coding regions (1085 bp total). Although relationships among some pairs of species and genera were well supported in the combined analysis, the base of the alcyoniid family tree remained an unresolved polytomy. We conclude that variation in the NADH subunit coding regions is adequate to resolve phylogenetic relationships among families and some genera of Octocorallia, but insufficient for most species - or population-level studies. Although the COI-COII intergenic spacer exhibits greater variability than the protein-coding regions and may contain useful species-specific markers, its short length limits its phylogenetic utility.     

The complete mitochondrial genome of Taeniogonalos taihorina (Bischoff) (Hymenoptera: Trigonalyidae) reveals a novel gene rearrangement pattern in the Hymenoptera

The family Trigonalyidae is considered to be one of the most basal lineages in the suborder Apocrita of Hymenoptera. Here, we determine the first complete mitochondrial genome of the Trigonalyidae, from the species Taeniogonalos taihorina (Bischoff, 1914). This mitochondrial genome is 15,927 bp long, with a high A + T-content of 84.60%. It contains all of the 37 typical animal mitochondrial genes and an A + T-rich region. The orders and directions of all genes are different from those of previously reported hymenopteran mitochondrial genomes. Eight tRNA genes, three protein-coding genes and the A + T-rich region were rearranged, with the dominant gene rearrangement events being translocation and local inversion. The arrangements of three tRNA clusters, trnY–trnM–trnI–trnQ, trnW–trnL2–trnC, and trnH–trnA–trnR–trnN–trnS–trnE–trnF, and the position of the cox1 gene, are novel to the Hymenoptera, even the insects. Six long intergenic spacers are present in the genome. The secondary structures of the RNA genes are normal, except for trnS2, in which the D-stem pairing is absent.     

Temporal, spatial, and ecological modes of evolution of Eurasian Mus based on mitochondrial and nuclear gene sequences

We sequenced mitochondrial (cytochrome b, 12S rRNA) and nuclear (IRBP, RAG1) genes for 17 species of the Old World murine genus Mus, drawn primarily from the Eurasian subgenus Mus. Phylogenetic analysis of the newly and previously available sequences support recognition of four subgenera within Mus (Mus, Coelomys, Nannomys, and Pyromys), with an unresolved basal polytomy. Our data further indicate that the subgenus Mus contains three distinct 'species groups': (1) a Mus booduga Species Group, also including Mus terricolor and Mus fragilicauda (probably also Mus famulus); (2) a Mus cervicolor Species Group, also including Mus caroli and Mus cookii; and (3) a Mus musculus Species Group, also including Mus macedonicus, Mus spicilegus, and Mus spretus. Species diversity in Eurasian Mus is probably explicable in terms of several phases of range expansion and vicariance, and by a propensity within the group to undergo biotope transitions. IRBP and RAG1 molecular clocks for Mus date the origin of subgenera to around 5-6 mya and the origin of Species Groups within subgenus Mus to around 2-3 mya. The temporal pattern of evolution among Eurasian Mus is more complex than that within the Eurasian temperate genus Apodemus.     

New primers for amplifying and sequencing the mitochondrial ND4/ND5 gene region of the Cypriniformes (Actinopterygii: Ostariophysi)

We provide 15 new primers for amplifying and sequencing the mitochondrial ND4/ND5 gene region of the Cypriniformes in an attempt to resolve relationships of this diverse group of freshwater fishes with extensive taxonomic sampling. Sequences from this region have the following desirable characteristics for phylogenetic analyses, some of which are lacking from the more commonly used cyt b and 12S/16S rRNA genes: they are (1) easy to align, (2) relatively long (ca. 3.4 kb), and (3) contain more phylogenetically informative variation at 1st and 2nd codon positions. Moreover, the ND4/ND5 gene region is easy to amplify and sequence when employing the protocol suggested herein.     

Complete nucleotide sequence and gene organization of the mitochondrial genome of Paa spinosa (Anura: Ranoidae)

The mt genome of Paa spinosa (Anura: Ranoidae) is a circular molecule of 18,012 bp in length, containing 38 genes (including an extra copy of tRNA-Met gene). This mt genome is characterized by three distinctive features: a cluster of rearranged tRNA genes (LTPF tRNA gene cluster), a tandem duplication of tRNA-Met gene (Met1 and Met2), and distinct repeat regions at both 5′ and 3′-sides in the control region. Comparing the locations and the sequences of all tRNA-Met genes among Ranoidae, and constructing NJ tree of the nucleotide of those tRNA-Met genes, we suggested a tandem duplication of tRNA-Met gene can be regarded as a synapomorphy of Dicroglossinae. To further investigate the phylogenetic relationships of anurans, phylogenetic analyses (BI, ML and MP) based on the nucleotide dataset and the corresponding amino acid dataset of 11 protein-coding genes (except ND5 and ATP8) arrived at the similar topology.