Single-copy Nuclear Genes Research Articles

Sequence analysis of single-copy genes in two wild olive subspecies: nucleotide diversity and potential use for testing admixture.

The wild olive distribution extends from the Mediterranean region to south Asia and Austral Africa. The species is also invasive, particularly in Australia. Here, we investigated the sequence variation at five nuclear single-copy genes in 41 native and invasive accessions of the Mediterranean and African olive subspecies. The nucleotide diversity was assessed and the phylogenetic relationships between alleles were depicted with haplotype networks. A Bayesian clustering method (STRUCTURE) was applied to identify the main gene pools. We found an average of 18.4 alleles per locus. Native Mediterranean and African olives only share one allele, which testifies for ancient admixture on the Red Sea hills. The presence of divergent alleles in the Mediterranean olive, as well as the identification of two main genetic clusters, suggests a complex origin with two highly differentiated gene pools from the eastern and western Mediterranean that recently admixed. In the invasive range, relatively high nucleotide diversity is observed as a consequence of the introduction of alleles from two subspecies. Our data confirm that four invasive individuals are early-generation hybrids. Finally, the utility of single-copy gene sequences in olive population genomic and phylogenetic studies is briefly discussed.

Phylogenetic relationships in the grass family (Poaceae) based on the nuclear single copy locus topoisomerase 6 compared with chloroplast DNA

Phylogenetic relationships within the grass family were studied using a newly obtained locus of the nuclear single copy gene topoisomerase 6 (Topo6) spanning the four exons 8–11 and the chloroplast matK gene. Data were evaluated using maximum parsimony, maximum likelihood and Bayesian methods. All analyses showed genera Streptochaeta and Anomochloa as early diverging, followed by Pharus as sister to the rest of the Poaceae, and monophyly of the subfamily Anomochlooideae was supported by the nuclear dataset. The remaining grasses formed a strongly supported and monophyletic group, which split into the major clades BEP and PACMAD in the Topo6 analyses. Monophyly of the BEP clade was strongly supported by the Topo6 data. The results showed clearly incongruity between the two sets of data, such as the different subfamilial relationships of Bambusoideae, Ehrhartoideae and Pooideae. Most of the analysed species are representatives of subfamily Pooideae, which was analysed in more detail by PCR fragment length differences of another Topo6 region spanning the exons 17–19. Monophyly of Pooideae was strongly supported by the matK data, whereas the nuclear data placed Brachyelytrum outside of the remaining Pooideae. Relationships within the early evolutionary lineages remained largely unresolved in the phylogenetic trees, but the ‘core’ Pooideae (Aveneae/Poeae tribe complex and Hordeeae) were highly supported in all analyses. The differences in amplification lengths illustrate the tribe and subtribe classification of Pooideae. The comparatively conserved structure of the newly studied Topo6 region makes it a promising marker from the nuclear genome that could be successfully PCR-amplified to study higher-level phylogenetic relationships within grasses and perhaps between families within the order Poales.

A simple strategy for development of single nucleotide polymorphisms from non-model species and its application in Panax.

Single nucleotide polymorphisms (SNPs) are widely employed in the studies of population genetics, molecular breeding and conservation genetics. In this study, we explored a simple route to develop SNPs from non-model species based on screening the library of single copy nuclear genes (SCNGs). Through application of this strategy in Panax, we identified 160 and 171 SNPs from P. quinquefolium and P. ginseng, respectively. Our results demonstrated that both P. ginseng and P. quinquefolium possessed a high level of nucleotide diversity. The number of haplotype per locus ranged from 1 to 12 for P. ginseng and from 1 to 9 for P. quinquefolium, respectively. The nucleotide diversity of total sites (πT) varied between 0.000 and 0.023 for P. ginseng and 0.000 and 0.035 for P. quinquefolium, respectively. These findings suggested that this approach is well suited for SNP discovery in non-model organisms and is easily employed in standard genetics laboratory studies.

Development of a multiplex system to assess DNA persistence in taphonomic studies

In this study, we have developed a PCR multiplex that can be used to assess DNA degradation and at the same time monitor for inhibition: primers have been designed to amplify human, pig, and rabbit DNA, allowing pig and rabbit to be used as experimental models for taphonomic research, but also enabling studies on human DNA persistence in forensic evidence. Internal amplified controls have been added to monitor for inhibition, allowing the effects of degradation and inhibition to be differentiated. Sequence data for single-copy nuclear recombination activation gene (RAG-1) from human, pig, and rabbit were aligned to identify conserved regions and primers were designed that targeted amplicons of 70, 194, 305, and 384 bp. Robust amplification in all three species was possible using as little as 0.3 ng of template DNA. These have been combined with primers that will amplify a bacterial DNA template within the PCR. The multiplex has been evaluated in a series of experiments to gain more knowledge of DNA persistence in soft tissues, which can be important when assessing what material to collect following events such as mass disasters or conflict, when muscle or bone material can be used to aid with the identification of human remains. The experiments used pigs as a model species. When whole pig bodies were exposed to the environment in Northwest England, DNA in muscle tissue persisted for over 24 days in the summer and over 77 days in the winter, with full profiles generated from these samples. In addition to time, accumulated degree days (ADD) were also used as a measure that combines both time and temperature-24 days was in summer equivalent to 295 ADD whereas 77 days in winter was equivalent to 494 ADD.

The coding region of the UFGT gene is a source of diagnostic SNP markers that allow single-locus DNA genotyping for the assessment of cultivar identity and ancestry in grapevine (Vitis vinifera L.).

BackgroundVitis vinifera L. is one of society’s most important agricultural crops with a broad genetic variability. The difficulty in recognizing grapevine genotypes based on ampelographic traits and secondary metabolites prompted the development of molecular markers suitable for achieving variety genetic identification.FindingsHere, we propose a comparison between a multi-locus barcoding approach based on six chloroplast markers and a single-copy nuclear gene sequencing method using five coding regions combined with a character-based system with the aim of reconstructing cultivar-specific haplotypes and genotypes to be exploited for the molecular characterization of 157 V. vinifera accessions. The analysis of the chloroplast target regions proved the inadequacy of the DNA barcoding approach at the subspecies level, and hence further DNA genotyping analyses were targeted on the sequences of five nuclear single-copy genes amplified across all of the accessions. The sequencing of the coding region of the UFGT nuclear gene (UDP-glucose: flavonoid 3-0-glucosyltransferase, the key enzyme for the accumulation of anthocyanins in berry skins) enabled the discovery of discriminant SNPs (1/34 bp) and the reconstruction of 130 V. vinifera distinct genotypes. Most of the genotypes proved to be cultivar-specific, and only few genotypes were shared by more, although strictly related, cultivars.ConclusionOn the whole, this technique was successful for inferring SNP-based genotypes of grapevine accessions suitable for assessing the genetic identity and ancestry of international cultivars and also useful for corroborating some hypotheses regarding the origin of local varieties, suggesting several issues of misidentification (synonymy/homonymy).

Single-Copy Nuclear Genes Place Haustorial Hydnoraceae within Piperales and Reveal a Cretaceous Origin of Multiple Parasitic Angiosperm Lineages

Extreme haustorial parasites have long captured the interest of naturalists and scientists with their greatly reduced and highly specialized morphology. Along with the reduction or loss of photosynthesis, the plastid genome often decays as photosynthetic genes are released from selective constraint. This makes it challenging to use traditional plastid genes for parasitic plant phylogenetics, and has driven the search for alternative phylogenetic and molecular evolutionary markers. Thus, evolutionary studies, such as molecular clock-based age estimates, are not yet available for all parasitic lineages. In the present study, we extracted 14 nuclear single copy genes (nSCG) from Illumina transcriptome data from one of the “strangest plants in the world”, Hydnora visseri (Hydnoraceae). A ∼15,000 character molecular dataset, based on all three genomic compartments, shows the utility of nSCG for reconstructing phylogenetic relationships in parasitic lineages. A relaxed molecular clock approach with the same multi-locus dataset, revealed an ancient age of ∼91 MYA for Hydnoraceae. We then estimated the stem ages of all independently originated parasitic angiosperm lineages using a published dataset, which also revealed a Cretaceous origin for Balanophoraceae, Cynomoriaceae and Apodanthaceae. With the exception of Santalales, older parasite lineages tend to be more specialized with respect to trophic level and have lower species diversity. We thus propose the “temporal specialization hypothesis” (TSH) implementing multiple independent specialization processes over time during parasitic angiosperm evolution.

Transcriptome sequences resolve deep relationships of the grape family.

Previous phylogenetic studies of the grape family (Vitaceae) yielded poorly resolved deep relationships, thus impeding our understanding of the evolution of the family. Next-generation sequencing now offers access to protein coding sequences very easily, quickly and cost-effectively. To improve upon earlier work, we extracted 417 orthologous single-copy nuclear genes from the transcriptomes of 15 species of the Vitaceae, covering its phylogenetic diversity. The resulting transcriptome phylogeny provides robust support for the deep relationships, showing the phylogenetic utility of transcriptome data for plants over a time scale at least since the mid-Cretaceous. The pros and cons of transcriptome data for phylogenetic inference in plants are also evaluated.

The Strepsiptera-Odyssey: the history of the systematic placement of an enigmatic parasitic insect order

The history of the phylogenetic placement of the parasitic insect order Strepsiptera is outlined. The first species was described in 1793 by P. Rossi and assigned to the hymenopteran family Ichneumonidae. A position close to the cucujiform beetle family Rhipiphoridae was suggested by several earlier authors. Others proposed a close relationship with Diptera or even a group Pupariata including Diptera, Strepsiptera and Coccoidea. A subordinate placement within the polyphagan series Cucujiformia close to the wood-associated Lymexylidae was favored by the coleopterist R.A. Crowson. W. Hennig considered a sistergroup relationship with Coleoptera as the most likely hypothesis but emphasized the uncertainty. Cladistic analyses of morphological data sets yielded very different placements, alternatively as sistergroup of Coleoptera, Antliophora, or all other holometabolan orders. Results based on ribosomal genes suggested a sistergroup relationship with Diptera (Halteria concept). A clade Coleopterida (Strepsiptera and Coleoptera) was supported in two studies based on different combinations of protein coding nuclear genes. Analyses of data sets comprising seven or nine genes (7 single copy nuclear genes), respectively, yielded either a subordinate placement within Coleoptera or a sistergroup relationship with Neuropterida. Several early hypotheses based on a typological approach − affinities with Diptera, Coleoptera, a coleopteran subgroup, or Neuropterida − were revived using either a Hennigian approach or formal analyses of morphological characters or different molecular data sets. A phylogenomic approach finally supported a sistergroup relationship with monophyletic Coleoptera.

Phylogeny of the cycads based on multiple single-copy nuclear genes: congruence of concatenated parsimony, likelihood and species tree inference methods.

Despite a recent new classification, a stable phylogeny for the cycads has been elusive, particularly regarding resolution of Bowenia, Stangeria and Dioon. In this study, five single-copy nuclear genes (SCNGs) are applied to the phylogeny of the order Cycadales. The specific aim is to evaluate several gene tree-species tree reconciliation approaches for developing an accurate phylogeny of the order, to contrast them with concatenated parsimony analysis and to resolve the erstwhile problematic phylogenetic position of these three genera. DNA sequences of five SCNGs were obtained for 20 cycad species representing all ten genera of Cycadales. These were analysed with parsimony, maximum likelihood (ML) and three Bayesian methods of gene tree-species tree reconciliation, using Cycas as the outgroup. A calibrated date estimation was developed with Bayesian methods, and biogeographic analysis was also conducted. Concatenated parsimony, ML and three species tree inference methods resolve exactly the same tree topology with high support at most nodes. Dioon and Bowenia are the first and second branches of Cycadales after Cycas, respectively, followed by an encephalartoid clade (Macrozamia-Lepidozamia-Encephalartos), which is sister to a zamioid clade, of which Ceratozamia is the first branch, and in which Stangeria is sister to Microcycas and Zamia. A single, well-supported phylogenetic hypothesis of the generic relationships of the Cycadales is presented. However, massive extinction events inferred from the fossil record that eliminated broader ancestral distributions within Zamiaceae compromise accurate optimization of ancestral biogeographical areas for that hypothesis. While major lineages of Cycadales are ancient, crown ages of all modern genera are no older than 12 million years, supporting a recent hypothesis of mostly Miocene radiations. This phylogeny can contribute to an accurate infrafamilial classification of Zamiaceae.

Phylogenetic analysis revealed reticulate evolution of allotetraploid Elymus ciliaris

Increasing evidence has shown the complex and dynamic nature of polyploids. Two single copy nuclear genes were used to explore genome evolutionary dynamics and the origin of tetraploid E. ciliaris: the phosphoglycerate kinase (PGK1) and the second largest subunit of RNA polymerase II (RPB2) together with a chloroplast gene encoding ribosomal protein S16 (RPS16). RPS16 data confirmed that the maternal origin of E. ciliaris is the St genome species. Both RPB2 and PGK1 data supported that E. ciliairs has multiple origins, and originated from the Pseudorogneria (St) and unknown donor (Y) diploids. The St genome in E. ciliaris species has a complex evolutionary history. Both RPB2 and PGK1 data suggested the absence of St genome in accession PI 377532 of E. ciliaris. However, cpDNA RPS16 clearly indicated that its maternal origin is the same as other E. ciliaris accessions, and is St genomic diploid species. Results suggest that there are two lineages of St genome present in E. ciliaris species; one is grouped with Pseudoroegneria diploid species, the other is grouped with Hordeum (H) species (named St?). The Japanese accession PI 377532 might have introgression either from HordeumH genome species or from ElymusStH genome species with replacement of at least some nuclear St-loci by H-loci. The correlation between genome differentiation and geographical distribution is also discussed.

Single-copy nuclear gene primers for Streptanthus and other Brassicaceae from genomic scans, published data, and ESTs.

• Premise of the study: We report 11 primer sets for nine single-copy nuclear genes in Streptanthus and other Thelypodieae (Brassicaceae) and their utility at tribal-level and species-level phylogenetics in this poorly resolved group.• Methods and Results: We selected regions based on a cross-referenced matrix of previous studies and public Brassica expressed sequence tags. To design primers, we used alignments of low-depth-coverage Illumina sequencing of genomic DNA for two species of Brassica mapped onto Arabidopsis thaliana. We report several primer combinations for five regions that consistently amplified a single band and yielded high-quality sequences for at least 70% of the species assayed, and for four additional regions whose utility might be clade specific.• Conclusions: Our primers will be useful in improving resolution at shallow depths across the Thelypodieae, and likely in other Brassicaceae.

Phylogenetic relationships and Y genome origin in Elymus L. sensu lato (Triticeae; Poaceae) based on single-copy nuclear Acc1 and Pgk1 gene sequences

To estimate the origin and genomic relationships of the polyploid species within Elymus L. sensu lato, two unlinked single-copy nuclear gene (Acc1 and Pgk1) sequences of eighteen tetraploids (StH and StY genomes) and fourteen hexaploids (StStH, StYP, StYH, and StYW genomes) were analyzed with those of 35 diploid taxa representing 18 basic genomes in Triticeae. Sequence and phylogenetic analysis suggested that: (1) the St, H, W, and P genomes were donated by Pseudoroegneria, Hordeum, Australopyrum, and Agropyron, respectively, while the Y genome is closely related to the Xp genome in Peridictyon sanctum; (2) different hexaploid Elymus s.l. species may derived their StY genome from different StY genome tetraploid species via independent origins; (3) due to incomplete lineage sorting and/or hybridization events, the genealogical conflict between the two gene trees suggest introgression involving some Elymus s.l. species, Pseudoroegneria, Agropyron and Aegilops/Triticum; (4) it is reasonable to recognize the StH genome species as Elymus sensu stricto, the StY genome species as Roegneria, the StYW genome species as Anthosachne, the StYH genome species as Campeiostachys, and the StYP genome species as Kengyilia. The occurrence of multiple origin and introgression could account for the rich diversity and ecological adaptation of Elymus s.l. species.

Phylogenetic reconstruction and diversification of the Triticeae (Poaceae) based on single-copy nuclear Acc1 and Pgk1 gene data

Two single-copy nuclear gene (Acc1 and Pgk1) sequences were used to estimate the phylogeny and diversification patterns of the Triticeae. Phylogenetic analyses and diversification patterns suggested that (1) a major radiation occurring in the Triticeae about 6.1–9.2 MYA might have been triggered by the late Miocene climate oscillations; (2) the Triticeae lineages from Mediterranean and Arctic-temperate regions were evolutionarily distinct; (3) the relationship between Thinopyrum bessarabicum and Triticum/Aegilops is more closely than that between Lophopyrum elongatum and Triticum/Aegilops; (4) Pseudoroegneria is closely related to Australopyrum and Lophopyrum; (5) Australopyrum might originate from colonization via South-east Asia during the late Miocene; (6) Habitats, mating systems, and climate oscillations may associated with the diversification rate shifts leading to the majority of Mediterranean lineage. Diversification rate shifts in Mediterranean lineage of the Triticeae not only spur the occurrence of many endemic lineages but also provide opportunity for polyploid crop origin and domestication.

Comparative transcriptome analysis and marker development of two closely related Primrose species (Primula poissonii and Primula wilsonii)

BackgroundPrimula species are important early spring garden plants with a centre of diversity and speciation in the East Himalaya-Hengduan Mountains in Western China. Studies on population genetics, speciation and phylogeny of Primula have been impeded by a lack of genomic resources. In the present study, we sequenced the transcriptomes of two closely related primrose species, Primula poissonii and Primula wilsonii, using short reads on the Illumina Genome Analyzer platform.ResultsWe obtained 55,284 and 55,011 contigs with N50 values of 938 and 1,085 for P. poissonii and P. wilsonii, respectively, and 6,654 pairs of putative orthologs were identified between the two species. Estimations of non-synonymous/synonymous substitution rate ratios for these orthologs indicated that 877 of the pairs may be under positive selection (Ka/Ks > 0.5), and functional enrichment analysis revealed that significant proportions of the orthologs were in the categories DNA repair, stress resistance, which may provide some hints as to how the two closely related Primula species adapted differentially to extreme environments, such as habitats characterized by aridity, high altitude and high levels of ionizing radiation. It was possible for the first time to estimate the divergence time between the radiated species pair, P. poissonii and P. wilsonii; this was found to be approximately 0.90 ± 0.57 Mya, which falls between the Donau and Gunz glaciation in the Middle Pleistocene. Primers based on 54 pairs of orthologous SSR-containing sequences between the two Primula species were designed and verified. About half of these pairs successfully amplified for both species. Of the 959 single copy nuclear genes shared by four model plants (known as APVO genes), 111 single copy nuclear genes were verified as being present in both Primula species and exon-anchored and intron-spanned primers were designed for use.ConclusionWe characterized the transcriptomes for the two Primula species, and produced an unprecedented amount of genomic resources for these important garden plants. Evolutionary analysis of these two Primula species not only revealed a more precise divergence time, but also provided some novel insights into how differential adaptations occurred in extreme habitats. Furthermore, we developed two sets of genetic markers, single copy nuclear genes and nuclear microsatellites (EST-SSR). Both these sets of markers will facilitate studies on the genetic improvement, population genetics and phylogenetics of this rapidly adapting taxon.

Elongation factor-1α, a putative single-copy nuclear gene, has divergent sets of paralogs in an arachnid

Identification of paralogy in candidate nuclear loci is an important prerequisite in phylogenetics and statistical phylogeography, but one that is often overlooked. One marker commonly assumed to be a single-copy gene and claimed to harbor great utility for inferring recent divergences is elongation factor-1α (EF-1α). To test this hypothesis, we systematically cloned EF-1α in three disjunct populations of the harvestman Metasiro americanus. Here we show that EF-1α has a large number of paralogs in this species. The paralogs do not evolve in a concerted manner, and the paralogs diverged prior to the population divergence. Moreover, the paralogs of M. americanus are not comparable to the highly divergent EF-1α paralogs found in bees and spiders, which are easily recognized and separated through the use of specific primers. We demonstrate statistically that our detection of paralogs cannot be attributed to amplification error. The presence of EF-1α paralogs in M. americanus prevents its use in statistical phylogeography, and the presence of out-paralogs argues against its use in phylogenetic inference among recently diverged clades. These data contradict the common assumption that EF-1α is for most or all taxa a single-copy gene, or that it has a small number of paralogs that are homogenized through gene conversion, unequal crossing over, or other processes.

Radically different phylogeographies and patterns of genetic variation in two European brown frogs, genus Rana

We reconstruct range–wide phylogeographies of two widespread and largely co-occurring Western Palearctic frogs, Rana temporaria and R. dalmatina. Based on tissue or saliva samples of over 1000 individuals, we compare a variety of genetic marker systems, including mitochondrial DNA, single-copy protein-coding nuclear genes, microsatellite loci, and single nucleotide polymorphisms (SNPs) of transcriptomes of both species. The two focal species differ radically in their phylogeographic structure, with R. temporaria being strongly variable among and within populations, and R. dalmatina homogeneous across Europe with a single strongly differentiated population in southern Italy. These differences were observed across the various markers studied, including microsatellites and SNP density, but especially in protein-coding nuclear genes where R. dalmatina had extremely low heterozygosity values across its range, including potential refugial areas. On the contrary, R. temporaria had comparably high range-wide values, including many areas of probable postglacial colonization. A phylogeny of R. temporaria based on various concatenated mtDNA genes revealed that two haplotype clades endemic to Iberia form a paraphyletic group at the base of the cladogram, and all other haplotypes form a monophyletic group, in agreement with an Iberian origin of the species. Demographic analysis suggests that R. temporaria and R. dalmatina have genealogies of roughly the same time to coalescence (TMRCA ∼3.5mya for both species), but R. temporaria might have been characterized by larger ancestral and current effective population sizes than R. dalmatina. The high genetic variation in R. temporaria can therefore be explained by its early range expansion out of Iberia, with subsequent cycles of differentiation in cryptic glacial refugial areas followed by admixture, while the range expansion of R. dalmatina into central Europe is a probably more recent event.

Multilocus estimation of divergence times and ancestral effective population sizes of Oryza species and implications for the rapid diversification of the genus

· Despite substantial investigations into Oryza phylogeny and evolution, reliable estimates of the divergence times and ancestral effective population sizes of major lineages in Oryza are challenging. · We sampled sequences of 106 single-copy nuclear genes from all six diploid genomes of Oryza to investigate the divergence times through extensive relaxed molecular clock analyses and estimated the ancestral effective population sizes using maximum likelihood and Bayesian methods. · We estimated that Oryza originated in the middle Miocene (c. 13-15 million years ago; Ma) and obtained an explicit time frame for two rapid diversifications in this genus. The first diversification involving the extant F-/G-genomes and possibly the extinct H-/J-/K-genomes occurred in the middle Miocene immediately after (within < 1 Myr) the origin of Oryza. The second giving rise to the A-/B-/C-genomes happened c. 5-6 Ma. We found that ancestral effective population sizes were much larger than those of extant species in Oryza. · We suggest that the climate fluctuations during the period from the middle Miocene to Pliocene may have contributed to the two rapid diversifications of Oryza species. Such information helps better understand the evolutionary history of Oryza and provides further insights into the pattern and mechanism of diversification in plants in general.

Disruption of de Novo Mitochondrial Thymidylate Synthesis Increases mtDNA Content

Mitochondrial DNA (mtDNA) synthesis is necessary for the replication and repair of damaged DNA; maintaining the proper balance of nucleotides is required to avoid mtDNA depletion and mutation. Mitochondrial deoxynucleotide triphosphates (dNTPs) are obtained from either salvage pathways or de novo biosynthesis within the mitochondria. In the mitochondria, de novo dTMP biosynthesis requires the enzymes serine hydroxymethyltransferase 2 (SHMT2), thymidylate synthase (TYMS), and dihydrofolate reductase‐like 1 (DHFRL1). In the nucleus, depletion of dTMP synthesis leads to misincorporation of deoxyuridine into nuclear DNA (nuDNA) resulting in genomic instability. SHMT2 in the mitochondria is responsible for transferring a one‐carbon unit from serine to tetrahydrofolate (THF), producing 5,10‐methyleneTHF and glycine in a reversible reaction; 5–10‐methyleneTHF is utilized by TYMS to create dTMP from dUMP.Our goal is to determine how mtDNA content is affected by disruptions of de novo mitochondrial dTMP synthesis, using two different approaches to decrease SHMT2 activity. First, SHMT2 expression in mammalian HeLa cells was decreased using siRNA technology. Second, HeLa cells were cultured in glycine concentrations ranging from 0mM to 10mM, which inhibits the SHMT2‐catalyzed synthesis of 5,10‐methyleneTHF. The mtDNA and nuDNA copy numbers were determined by qPCR using primers specific for the mitochondrial tRNALeu(UUR) gene and a single copy nuclear gene, β2‐microglobulin. Effects of SHMT2 inhibition on deoxyuridine misincorporation into mtDNA are being assessed.Our preliminary results suggest that mitochondrial de novo dTMP synthesis is affected by SHMT2 expression and glycine concentration. Knockdown of SHMT2 results in an increase (2.8 fold) in mtDNA copy number. Similarly, mtDNA copy number increases with increasing glycine concentration in culture media (4 fold). Supported by NIH R37DK58144.

Phylogenetic Analysis and DNA-based Species Confirmation in Anopheles (Nyssorhynchus)

Specimens of neotropical Anopheles (Nyssorhynchus) were collected and identified morphologically. We amplified three genes for phylogenetic analysis–the single copy nuclear white and CAD genes, and the COI barcode region. Since we had multiple specimens for most species we were able to test how well the single or combined genes were able to corroborate morphologically defined species by placing the species into exclusive groups. We found that single genes, including the COI barcode region, were poor at confirming species, but that the three genes combined were able to do so much better. This has implications for species identification, species delimitation, and species discovery, and we caution that single genes are not enough. Higher level groupings were partially resolved with some well-supported groupings, whereas others were found to be either polyphyletic or paraphyletic. There were examples of known groups, such as the Myzorhynchella Section, which were poorly supported with single genes but were well supported with combined genes. From this we can infer that more sequence data will be needed in order to show more higher-level groupings with good support. We got unambiguously good support (0.94–1.0 Bayesian posterior probability) from all DNA-based analyses for a grouping of An. dunhami with An. nuneztovari and An. goeldii, and because of this and because of morphological similarities we propose that An. dunhami be included in the Nuneztovari Complex. We obtained phylogenetic corroboration for new species which had been recognised by morphological differences; these will need to be formally described and named.

Phylogenetic analysis in some Hordeum species (Triticeae; Poaceae) based on two single-copy nuclear genes encoding acetyl-CoA carboxylase

Abstract We investigate the phylogenetic relationship, and evolutionary history of 18 diploid and polyploid Hordeum species including 22 taxa based on two single-copy nuclear ACC1 and ACC2 genes using maximum parsimony, maximum likelihood, and Bayesian inference. The results of molecular phylogenetic analysis demonstrated genetic relationships among taxa and origin of polyploids. Our phylogenetic analyses revealed a clear alloploid origin of Hordeum capense , with Eurasian Hordeum marinum subsp. gussoneanum as the Xa genome donor and diploid Asian Hordeum roshevitzii as the H genome donor. The formation of hexaploid Hordeum lechleri likely involves hybridization between tetraploid Hordeum brachyantherum subsp. brachyantherum and a diploid possessing the I genome. The Acc1 and Acc2 gene data analyses suggested that Siberian Hordeum bogdanii might have be the common ancestor of the diploid New World Hordeum species. Perennial diploid South American species, Hordeum comosum was the first-diverging group within the clade of diploid American species in the analyses.