Genes — Open Access Journal

Measuring the magnitude of differentiation between populations based on genetic markers is commonplace in ecology, evolution, and conservation biology. The predominant differentiation metric used for this purpose is F_ST. Based on a qualitative survey, numerical analyses, simulations, and empirical data, I here argue that F_ST does not express the relationship to allele frequency differentiation between populations generally considered interpretable and desirable by researchers. In particular, F_ST (1) has low sensitivity when population differentiation is weak, (2) is contingent on the minor allele frequency across the populations, (3) can be strongly affected by asymmetry in sample sizes, and (4) can differ greatly among the available estimators. Together, these features can complicate pattern recognition and interpretation in population genetic and genomic analysis, as illustrated by empirical examples, and overall compromise the comparability of population differentiation among markers and study systems. I argue that a simple differentiation metric displaying intuitive properties, the absolute allele frequency difference AFD, provides a valuable alternative to F_ST. I provide a general definition of AFD applicable to both bi- and multi-allelic markers and conclude by making recommendations on the sample sizes needed to achieve robust differentiation estimates using AFD. Full article

Wheat is a staple food commodity grown worldwide, and wheat starch is a valuable source of energy and carbon that constitutes 80% of the grain weight. Manipulation of genes involved in starch synthesis significantly affects wheat grain weight and yield. TaSSIV plays an important role in starch synthesis and its main function is granule formation. To mine and stack more favorable alleles, single nucleotide polymorphisms (SNPs) of TaSSIV-A, B, and D were investigated across 362 wheat accessions by Ecotype-Targeting Induced Local Lesions IN Genome (EcoTILLING). As a result, a total of 38 SNPs in the amplified regions of three TaSSIV genes were identified, of which 10, 15, and 13 were in TaSSIV-A, B, and D, respectively. These 38 SNPs were evaluated by using KASP and six SNPs showed an allele frequency >5% whereas the rest were <5%, i.e., considered to be minor alleles. In the Chinese mini core collection, three haplotypes were detected for TaSSIV–A and three for TaSSIV–B. The results of an association study in the Chinese mini core collection with thousand grain weight (TGW) and spike length (SPL) showed that Hap-2-1A was significantly associated with TGW and Hap-3-1B with SPL. Allelic frequency and geographic distribution indicated that the favored haplotype (Hap-2-1A) has been positively selected in Chinese wheat breeding. These results suggested that the Kompetitive Allele Specific PCR (KASP) markers can be applied in starch improvement to ultimately improve wheat yield by marker assisted selection in wheat breeding. Full article

The Hyrcanian Forests present a unique Tertiary relict ecosystem, covering the northern Elburz and Talysh Ranges (Iran, Azerbaijan), a poorly investigated, unique biodiversity hotspot with many cryptic species. Since the 1970s, two nominal species of Urodela, Hynobiidae, Batrachuperus (later: Paradactylodon) have been described: Paradactylodon persicus from northwestern and P. gorganensis from northeastern Iran. Although P. gorganensis has been involved in studies on phylogeny and development, there is little data on the phylogeography, systematics, and development of the genus throughout the Hyrcanian Forests; genome-wide resources have been entirely missing. Given the huge genome size of hynobiids, making whole genome sequencing hardly affordable, we aimed to publish the first transcriptomic resources for Paradactylodon from an embryo and a larva (9.17 Gb RNA sequences; assembled to 78,918 unigenes). We also listed 32 genes involved in vertebrate sexual development and sex determination. Photographic documentation of the development from egg sacs across several embryonal and larval stages until metamorphosis enabled, for the first time, comparison of the ontogeny with that of other hynobiids and new histological and transcriptomic insights into early gonads and timing of their differentiation. Transcriptomes from central Elburz, next-generation sequencing (NGS) libraries of archival DNA of topotypic P. persicus, and GenBank-sequences of eastern P. gorganensis allowed phylogenetic analysis with three mitochondrial genomes, supplemented by PCR-amplified mtDNA-fragments from 17 museum specimens, documenting <2% uncorrected intraspecific genetic distance. Our data suggest that these rare salamanders belong to a single species P. persicus s.l. Humankind has a great responsibility to protect this species and the unique biodiversity of the Hyrcanian Forest ecosystems. Full article

Temperature is one of the most important environmental factors affecting flowering in plants. Adonis amurensis, a perennial herbaceous flower that blooms in early spring in northeast China where the temperature can drop to −15 °C, is an ideal model for studying the molecular mechanisms of flowering at extremely low temperatures. This study first investigated global gene expression profiles at different developmental stages of flowering in A. amurensis by RNA-seq transcriptome and iTRAQ proteomics. Finally, 123 transcription factors (TFs) were detected in both the transcriptome and the proteome. Of these, 66 TFs belonging to 14 families may play a key role in multiple signaling pathways of flowering in A. amurensis. The TFs FAR1, PHD, and B3 may be involved in responses to light and temperature, while SCL, SWI/SNF, ARF, and ERF may be involved in the regulation of hormone balance. SPL may regulate the age pathway. Some members of the TCP, ZFP, MYB, WRKY, and bHLH families may be involved in the transcriptional regulation of flowering genes. The MADS-box TFs are the key regulators of flowering in A. amurensis. Our results provide a direction for understanding the molecular mechanisms of flowering in A. amurensis at low temperatures. Full article

Today, elasmobranchs are one the most threatened vertebrate groups worldwide. In fact, at least 90% of elasmobranch species are listed in the International Union for Conservation of Nature (IUCN) Red List, while more than 40% are data-deficient. Although these vertebrates are mainly affected by unsustainable fishery activities, bycatch is also one of the major threats to sharks and batoids worldwide, and represents a challenge for both sustainable fishery management and for biodiversity and conservational efforts. Thus, in this study, DNA barcode methodology was used to identify the bycatch composition of batoid species from small-scale industrial fisheries in the southwest Atlantic and artisanal fisheries from southeast Brazil. A total of 228 individuals belonging to four Chondrichthyes orders, seven families, and at least 17 distinct batoid species were sequenced; among these individuals, 131 belonged to species protected in Brazil, 101 to globally threatened species, and some to species with trade restrictions provided by Appendix II of the Convention on International Trade in Endangered Species (CITES). These results highlight the impacts on marine biodiversity of bycatch by small-scale industrial and unmanaged artisanal fisheries from the southwest Atlantic, and support the implementation of DNA-based methodologies for species-specific identification in data-poor fisheries as a powerful tool for improving the quality of fisheries’ catch statistics and for keeping precise bycatch records. Full article

Salt stress has detrimental effects on plant growth and development. MicroRNAs (miRNAs) are a class of noncoding RNAs that are involved in post-transcriptional gene expression regulation. In this study, small RNA sequencing was employed to identify the salt stress-responsive miRNAs of the salt-sensitive Hassawi-3 and the salt-tolerant ILB4347 genotypes of faba bean, growing under salt stress. A total of 527 miRNAs in Hassawi-3 plants, and 693 miRNAs in ILB4347 plants, were found to be differentially expressed. Additionally, 284 upregulated and 243 downregulated miRNAs in Hassawi-3, and 298 upregulated and 395 downregulated miRNAs in ILB4347 plants growing in control and stress conditions were recorded. Target prediction and annotation revealed that these miRNAs regulate specific salt-responsive genes, which primarily included genes encoding transcription factors and laccases, superoxide dismutase, plantacyanin, and F-box proteins. The salt-responsive miRNAs and their targets were functionally enriched by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, which showed that the miRNAs were involved in salt stress-related biological pathways, including the ABC transporter pathway, MAPK signaling pathway, plant hormone signal transduction, and the phosphatidylinositol signaling system, among others, suggesting that the miRNAs play an important role in the salt stress tolerance of the ILB4347 genotype.These results offer a novel understanding of the regulatory role of miRNAs in the salt response of the salt-tolerant ILB4347 and the salt-sensitive Hassawi-3 faba bean genotypes. Full article

The sex of an animal influences its economic traits, especially in species displaying sexual dimorphism. The Chinese soft-shelled turtle, Pelodiscus sinensis, is an economically important aquatic species that shows significant male sexual dimorphism, with a large body size, faster growth, a thick and wide calipash, and lower body fat. In this study, ten male and ten female turtles were subjected to restriction site-associated DNA sequencing (RAD-seq) using the Hi-Seq 4000 sequencing platform to isolate female-specific DNA fragments. We identified 5967 bp and 6532 bp fragments using genome walking. Three female-specific markers designed from these two fragments were confirmed to separate the sexes of Pelodiscus sinensis perfectly. One of the female-specific markers showed dosage association in female and male individuals. Individuals from different populations (n = 296) were used to validate that the female-specific markers could identify the genetic sex of Pelodiscus sinensis with 100% accuracy. The results of the present study demonstrated that RAD-seq was useful to develop sex-related markers in animals, and verified that the sex determination system of Pelodiscus sinensis belonged to the ZZ/ZW heterogametic system. Importantly, the developed markers could lead to a method for sex-controlled breeding in the Chinese soft-shelled turtle. Full article

Avian genomes have perplexed researchers by being conservative in both size and rearrangements, while simultaneously holding the blueprints for a massive species radiation during the last 65 million years (My). Transposable elements (TEs) in bird genomes are relatively scarce but have been implicated as important hotspots for chromosomal inversions. In zebra finch (Taeniopygia guttata), long terminal repeat (LTR) retrotransposons have proliferated and are positively associated with chromosomal breakpoint regions. Here, we present the genome, karyotype and transposons of blue-capped cordon-bleu (Uraeginthus cyanocephalus), an African songbird that diverged from zebra finch at the root of estrildid finches 10 million years ago (Mya). This constitutes the third linked-read sequenced genome assembly and fourth in-depth curated TE library of any bird. Exploration of TE diversity on this brief evolutionary timescale constitutes a considerable increase in resolution for avian TE biology and allowed us to uncover 4.5 Mb more LTR retrotransposons in the zebra finch genome. In blue-capped cordon-bleu, we likewise observed a recent LTR accumulation indicating that this is a shared feature of Estrildidae. Curiously, we discovered 25 new endogenous retrovirus-like LTR retrotransposon families of which at least 21 are present in zebra finch but were previously undiscovered. This highlights the importance of studying close relatives of model organisms. Full article

Infection with canine heartworm (Dirofilaria immitis), spread via mosquito vectors, causes coughing, asthma, pneumonia, and bronchitis in humans and other animals. The disease is especially severe and often fatal in dogs and represents a serious threat to public health worldwide. Cysteine protease inhibitors (CPIs), also known as cystatins, are major immunomodulators of the host immune response during nematode infections. Herein, we cloned and expressed the cystatin Di-CPI from D. immitis. Sequence analysis revealed two specific cystatin-like domains, a Q-x-V-x-G motif, and a SND motif. Phylogenetic analysis indicates that Di-CPI is a member of the second subgroup of nematode type II cystatins. Probing of D. immitis total proteins with anti-rDi-CPI polyclonal antibody revealed a weak signal, and immunofluorescence-based histochemical analysis showed that native Di-CPI is mainly localized in the cuticle of male and female worms and the gut of male worms. Treatment of canine peripheral blood mononuclear cells (PMBCs) with recombinant Di-CPI induced a Th2-type immune response characterized by high expression of the anti-inflammatory factor interleukin-10. Proliferation assays showed that Di-CPI inhibits the proliferation of canine PMBCs by 15%. Together, the results indicate that Di-CPI might be related to cellular hyporesponsiveness in dirofilariasis and may help D. immitis to evade the host immune system. Full article

The concept of the human oral microbiome was applied to understand health and disease, lifestyles, and dietary habits throughout part of human history. In the present study, we augment the understanding of ancient oral microbiomes by characterizing human dental calculus samples recovered from the ancient Abbey of Badia Pozzeveri (central Italy), with differences in socioeconomic status, time period, burial type, and sex. Samples dating from the Middle Ages (11th century) to the Industrial Revolution era (19th century) were characterized using high-throughput sequencing of the 16S ribosomal RNA (rRNA) gene V4 region. Consistent with previous studies, individuals from Badia Pozzeveri possessed commensal oral bacteria that resembled modern oral microbiomes. These results suggest that members of the oral microbiome are ubiquitous despite differences in geographical regions, time period, sex, and socioeconomic status. The presence of fecal bacteria could be in agreement with poor hygiene practices, consistent with the time period. Respiratory tract, nosocomial, and other rare pathogens detected in the dental calculus samples are intriguing and could suggest subject-specific comorbidities that could be reflected in the oral microbiome. Full article

Motivation: Researchers in genomics are increasingly interested in epigenetic factors such as DNA methylation because they play an important role in regulating gene expression without changes in the sequence of DNA. Abnormal DNA methylation is associated with many human diseases. Results: We propose two different approaches to test for differentially methylated regions (DMRs) associated with complex traits, while accounting for correlations among CpG sites in the DMRs. The first approach is a nonparametric method using a kernel distance statistic and the second one is a likelihood-based method using a binomial spatial scan statistic. The kernel distance method uses the kernel function, while the binomial scan statistic approach uses a mixed-effects model to incorporate correlations among CpG sites. Extensive simulations show that both approaches have excellent control of type I error, and both have reasonable statistical power. The binomial scan statistic approach appears to have higher power, while the kernel distance method is computationally faster. The proposed methods are demonstrated using data from a chronic lymphocytic leukemia (CLL) study. Full article

A growing tumor is constantly secreting inflammatory chemokines and cytokines that induce release of immature myeloid cells, including myeloid-derived suppressor cells (MDSCs) and macrophages, from the bone marrow. These cells not only promote tumor growth, but also prepare distant organs for tumor metastasis. On the other hand, the myeloid-derived cells also have phagocytic potential, and can serve as vehicles for drug delivery. We have previously identified thioaptamers that bind a subset of MDSCs with high affinity and specificity. In the current study, we applied one of the thioaptamers as a probe to track myeloid cell distribution in the bone, liver, spleen and tumor in multiple murine models of breast cancer including the 4T1 syngeneic model and MDA-MB-231 and SUM159 xenograft models. Information generated from this study will facilitate further understanding of tumor growth and metastasis, and predict biodistribution patterns of cell-mediated drug delivery. Full article

Alternative splicing (AS) is a post-transcriptional regulatory process that enhances transcriptome diversity, thereby affecting plant growth, development, and stress responses. To identify the new transcripts and changes in the isoform-level AS landscape of rapeseed (Brassica napus) infected with the fungal pathogen Leptosphaeria maculans, we compared eight RNA-seq libraries prepared from mock-inoculated and inoculated B. napus cotyledons and stems. The AS events that occurred in stems were almost the same as those in cotyledons, with intron retention representing the most common AS pattern. We identified 1892 differentially spliced genes between inoculated and uninoculated plants. We performed a weighted gene co-expression network analysis (WGCNA) to identify eight co-expression modules and their Hub genes, which are the genes most connected with other genes within each module. There are nine Hub genes, encoding nine transcription factors, which represent key regulators of each module, including members of the NAC, WRKY, TRAF,AP2/ERF-ERF, C2H2,C2C2-GATA, HMG, bHLH, and C2C2-CO-likefamilies. Finally, 52 and 117 alternatively spliced genes in cotyledons and stems were also differentially expressed between mock-infected and infected materials, such as HMG and C2C2-Dof; which have dual regulatory mechanisms in response to L. maculans. The splicing of the candidate genes identified in this study could be exploited to improve resistance to L. maculans. Full article

Background: Exercise is one of the best nonpharmacologic therapies to treat hypertension. The blood pressure (BP) response to exercise is heritable. Yet, the genetic basis for the antihypertensive effects of exercise remains elusive. Methods: To assemble a prioritized gene signature, we performed a systematic review with a series of Boolean searches in PubMed (including Medline) from earliest coverage. The inclusion criteria were human genes in major BP regulatory pathways reported to be associated with: (1) the BP response to exercise; (2) hypertension in genome-wide association studies (GWAS); (3) the BP response to pharmacotherapy; (4a) physical activity and/or obesity in GWAS; and (4b) BP, physical activity, and/or obesity in non-GWAS. Included GWAS reports disclosed the statistically significant thresholds used for multiple testing. Results: The search yielded 1422 reports. Of these, 57 trials qualified from which we extracted 11 genes under criteria 1, 18 genes under criteria 2, 28 genes under criteria 3, 27 genes under criteria 4a, and 29 genes under criteria 4b. We also included 41 genes identified from our previous work. Conclusions: Deep-sequencing the exons of this systematically assembled signature of genes represents a cost and time efficient approach to investigate the genomic basis for the antihypertensive effects of exercise. Full article

Tunicates or urochordates—comprising ascidians, larvaceans, and salps—are the only metazoans that can synthesize cellulose, a biological function usually associated with bacteria and plants but not animals. Tunicate cellulose or tunicine is a major component of the outer acellular coverage (tunic) of the entire body of these organisms. Previous studies have suggested that the prokaryotic cellulose synthase gene (CesA) was horizontally transferred into the genome of a tunicate ancestor. However, no convenient tools have been devised to determine whether only tunicates harbor CesA. ORTHOSCOPE is a recently developed tool used to identify orthologous genes and to examine the phylogenic relationship of molecules within major metazoan taxa. The present analysis with this tool revealed the presence of CesA orthologs in all sequenced tunicate genomes but an absence in other metazoan genomes. This supports an evolutionary origin of animal cellulose and provides insights into the evolution of this animal taxon. Full article

: Early stage prediction of economic trait performance is important and directly linked to profitability of farm pig production. Genome-wide association study (GWAS) has been applied to find causative genomic regions of traits. This study established a regulatory gene network using GWAS for critical economic pig characteristics, centered on easily measurable body fat thickness in live animals. We genotyped 2,681 pigs using Illumina Porcine SNP60, followed by GWAS to calculate Bayes factors for 47,697 single nucleotide polymorphisms (SNPs) of seven traits. Using this information, SNPs were annotated with specific genes near genome locations to establish the association weight matrix.The entire network consisted of 226 nodes and 6,921 significant edges. For in silico validation of their interactions, we conducted regulatory sequence analysis of predicted target genes of transcription factors (TFs). Three key regulatory TFs were identified to guarantee maximum coverage: AT-rich interaction domain 3B (ARID3B), glial cell missing homolog 1 (GCM1), and GLI family zinc finger 2 (GLI2).We identified numerous genes targeted by ARID3B, associated with cellular processes. GCM1 and GLI2 were involved in developmental processes, and their shared target genes regulated multicellular organismal process. This system biology-based function analysis might contribute to enhancing understanding of economic pig traits. Full article

Proturans are small, wingless, soil-dwelling arthropods, generally associated with the early diversification of Hexapoda. Their bizarre morphology, together with conflicting results of molecular studies, has nevertheless made their classification ambiguous. Furthermore, their limited dispersal capability (due to the primarily absence of wings) and their euedaphic lifestyle have greatly complicated species-level identification. Mitochondrial and nuclear markers have been applied herein to investigate and summarize proturan systematics at different hierarchical levels. Two new mitochondrial genomes are described and included in a phylum-level phylogenetic analysis, but the position of Protura could not be resolved with confidence due to an accelerated rate of substitution and extensive gene rearrangements. Mitochondrial and nuclear loci were also applied in order to revise the intra-class systematics, recovering three proturan orders and most of the families/subfamilies included as monophyletic, with the exception of the subfamily Acerentominae. At the species level, most morphologically described species were confirmed using molecular markers, with some exceptions, and the advantages of including nuclear, as well as mitochondrial, markers and morphology are discussed. At all levels, an enlarged taxon sampling and the integration of data from different sources may be of significant help in solving open questions that still persist on the evolutionary history of Protura. Full article

Breeding efforts in the American cranberry (Vaccinium macrocarpon Ait.), a North American perennial fruit crop of great importance, have been hampered by the limited genetic and phenotypic variability observed among cultivars and experimental materials. Most of the cultivars commercially used by cranberry growers today were derived from a few wild accessions bred in the 1950s. In different crops, wild germplasm has been used as an important genetic resource to incorporate novel traits and increase the phenotypic diversity of breeding materials. Vaccinium microcarpum (Turcz. ex Rupr.) Schmalh. and V. oxycoccos L., two closely related species, may be cross-compatible with the American cranberry, and could be useful to improve fruit quality such as phytochemical content. Furthermore, given their northern distribution, they could also help develop cold hardy cultivars. Although these species have previously been analyzed in diversity studies, genomic characterization and comparative studies are still lacking. In this study, we sequenced and assembled the organelle genomes of the cultivated American cranberry and its wild relative, V. microcarpum. PacBio sequencing technology allowed us to assemble both mitochondrial and plastid genomes at very high coverage and in a single circular scaffold. A comparative analysis revealed that the mitochondrial genome sequences were identical between both species and that the plastids presented only two synonymous single nucleotide polymorphisms (SNPs). Moreover, the Illumina resequencing of additional accessions of V. microcarpum and V. oxycoccos revealed high genetic variation in both species. Based on these results, we provided a hypothesis involving the extension and dynamics of the last glaciation period in North America, and how this could have shaped the distribution and dispersal of V. microcarpum. Finally, we provided important data regarding the polyploid origin of V. oxycoccos. Full article

Root morphology is essential for plant survival. NO₃⁻ is not only a nutrient, but also a signal substance affecting root growth in plants. However, the mechanism of NO₃⁻-mediated root growth in rice remains unclear. In this study, we investigated the effect of OsNRT2.1 on root elongation and nitrate signaling-mediated auxin transport using OsNRT2.1 overexpression lines. We observed that the overexpression of OsNRT2.1 increased the total root length in rice, including the seminal root length, total adventitious root length, and total lateral root length in seminal roots and adventitious roots under 0.5-mM NO₃^- conditions, but not under 0.5-mM NH₄⁺ conditions. Compared with wild type (WT), the ¹⁵NO₃⁻ influx rate of OsNRT2.1 transgenic lines increased by 24.3%, and the expressions of auxin transporter genes (OsPIN1a/b/c and OsPIN2) also increased significantly under 0.5-mM NO₃⁻ conditions. There were no significant differences in root length, ß-glucuronidase (GUS) activity, and the expressions of OsPIN1a/b/c and OsPIN2 in the pDR5::GUS transgenic line between 0.5-mM NO₃⁻ and 0.5-mM NH₄⁺ treatments together with N-1-naphthylphalamic acid (NPA) treatment. When exogenous NPA was added to 0.5-mM NO₃⁻ nutrient solution, there were no significant differences in the total root length and expressions of OsPIN1a/b/c and OsPIN2 between transgenic plants and WT, although the ¹⁵NO₃⁻ influx rate of OsNRT2.1 transgenic lines increased by 25.2%. These results indicated that OsNRT2.1 is involved in the pathway of nitrate-dependent root elongation by regulating auxin transport to roots; i.e., overexpressing OsNRT2.1 promotes an effect on root growth upon NO₃⁻ treatment that requires active polar auxin transport. Full article