Menu
July 7, 2019  |  

Complete fusion of a transposon and herpesvirus created the Teratorn mobile element in medaka fish.

Mobile genetic elements (e.g., transposable elements and viruses) display significant diversity with various life cycles, but how novel elements emerge remains obscure. Here, we report a giant (180-kb long) transposon, Teratorn, originally identified in the genome of medaka, Oryzias latipes. Teratorn belongs to the piggyBac superfamily and retains the transposition activity. Remarkably, Teratorn is largely derived from a herpesvirus of the Alloherpesviridae family that could infect fish and amphibians. Genomic survey of Teratorn-like elements reveals that some of them exist as a fused form between piggyBac transposon and herpesvirus genome in teleosts, implying the generality of transposon-herpesvirus fusion. We propose that Teratorn was created by a unique fusion of DNA transposon and herpesvirus, leading to life cycle shift. Our study supports the idea that recombination is the key event in generation of novel mobile genetic elements. Teratorn is a large mobile genetic element originally identified in the small teleost fish medaka. Here, the authors show that Teratorn is derived from the fusion of a piggyBac superfamily DNA transposon and an alloherpesvirus and that it is widely found across teleost fish.


July 7, 2019  |  

Variations in 5S rDNAs in diploid and tetraploid offspring of red crucian carp × common carp.

The allotetraploid hybrid fish (4nAT) that was created in a previous study through an intergeneric cross between red crucian carp (Carassius auratus red var., ?) and common carp (Cyprinus carpio L., ?) provided an excellent platform to investigate the effect of hybridization and polyploidization on the evolution of 5S rDNA. The 5S rDNAs of paternal common carp were made up of a coding sequence (CDS) and a non-transcribed spacer (NTS) unit, and while the 5S rDNAs of maternal red crucian carp contained a CDS and a NTS unit, they also contained a variable number of interposed regions (IPRs). The CDSs of the 5S rDNAs in both parental fishes were conserved, while their NTS units seemed to have been subjected to rapid evolution.The diploid hybrid 2nF1 inherited all the types of 5S rDNAs in both progenitors and there were no signs of homeologous recombination in the 5S rDNAs of 2nF1 by sequencing of PCR products. We obtained two segments of 5S rDNA with a total length of 16,457 bp from allotetraploid offspring 4nAT through bacterial artificial chromosome (BAC) sequencing. Using this sequence together with the 5S rDNA sequences amplified from the genomic DNA of 4nAT, we deduced that the 5S rDNAs of 4nAT might be inherited from the maternal progenitor red crucian carp. Additionally, the IPRs in the 5S rDNAs of 4nAT contained A-repeats and TA-repeats, which was not the case for the IPRs in the 5S rDNAs of 2nF1. We also detected two signals of a 200-bp fragment of 5S rDNA in the chromosomes of parental progenitors and hybrid progenies by fluorescence in situ hybridization (FISH).We deduced that during the evolution of 5S rDNAs in different ploidy hybrid fishes, interlocus gene conversion events and tandem repeat insertion events might occurred in the process of polyploidization. This study provided new insights into the relationship among the evolution of 5S rDNAs, hybridization and polyploidization, which were significant in clarifying the genome evolution of polyploid fish.


July 7, 2019  |  

Heterogeneity of the Epstein-Barr virus major internal repeat reveals evolutionary mechanisms of EBV and a functional defect in the prototype EBV strain B95-8.

Epstein-Barr virus (EBV) is a ubiquitous pathogen of humans that can cause several types of lymphoma and carcinoma. Like other herpesviruses, EBV has diversified both through co-evolution with its host, and genetic exchange between virus strains. Sequence analysis of the EBV genome is unusually challenging, because of the large number and length of repeat regions within the virus. Here we describe the sequence assembly and analysis of the large internal repeat of EBV (IR1 or BamW repeats) from over 70 strains.Diversity of the latency protein EBNA-LP resides predominantly within the exons downstream of IR1. The integrity of the putative BWRF1 ORF is retained in over 80% of strains, and deletions truncating IR1 always spare BWRF1. Conserved regions include the IR1 latency promoter (Wp), and one zone upstream of and two within BWRF1.IR1 is heterogeneous in 70% of strains, and this heterogeneity arises from sequence exchange between strains as well as spontaneous mutation, with inter-strain recombination more common in tumour-derived viruses. This genetic exchange often incorporates regions of <1kb, and allelic gene conversion changes the frequency of small regions within the repeat, but not close to the flanks. These observations suggest that IR1 - and by extension EBV - diversifies through both recombination and breakpoint repair, while concerted evolution of IR1 is driven by gene conversion of small regions. Finally, the prototype EBV strain B95-8 contains four non-consensus variants within a single IR1 repeat unit, including a STOP codon in EBNA-LP. Repairing IR1 improves EBNA-LP levels and the quality of transformation by the B95-8 BAC.IMPORTANCE Epstein-Barr virus (EBV) infects the majority of the world population, but only causes illness in a small minority. Nevertheless, over 1% of cancers worldwide are attributable to EBV. Recent sequencing projects investigating virus diversity, to see if different strains have different disease impacts, have excluded regions of repeating sequence, as they are more technically challenging. Here we analyse the sequence of the largest repeat in EBV (IR1). We first characterised the variations in protein sequences encoded across IR1. In studying variations within the repeat of each strain, we identified a mutation in the main laboratory strain of EBV that impairs virus function, and suggest that tumour-associated viruses may be more likely to contain DNA mixed from two strains. Patterns of this mixing suggest that sequences can spread between strains (and also within the repeat) by copying sequence from another strain (or repeat unit) to repair DNA damage. Copyright © 2017 Ba abdullah et al.


July 7, 2019  |  

The unusual S locus of Leavenworthia is composed of two sets of paralogous loci.

The Leavenworthia self-incompatibility locus (S locus) consists of paralogs (Lal2, SCRL) of the canonical Brassicaceae S locus genes (SRK, SCR), and is situated in a genomic position that differs from the ancestral one in the Brassicaceae. Unexpectedly, in a small number of Leavenworthia alabamica plants examined, sequences closely resembling exon 1 of SRK have been found, but the function of these has remained unclear. BAC cloning and expression analyses were employed to characterize these SRK-like sequences. An SRK-positive Bacterial Artificial Chromosome clone was found to contain complete SRK and SCR sequences located close by one another in the derived genomic position of the Leavenworthia S locus, and in place of the more typical Lal2 and SCRL sequences. These sequences are expressed in stigmas and anthers, respectively, and crossing data show that the SRK/SCR haplotype is functional in self-incompatibility. Population surveys indicate that < 5% of Leavenworthia S loci possess such alleles. An ancestral translocation or recombination event involving SRK/SCR and Lal2/SCRL likely occurred, together with neofunctionalization of Lal2/SCRL, and both haplotype groups now function as Leavenworthia S locus alleles. These findings suggest that S locus alleles can have distinctly different evolutionary origins.© 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.


July 7, 2019  |  

Contributions of Zea mays subspecies mexicana haplotypes to modern maize.

Maize was domesticated from lowland teosinte (Zea mays ssp. parviglumis), but the contribution of highland teosinte (Zea mays ssp. mexicana, hereafter mexicana) to modern maize is not clear. Here, two genomes for Mo17 (a modern maize inbred) and mexicana are assembled using a meta-assembly strategy after sequencing of 10 lines derived from a maize-teosinte cross. Comparative analyses reveal a high level of diversity between Mo17, B73, and mexicana, including three Mb-size structural rearrangements. The maize spontaneous mutation rate is estimated to be 2.17?×?10-8 ~3.87?×?10-8 per site per generation with a nonrandom distribution across the genome. A higher deleterious mutation rate is observed in the pericentromeric regions, and might be caused by differences in recombination frequency. Over 10% of the maize genome shows evidence of introgression from the mexicana genome, suggesting that mexicana contributed to maize adaptation and improvement. Our data offer a rich resource for constructing the pan-genome of Zea mays and genetic improvement of modern maize varieties.


July 7, 2019  |  

Map-based cloning of the fertility restoration locus Rfm1 in cultivated barley (Hordeum vulgare)

Hybridization technology has proven valuable in enhancing yields in many crops, but was only recently adopted in the small grain cereals. Hybrid varieties in barley (Hordeum vulgare) rely on the cytoplasmic male sterility (CMS) system msm1 derived from Hordeum vulgare ssp. spontaneum. The major restorer gene described for the msm1 system is known as Rfm1 and maps to the top of chromosome 6H. To gain further insight into mechanisms underlying male fertility restoration in barley, we used a map-based cloning approach to identify the nuclear gene involved in the restoration mechanism of this hybridization system. Taking advantage of the available genomic resources in barley in combination with a custom-made non-gridded BAC library developed from a restorer line, we cloned and sequenced the Rfm1 restorer locus. The characterization and annotation of the nucleotide sequence for the Rfm1 restorer allele allowed for the identification of the candidate gene for Rfm1. The Rfm1 locus carries a tandem repeat of a gene encoding a pentatricopeptide repeat (PPR) protein. Surprisingly, Rfm1 belongs to the PLS-DYW subfamily of PPR genes known for their involvement in RNA editing in plants organelles, but that to date have not been identified as restorer genes.


July 7, 2019  |  

Disease onset in X-linked dystonia-parkinsonism correlates with expansion of a hexameric repeat within an SVA retrotransposon in TAF1.

X-linked dystonia-parkinsonism (XDP) is a neurodegenerative disease associated with an antisense insertion of a SINE-VNTR-Alu (SVA)-type retrotransposon within an intron ofTAF1This unique insertion coincides with six additional noncoding sequence changes inTAF1, the gene that encodes TATA-binding protein-associated factor-1, which appear to be inherited together as an identical haplotype in all reported cases. Here we examined the sequence of this SVA in XDP patients (n= 140) and detected polymorphic variation in the length of a hexanucleotide repeat domain, (CCCTCT)nThe number of repeats in these cases ranged from 35 to 52 and showed a highly significant inverse correlation with age at disease onset. Because other SVAs exhibit intrinsic promoter activity that depends in part on the hexameric domain, we assayed the transcriptional regulatory effects of varying hexameric lengths found in the unique XDP SVA retrotransposon using luciferase reporter constructs. When inserted sense or antisense to the luciferase reading frame, the XDP variants repressed or enhanced transcription, respectively, to an extent that appeared to vary with length of the hexamer. Further in silico analysis of this SVA sequence revealed multiple motifs predicted to form G-quadruplexes, with the greatest potential detected for the hexameric repeat domain. These data directly link sequence variation within the XDP-specific SVA sequence to phenotypic variability in clinical disease manifestation and provide insight into potential mechanisms by which this intronic retroelement may induce transcriptional interference inTAF1expression. Copyright © 2017 the Author(s). Published by PNAS.


July 7, 2019  |  

Two orangutan species have evolved different KIR alleles and haplotypes.

The immune and reproductive functions of human NK cells are regulated by interactions of the C1 and C2 epitopes of HLA-C with C1-specific and C2-specific lineage III killer cell Ig-like receptors (KIR). This rapidly evolving and diverse system of ligands and receptors is restricted to humans and great apes. In this context, the orangutan has particular relevance because it represents an evolutionary intermediate, one having the C1 epitope and corresponding KIR but lacking the C2 epitope. Through a combination of direct sequencing, KIR genotyping, and data mining from the Great Ape Genome Project, we characterized the KIR alleles and haplotypes for panels of 10 Bornean orangutans and 19 Sumatran orangutans. The orangutan KIR haplotypes have between 5 and 10 KIR genes. The seven orangutan lineage III KIR genes all locate to the centromeric region of the KIR locus, whereas their human counterparts also populate the telomeric region. One lineage III KIR gene is Bornean specific, one is Sumatran specific, and five are shared. Of 12 KIR gene-content haplotypes, 5 are Bornean specific, 5 are Sumatran specific, and 2 are shared. The haplotypes have different combinations of genes encoding activating and inhibitory C1 receptors that can be of higher or lower affinity. All haplotypes encode an inhibitory C1 receptor, but only some haplotypes encode an activating C1 receptor. Of 130 KIR alleles, 55 are Bornean specific, 65 are Sumatran specific, and 10 are shared. Copyright © 2017 by The American Association of Immunologists, Inc.


July 7, 2019  |  

Assembly and characterization of the MHC class I region of the Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis).

The Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis; YFP) is the sole freshwater subspecies of N. asiaeorientalis and is now critically endangered. Major histocompatibility complex (MHC) is a family of highly polymorphic genes that play an important immunological role in antigen presentation in the vertebrates. Currently, however, little is known about MHC region in the genome of the YFP, which hampers conservation genetics and evolutionary ecology study using MHC genes. In this work, a nucleotide sequence of 774,811 bp covering the YFP MHC class I region was obtained by screening a YFP bacterial artificial chromosome (BAC) library, followed by sequencing and assembly of positive BAC clones. A total of 45 genes were successfully annotated, of which four were MHC class I genes. There are high similarities among the four YFP MHC class I genes (>94 %). Divergence in the coding region of the four YFP MHC class I genes is mainly localized to exons 2 and 3, which encode the antigen-binding sites of MHC class I genes. Additionally, comparison of the MHC structure in YFP to those of cattle, sheep, and pig showed that MHC class I genes are located in genome regions with regard to the conserved genes, and the YFP contains the fewest MHC class I genes among these species. This is the first report characterizing a cetacean MHC class I region and describing its organization, which would be valuable for further investigation of adaptation in natural populations of the YFP and other cetaceans.


July 7, 2019  |  

Refinement of the canine CD1 locus topology and investigation of antibody binding to recombinant canine CD1 isoforms.

CD1 molecules are antigen-presenting glycoproteins primarily found on dendritic cells (DCs) responsible for lipid antigen presentation to CD1-restricted T cells. Despite their pivotal role in immunity, little is known about CD1 protein expression in dogs, notably due to lack of isoform-specific antibodies. The canine (Canis familiaris) CD1 locus was previously found to contain three functional CD1A genes: canCD1A2, canCD1A6, and canCD1A8, where two variants of canCD1A8, canCD1A8.1 and canCD1A8.2, were assumed to be allelic variants. However, we hypothesized that these rather represented two separate genes. Sequencing of three overlapping bacterial artificial chromosomes (BACs) spanning the entire canine CD1 locus revealed canCD1A8.2 and canCD1A8.1 to be located in tandem between canCD1A7 and canCD1C, and canCD1A8.1 was consequently renamed canCD1A9. Green fluorescent protein (GFP)-fused canine CD1 transcripts were recombinantly expressed in 293T cells. All proteins showed a highly positive GFP expression except for canine CD1d and a splice variant of canine CD1a8 lacking exon 3. Probing with a panel of anti-CD1 monoclonal antibodies (mAbs) showed that Ca13.9H11 and Ca9.AG5 only recognized canine CD1a8 and CD1a9 isoforms, and Fe1.5F4 mAb solely recognized canine CD1a6. Anti-CD1b mAbs recognized the canine CD1b protein, but also bound CD1a2, CD1a8, and CD1a9. Interestingly, Ca9.AG5 showed allele specificity based on a single nucleotide polymorphism (SNP) located at position 321. Our findings have refined the structure of the canine CD1 locus and available antibody specificity against canine CD1 proteins. These are important fundamentals for future investigation of the role of canine CD1 in lipid immunity.


July 7, 2019  |  

The genome analysis of Candidatus Burkholderia crenata reveals that secondary metabolism may be a key function of the Ardisia crenata leaf nodule symbiosis.

A majority of Ardisia species harbour Burkholderia sp. bacteria within specialized leaf nodules. The bacteria are transmitted hereditarily and have not yet been cultured outside of their host. Because the plants cannot develop beyond the seedling stage without their symbionts, the symbiosis is considered obligatory. We sequenced for the first time the genome of Candidatus Burkholderia crenata (Ca. B. crenata), the leaf nodule symbiont of Ardisia crenata. The genome of Ca. B. crenata is the smallest Burkholderia genome to date. It contains a large amount of insertion sequences and pseudogenes and displays features consistent with reductive genome evolution. The genome does not encode functions commonly associated with plant symbioses such as nitrogen fixation and plant hormone metabolism. However, we identified unique genes with a predicted role in secondary metabolism in the genome of Ca. B. crenata. Specifically, we provide evidence that the bacterial symbionts are responsible for the synthesis of compound FR900359, a cyclic depsipeptide with biomedical properties previously isolated from leaves of A.?crenata. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.


July 7, 2019  |  

The Cer-cqu gene cluster determines three key players in a ß-diketone synthase polyketide pathway synthesizing aliphatics in epicuticular waxes.

Aliphatic compounds on plant surfaces, called epicuticular waxes, are the first line of defense against pathogens and pests, contribute to reducing water loss and determine other important phenotypes. Aliphatics can form crystals affecting light refraction, resulting in a color change and allowing identification of mutants in their synthesis or transport. The present study discloses three such Eceriferum (cer) genes in barley – Cer-c, Cer-q and Cer-u – known to be tightly linked and functioning in a biochemical pathway forming dominating amounts of ß-diketone and hydroxy-ß-diketones plus some esterified alkan-2-ols. These aliphatics are present in many Triticeae as well as dicotyledons such as Eucalyptus and Dianthus. Recently developed genomic resources and mapping populations in barley defined these genes to a small region on chromosome arm 2HS. Exploiting Cer-c and -u potential functions pinpointed five candidates, of which three were missing in apparent cer-cqu triple mutants. Sequencing more than 50 independent mutants for each gene confirmed their identification. Cer-c is a chalcone synthase-like polyketide synthase, designated diketone synthase (DKS), Cer-q is a lipase/carboxyl transferase and Cer-u is a P450 enzyme. All were highly expressed in pertinent leaf sheath tissue of wild type. A physical map revealed the order Cer-c, Cer-u, Cer-q with the flanking genes 101kb apart, confirming they are a gene cluster, Cer-cqu. Homology-based modeling suggests that many of the mutant alleles affect overall protein structure or specific active site residues. The rich diversity of identified mutations will facilitate future studies of three key enzymes involved in synthesis of plant apoplast waxes. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.


July 7, 2019  |  

Filling in the gap of human chromosome 4: Single Molecule Real Time sequencing of macrosatellite repeats in the facioscapulohumeral muscular dystrophy locus.

A majority of facioscapulohumeral muscular dystrophy (FSHD) is caused by contraction of macrosatellite repeats called D4Z4 that are located in the subtelomeric region of human chromosome 4q35. Sequencing the FSHD locus has been technically challenging due to its long size and nearly identical nature of repeat elements. Here we report sequencing and partial assembly of a BAC clone carrying an entire FSHD locus by a single molecule real time (SMRT) sequencing technology which could produce long reads up to about 18 kb containing D4Z4 repeats. De novo assembly by Hierarchical Genome Assembly Process 1 (HGAP.1) yielded a contig of 41 kb containing all but a part of the most distal D4Z4 element. The validity of the sequence model was confirmed by an independent approach employing anchored multiple sequence alignment by Kalign using reads containing unique flanking sequences. Our data will provide a basis for further optimization of sequencing and assembly conditions of D4Z4.


July 7, 2019  |  

High quality maize centromere 10 sequence reveals evidence of frequent recombination events.

The ancestral centromeres of maize contain long stretches of the tandemly arranged CentC repeat. The abundance of tandem DNA repeats and centromeric retrotransposons (CR) has presented a significant challenge to completely assembling centromeres using traditional sequencing methods. Here, we report a nearly complete assembly of the 1.85 Mb maize centromere 10 from inbred B73 using PacBio technology and BACs from the reference genome project. The error rates estimated from overlapping BAC sequences are 7 × 10(-6) and 5 × 10(-5) for mismatches and indels, respectively. The number of gaps in the region covered by the reassembly was reduced from 140 in the reference genome to three. Three expressed genes are located between 92 and 477 kb from the inferred ancestral CentC cluster, which lies within the region of highest centromeric repeat density. The improved assembly increased the count of full-length CR from 5 to 55 and revealed a 22.7 kb segmental duplication that occurred approximately 121,000 years ago. Our analysis provides evidence of frequent recombination events in the form of partial retrotransposons, deletions within retrotransposons, chimeric retrotransposons, segmental duplications including higher order CentC repeats, a deleted CentC monomer, centromere-proximal inversions, and insertion of mitochondrial sequences. Double-strand DNA break (DSB) repair is the most plausible mechanism for these events and may be the major driver of centromere repeat evolution and diversity. In many cases examined here, DSB repair appears to be mediated by microhomology, suggesting that tandem repeats may have evolved to efficiently repair frequent DSBs in centromeres.


July 7, 2019  |  

BAC-pool sequencing and assembly of 19 Mb of the complex sugarcane genome.

Sequencing plant genomes are often challenging because of their complex architecture and high content of repetitive sequences. Sugarcane has one of the most complex genomes. It is highly polyploid, preserves intact homeologous chromosomes from its parental species and contains >55% repetitive sequences. Although bacterial artificial chromosome (BAC) libraries have emerged as an alternative for accessing the sugarcane genome, sequencing individual clones is laborious and expensive. Here, we present a strategy for sequencing and assembly reads produced from the DNA of pooled BAC clones. A set of 178 BAC clones, randomly sampled from the SP80-3280 sugarcane BAC library, was pooled and sequenced using the Illumina HiSeq2000 and PacBio platforms. A hybrid assembly strategy was used to generate 2,451 scaffolds comprising 19.2 MB of assembled genome sequence. Scaffolds of =20 Kb corresponded to 80% of the assembled sequences, and the full sequences of forty BACs were recovered in one or two contigs. Alignment of the BAC scaffolds with the chromosome sequences of sorghum showed a high degree of collinearity and gene order. The alignment of the BAC scaffolds to the 10 sorghum chromosomes suggests that the genome of the SP80-3280 sugarcane variety is ~19% contracted in relation to the sorghum genome. In conclusion, our data show that sequencing pools composed of high numbers of BAC clones may help to construct a reference scaffold map of the sugarcane genome.


Talk with an expert

If you have a question, need to check the status of an order, or are interested in purchasing an instrument, we're here to help.