Generation of Lamprey Monoclonal Antibodies (Lampribodies) Using the Phage Display System.

The variable lymphocyte receptors (VLRs) consist of leucine rich repeats (LRRs) and comprise the humoral antibodies produced by lampreys and hagfishes. The diversity of the molecules is generated by stepwise genomic rearrangements of LRR cassettes dispersed throughout the VLRB locus. Previously, target-specific monovalent VLRB antibodies were isolated from sea lamprey larvae after immunization with model antigens. Further, the cloned VLR cDNAs from activated lamprey leukocytes were transfected into human cell lines or yeast to select best binders. Here, we expand on the overall utility of the VLRB technology by introducing it into a filamentous phage display system. We first tested the efficacy of isolating phage into which known VLRB molecules were cloned after a series of dilutions. These experiments showed that targeted VLRB clones could easily be recovered even after extensive dilutions (1 to 109). We further utilized the system to isolate target-specific "lampribodies" from phage display libraries from immunized animals and observed an amplification of binders with relative high affinities by competitive binding. The lampribodies can be individually purified and ostensibly utilized for applications for which conventional monoclonal antibodies are employed

Molecular cytogenetic differentiation of paralogs of Hox paralogs in duplicated and re-diploidized genome of the North American paddlefish (Polyodon spathula).

BackgroundAcipenseriformes is a basal lineage of ray-finned fishes and comprise 27 extant species of sturgeons and paddlefishes. They are characterized by several specific genomic features as broad ploidy variation, high chromosome numbers, presence of numerous microchromosomes and propensity to interspecific hybridization. The presumed palaeotetraploidy of the American paddlefish was recently validated by molecular phylogeny and Hox genes analyses. A whole genome duplication in the paddlefish lineage was estimated at approximately 42 Mya and was found to be independent from several genome duplications evidenced in its sister lineage, i.e. sturgeons. We tested the ploidy status of available chromosomal markers after the expected rediploidization. Further we tested, whether paralogs of Hox gene clusters originated from this paddlefish specific genome duplication are cytogenetically distinguishable.ResultsWe found that both paralogs HoxA alpha and beta were distinguishable without any overlapping of the hybridization signal - each on one pair of large metacentric chromosomes. Of the HoxD, only the beta paralog was unequivocally identified, whereas the alpha paralog did not work and yielded only an inconclusive diffuse signal. Chromosomal markers on three diverse ploidy levels reflecting different stages of rediploidization were identified: quadruplets retaining their ancestral tetraploid condition, semi-quadruplets still reflecting the ancestral tetraploidy with clear signs of advanced rediploidization, doublets were diploidized with ancestral tetraploidy already blurred. Also some of the available microsatellite data exhibited diploid allelic band patterns at their loci whereas another locus showed more than two alleles.ConclusionsOur exhaustive staining of paddlefish chromosomes combined with cytogenetic mapping of ribosomal genes and Hox paralogs and with microsatellite data, brings a closer look at results of the process of rediploidization in the course of paddlefish genome evolution. We show a partial rediploidization represented by a complex mosaic structure comparable with segmental paleotetraploidy revealed in sturgeons (Acipenseridae). Sturgeons and paddlefishes with their high propensity for whole genome duplication thus offer suitable animal model systems to further explore evolutionary processes that were shaping the early evolution of all vertebrates

Sequence and organization of coelacanth neurohypophysial hormone genes: Evolutionary history of the vertebrate neurohypophysial hormone gene locus

<p>Abstract</p> <p>Background</p> <p>The mammalian neurohypophysial hormones, vasopressin and oxytocin are involved in osmoregulation and uterine smooth muscle contraction respectively. All jawed vertebrates contain at least one homolog each of vasopressin and oxytocin whereas jawless vertebrates contain a single neurohypophysial hormone called vasotocin. The vasopressin homolog in non-mammalian vertebrates is vasotocin; and the oxytocin homolog is mesotocin in non-eutherian tetrapods, mesotocin and [Phe²]mesotocin in lungfishes, and isotocin in ray-finned fishes. The genes encoding vasopressin and oxytocin genes are closely linked in the human and rodent genomes in a tail-to-tail orientation. In contrast, their pufferfish homologs (vasotocin and isotocin) are located on the same strand of DNA with isotocin gene located upstream of vasotocin gene separated by five genes, suggesting that this locus has experienced rearrangements in either mammalian or ray-finned fish lineage, or in both lineages. The coelacanths occupy a unique phylogenetic position close to the divergence of the mammalian and ray-finned fish lineages.</p> <p>Results</p> <p>We have sequenced a coelacanth (<it>Latimeria menadoensis</it>) BAC clone encompassing the neurohypophysial hormone genes and investigated the evolutionary history of the vertebrate neurohypophysial hormone gene locus within a comparative genomics framework. The coelacanth contains vasotocin and mesotocin genes like non-mammalian tetrapods. The coelacanth genes are present on the same strand of DNA with no intervening genes, with the vasotocin gene located upstream of the mesotocin gene. Nucleotide sequences of the second exons of the two genes are under purifying selection implying a regulatory function. We have also analyzed the neurohypophysial hormone gene locus in the genomes of opossum, chicken and <it>Xenopus tropicalis</it>. The opossum contains two tandem copies of vasopressin and mesotocin genes. The vasotocin and mesotocin genes in chicken and <it>Xenopus</it>, and the vasopressin and mesotocin genes in opossum are linked tail-to-head similar to their orthologs in coelacanth and unlike their homologs in human and rodents.</p> <p>Conclusion</p> <p>Our results indicate that the neurohypophysial hormone gene locus has experienced independent rearrangements in both placental mammals and teleost fishes. The coelacanth genome appears to be more stable than mammalian and teleost fish genomes. As such, it serves as a valuable outgroup for studying the evolution of mammalian and teleost fish genomes.</p

Numerical Modeling of Dynamic Loss in HTS-Coated Conductors Under Perpendicular Magnetic Fields

© 2017 IEEE. High-Tc superconducting (HTS)-coated conductors are a promising option for the next-generation power devices. However, their thin-film geometry incurs dynamic loss when exposed to a perpendicular external ac magnetic field, which is difficult to predicate and estimate. In this paper, we propose and verify a numerical simulation model to predict the dynamic loss in HTS-thin-coated conductors by taking into account their Jc-B dependence and I-V characteristics. The model has been tested on a SuperPower YBCO-coated conductor, and we observed a linear increase of dynamic loss along the increasing field amplitude after the threshold field. Our simulation results agree closely with experimental measurements as well as an analytical model. Furthermore, the model can predict the nonlinear increase of dynamic loss at high current, while the analytical model deviates from the measurement results and still shows a linear correlation between the dynamic loss and the external magnetic field. In addition, we have used this model to simulate the distributions of magnetic field and current density when dynamic loss occurs. Results clearly show the flux traversing the coated conductor, which causes dynamic loss. These distributions have also been used to analyze the change of dynamic loss when either the transport current or the magnetic field increase individually, while the other factor remains constant. The simulation analysis on dynamic loss is done for the first time in this paper, and our results clearly demonstrate how dynamic loss changes as well as its dependence on transport current and magnetic field. © 2018 IEEE. Personal use of this material is permitted. Permission from IEEE must be obtained for all other uses, in any current or future media, including reprinting/republishing this material for advertising or promotional purposes, creating new collective works, for resale or redistribution to servers or lists, or reuse of any copyrighted component of this work in other works

Discovery of large genomic inversions using long range information.

BackgroundAlthough many algorithms are now available that aim to characterize different classes of structural variation, discovery of balanced rearrangements such as inversions remains an open problem. This is mainly due to the fact that breakpoints of such events typically lie within segmental duplications or common repeats, which reduces the mappability of short reads. The algorithms developed within the 1000 Genomes Project to identify inversions are limited to relatively short inversions, and there are currently no available algorithms to discover large inversions using high throughput sequencing technologies.ResultsHere we propose a novel algorithm, VALOR, to discover large inversions using new sequencing methods that provide long range information such as 10X Genomics linked-read sequencing, pooled clone sequencing, or other similar technologies that we commonly refer to as long range sequencing. We demonstrate the utility of VALOR using both pooled clone sequencing and 10X Genomics linked-read sequencing generated from the genome of an individual from the HapMap project (NA12878). We also provide a comprehensive comparison of VALOR against several state-of-the-art structural variation discovery algorithms that use whole genome shotgun sequencing data.ConclusionsIn this paper, we show that VALOR is able to accurately discover all previously identified and experimentally validated large inversions in the same genome with a low false discovery rate. Using VALOR, we also predicted a novel inversion, which we validated using fluorescent in situ hybridization. VALOR is available at https://github.com/BilkentCompGen/VALOR

Sex Chromosome Evolution in Amniotes: Applications for Bacterial Artificial Chromosome Libraries

Variability among sex chromosome pairs in amniotes denotes a dynamic history. Since amniotes diverged from a common ancestor, their sex chromosome pairs and, more broadly, sex-determining mechanisms have changed reversibly and frequently. These changes have been studied and characterized through the use of many tools and experimental approaches but perhaps most effectively through applications for bacterial artificial chromosome (BAC) libraries. Individual BAC clones carry 100–200 kb of sequence from one individual of a target species that can be isolated by screening, mapped onto karyotypes, and sequenced. With these techniques, researchers have identified differences and similarities in sex chromosome content and organization across amniotes and have addressed hypotheses regarding the frequency and direction of past changes. Here, we review studies of sex chromosome evolution in amniotes and the ways in which the field of research has been affected by the advent of BAC libraries

Evolution of Otx paralogue usages in early patterning of the vertebrate head

AbstractTo assess evolutional changes in the expression pattern of Otx paralogues, expression analyses were undertaken in fugu, bichir, skate and lamprey. Together with those in model vertebrates, the comparison suggested that a gnathostome ancestor would have utilized all of Otx1, Otx2 and Otx5 paralogues in organizer and anterior mesendoderm for head development. In this animal, Otx1 and Otx2 would have also functioned in specification of the anterior neuroectoderm at presomite stage and subsequent development of forebrain/midbrain at somite stage, while Otx5 expression would have already been specialized in epiphysis and eyes. Otx1 and Otx2 functions in anterior neuroectoderm and brain of the gnathostome ancestor would have been differentially maintained by Otx1 in a basal actinopterygian and by Otx2 in a basal sarcopterygian. Otx5 expression in head organizer and anterior mesendoderm seems to have been lost in the teleost lineage after divergence of bichir, and also from the amniotes after divergence of amphibians as independent events. Otx1 expression was lost from the organizer in the tetrapod lineage. In contrast, in a teleost ancestor prior to whole genome duplication, Otx1 and Otx2 would have both been expressed in the dorsal margin of blastoderm, embryonic shield, anterior mesendoderm, anterior neuroectoderm and forebrain/midbrain, at respective stages of head development. Subsequent whole genome duplication and the following genome changes would have caused different Otx paralogue usages in each teleost lineage. Lampreys also have three Otx paralogues; their sequences are highly diverged from gnathostome cognates, but their expression pattern is well related to those of skate Otx cognates

The dynamic resistance of YBCO coated conductor wire: Effect of DC current magnitude and applied field orientation

Dynamic resistance, which occurs when a HTS coated conductor carries a DC current under an AC magnetic field, can have critical implications for the design of HTS machines. Here, we report measurements of dynamic resistance in a commercially available SuperPower 4 mm-wide YBCO coated conductor, carrying a DC current under an applied AC magnetic field of arbitrary orientation. The reduced DC current, I t/I c0, ranged from 0.01 to 0.9, where I t is the DC current level and I c0 is the self-field critical current of the conductor. The field angle (the angle between the magnetic field and the normal vector of the conductor wide-face) was varied between 0° and 90° at intervals of 10°. We show that the effective width of the conductor under study is ∼12% less than the physical wire width, and we attribute this difference to edge damage of the wire during or after manufacture. We then examine the measured dynamic resistance of this wire under perpendicular applied fields at very low DC current levels. In this regime we find that the threshold field, B th, of the conductor is well described by the nonlinear equation of Mikitik and Brandt. However, this model consistently underestimates the threshold field at higher current levels. As such, the dynamic resistance in a coated conductor under perpendicular magnetic fields is best described using two different equations for each of the low and high DC current regimes, respectively. At low DC currents where I t/I c0 ≤ 0.1, the nonlinear relationship of Mikitik and Brandt provides the closest agreement with experimental data. However, in the higher current regime where I t/I c0 ≥ 0.2, closer agreement is obtained using a simple linear expression which assumes a current-independent penetration field. We further show that for the conductor studied here, the measured dynamic resistance at different field angles is dominated by the perpendicular magnetic field component, with negligible contribution from the parallel component. Our findings now enable the dynamic resistance of a single conductor to be analytically determined for a very wide range of DC currents and at all applied field angles. This is the Accepted Manuscript version of an article accepted for publication in 'Superconductor Science and Technology'. IOP Publishing Ltd is not responsible for any errors or omissions in this version of the manuscript or any version derived from it. The Version of Record is available online at https://doi.org/10.1088/1361-6668/aaa49e