First CLAS12 Measurement of Deeply Virtual Compton Scattering Beam-Spin Asymmetries in the Extended Valence Region.

Deeply virtual Compton scattering (DVCS) allows one to probe generalized parton distributions describing the 3D structure of the nucleon. We report the first measurement of the DVCS beam-spin asymmetry using the CLAS12 spectrometer with a 10.2 and 10.6 GeV electron beam scattering from unpolarized protons. The results greatly extend the Q^{2} and Bjorken-x phase space beyond the existing data in the valence region and provide 1600 new data points measured with unprecedented statistical uncertainty, setting new, tight constraints for future phenomenological studies.

Observation of Correlations between Spin and Transverse Momenta in Back-to-Back Dihadron Production at CLAS12.

We report the first measurements of deep inelastic scattering spin-dependent azimuthal asymmetries in back-to-back dihadron electroproduction in the deep inelastic scattering process. In this reaction, two hadrons are produced in opposite hemispheres along the z axis in the virtual photon-target nucleon center-of-mass frame, with the first hadron produced in the current-fragmentation region and the second in the target-fragmentation region. The data were taken with longitudinally polarized electron beams of 10.2 and 10.6 GeV incident on an unpolarized liquid-hydrogen target using the CLAS12 spectrometer at Jefferson Lab. Observed nonzero sinΔϕ modulations in ep→e^{'}pπ^{+}X events, where Δϕ is the difference of the azimuthal angles of the proton and pion in the virtual photon and target nucleon center-of-mass frame, indicate that correlations between the spin and transverse momenta of hadrons produced in the target- and current-fragmentation regions may be significant. The measured beam-spin asymmetries provide a first access in dihadron production to a previously unexplored leading-twist spin- and transverse-momentum-dependent fracture function. The fracture functions describe the hadronization of the target remnant after the hard scattering of a virtual photon off a quark in the target particle and provide a new avenue for studying nucleonic structure and hadronization.

First Measurement of Λ Electroproduction off Nuclei in the Current and Target Fragmentation Regions.

We report results of Λ hyperon production in semi-inclusive deep-inelastic scattering off deuterium, carbon, iron, and lead targets obtained with the CLAS detector and the Continuous Electron Beam Accelerator Facility 5.014 GeV electron beam. These results represent the first measurements of the Λ multiplicity ratio and transverse momentum broadening as a function of the energy fraction (z) in the current and target fragmentation regions. The multiplicity ratio exhibits a strong suppression at high z and an enhancement at low z. The measured transverse momentum broadening is an order of magnitude greater than that seen for light mesons. This indicates that the propagating entity interacts very strongly with the nuclear medium, which suggests that propagation of diquark configurations in the nuclear medium takes place at least part of the time, even at high z. The trends of these results are qualitatively described by the Giessen Boltzmann-Uehling-Uhlenbeck transport model, particularly for the multiplicity ratios. These observations will potentially open a new era of studies of the structure of the nucleon as well as of strange baryons.

Observation of Azimuth-Dependent Suppression of Hadron Pairs in Electron Scattering off Nuclei.

We present the first measurement of dihadron angular correlations in electron-nucleus scattering. The data were taken with the CLAS detector and a 5.0 GeV electron beam incident on deuterium, carbon, iron, and lead targets. Relative to deuterium, the nuclear yields of charged-pion pairs show a strong suppression for azimuthally opposite pairs, no suppression for azimuthally nearby pairs, and an enhancement of pairs with large invariant mass. These effects grow with increased nuclear size. The data are qualitatively described by the gibuu model, which suggests that hadrons form near the nuclear surface and undergo multiple scattering in nuclei.These results show that angular correlation studies can open a new way to elucidate how hadrons form and interact inside nuclei.

An extended anchored linkage map and virtual mapping for the American mink genome based on homology to human and dog.

In this report we present an extended linkage map of the American mink (Neovison vison) consisting of 157 microsatellite markers and comprising at least one linkage group for each of the autosomes. Each linkage group has been assigned to a chromosome and oriented by fluorescence in situ hybridization (FISH) and/or by means of human/dog/mink comparative homology. The average interval between markers is 8.5 cM and the linkage groups collectively span 1340 cM. In addition, 217 and 275 mink microsatellites have been placed on human and dog genomes, respectively. In conjunction with the existing comparative human/dog/mink data, these assignments represent useful virtual maps for the American mink genome. Comparison of the current human/dog assembled sequential map with the existing Zoo-FISH-based human/dog/mink maps helped to refine the human/dog/mink comparative map. Furthermore, comparison of the human and dog genome assemblies revealed a number of large synteny blocks, some of which are corroborated by data from the mink linkage map.

Novel avian leukosis virus-related endogenous proviruses from layer chickens: characterization and development of locus-specific assays.

During the course of evolution, vertebrate genomes have been invaded and colonized by retroviruses. In humans, for example, endogenous retroviruses (long terminal repeat elements) occupy roughly twice as much sequence space as essential genes. There are numerous reports in the literature implicating endogenous proviruses in the modulation of host physiology. The fact that many of these host-virus interactions take place in a proviral locus-specific manner speaks to the need for rapid assays for element profiling. This report deals with the identification of novel elements belonging to a family of endogenous retroviruses, designated ALVE, that reside in the genome of the chicken and that are closely related to exogenous avian leukosis viruses. The study of ALVE elements in the chicken genome serves as a model system for understanding the interplay between endogenous viruses and their vertebrate hosts in general, including humans. In this report, we present locus-specific, diagnostic PCR-based assays for 2 novel ALVE elements. In addition, we characterize the proviral structures and examine the genomic environments of both novel elements along with a previously described element known as ALVE-NSAC-3.

Discovery of an expanded set of avian leukosis subgroup E proviruses in chickens using Vermillion, a novel sequence capture and analysis pipeline [corrected].

Transposable elements (TEs), such as endogenous retroviruses (ERVs), are common in the genomes of vertebrates. ERVs result from retroviral infections of germ-line cells, and once integrated into host DNA they become part of the host's heritable genetic material. ERVs have been ascribed positive effects on host physiology such as the generation of novel, adaptive genetic variation and resistance to infection, as well as negative effects as agents of tumorigenesis and disease. The avian leukosis virus subgroup E family (ALVE) of endogenous viruses of chickens has been used as a model system for studying the effects of ERVs on host physiology, and approximately 30 distinct ALVE proviruses have been described in the Gallus gallus genome. In this report we describe the development of a software tool, which we call Vermillion, and the use of this tool in combination with targeted next-generation sequencing (NGS) to increase the number of known proviruses belonging to the ALVE family of ERVs in the chicken genome by 4-fold, including expanding the number of known ALVE elements on chromosome 1 (Gga1) from the current 9 to a total of 40. Although we focused on the discovery of ALVE elements in chickens, with appropriate selection of target sequences Vermillion can be used to develop profiles of other families of ERVs and TEs in chickens as well as in species other than the chicken.

Glucose Repression of Amylase Gene Expression in DROSOPHILA MELANOGASTER.

We have previously shown that dietary glucose can reduce amylase activity in both adults and larvae of Drosophila; this reduction in enzyme activity reflects a reduction in the quantity of amylase protein, rather than an inhibition of enzyme activity. Here, we report that we have now defined conditions in which the repressive effect of glucose can be greater than 100-fold. Moreover, this repression is partially counteracted by the addition of exogenous cyclic AMP. We also show that there is a direct correlation between changes in amylase activity and changes in the amount of translatable mRNA as assayed in microinjected Xenopus oocytes. This means that the glucose repression is occurring at a pretranslational stage.

The Interaction of Genetic and Environmental Factors in the Control of Amylase Gene Expression in DROSOPHILA MELANOGASTER.

A number of previous studies have established that amylase activity can vary between Drosophila strains which are maintained under identical laboratory conditions. In addition, we have recently shown that all strains examined so far are subject to glucose repression of amylase activity. In this study, we show that the degree of glucose repression can vary between strains. Moreover, the glucose repression effect is much more pronounced in larvae than in adult flies. Our results lead to the conclusion that the strain-specific differences in activity and the dietary effects are not independent phenomena. These results have implications for the interpretation of many studies on amylase activity variation, including those experiments which have been designed to link amylase activity variations with fitness differences in nature. A question that naturally arises concerns the molecular basis for these strain-specific variations in the degree of glucose repression of this eukaryotic gene.

A short 5'-flanking region mediates glucose repression of amylase gene expression in Drosophila melanogaster.

Expression of the alpha-amylase gene is highly repressed by dietary glucose in Drosophila melanogaster larvae. Here, we show that glucose repression is controlled by DNA sequences that are located upstream of the transcribed region. Recombinant gene constructions, in which the amylase promoter sequences were fused with the transcribed region of the Adh gene, were expressed in transgenic Drosophila larvae. The expression of ADH from the recombinant gene was shown to be subject to glucose repression. The function of potential regulatory cis-acting elements within the glucose responsive upstream region was examined by deletion analysis and by site-directed mutagenesis, coupled with expression assays in transformed larvae. The upstream deletion analysis showed that essential elements, both for overall activity and for glucose repression of the amylase gene, are located within a 109-bp region upstream of the transcription start site. Site-directed mutagenesis of these upstream sequences showed that the TATA motif, at position -31, and a novel 36-bp element, at position -109, were necessary for full activity of the amylase promoter. None of the introduced mutations resulted in loss of glucose responsiveness. These results indicate that glucose repression, in Drosophila, is mediated by transcriptional mechanisms that involve multiple, functionally redundant DNA elements.

Prediction of heterosis from DNA fingerprints in chickens.

To assess the value of DNA fingerprints for the prediction of heterosis in chickens, retrospective analyses of data from three crossbreeding experiments and DNA fingerprints (DEP) of parental strains were conducted using two minisatellite and one middle-repetitive DNA probes. DEP bands were assessed on pooled DNA samples of 10-15 individuals per parental genetic group. The number of DEP bands evaluated in the experiments ranged from 81 to 139. The probes varied in their predictive value, but predictability of heterosis generally increased with multiple probes. Highly significant correlations (0.68-0.87) between band sharing ratios (SH) and heterosis were found in 25 crosses of White Leghorns in the first egg production cycle for age at sexual maturity, egg production, and mature body weight: traits with heterosis of 10% or more of the means. Regressions on SH explained 78.4% of the variation in heterosis in age at sexual maturity, 60.2% in egg production and 46.4% in mature body weight. For "broiler" traits with heterosis of < 1%, none of the correlations, based on 13 crosses, were significant. It was concluded that multilocus probe DFP of pooled DNA samples show promise as predictors of heterosis.

Cloning and expression of a chicken alpha-amylase gene.

We have isolated and sequenced a genomic clone for a pancreatic alpha-amylase gene (amy) of the chicken (Gallus gallus). The gene is interrupted by nine introns, spans over 4 kb, and encodes a protein (AMY) of 512 aa that is 83% identical to the human pancreatic alpha-amylase enzyme. Southern blot analysis of chicken DNA revealed two distinct pancreatic amy loci. In addition, we have generated a cDNA from chicken pancreatic RNA corresponding to the coding sequence of the genomic clone. The cDNA was inserted into a yeast expression vector, and the resulting construct used to transform Saccharomyces cerevisiae cells. Transformed yeast cells synthesized and secreted active AMY enzyme, and the gel migration pattern of the alpha-amylase produced by the yeast cells was identical to that of the native chicken enzyme.

A comparative map of bovine chromosome 19 based on a combination of mapping on a bacterial artificial chromosome scaffold map, a whole genome radiation hybrid panel and the human draft sequence.

We have constructed a medium density physical map of bovine chromosome 19 using a combination of mapping loci on both a bovine bacterial artificial chromosome (BAC) scaffold map and a whole genome radiation hybrid (WGRH) panel. The resulting map contains 70 loci spanning the length of bovine chromosome 19. Three contiguous groups of BACs were identified on the basis of multiple loci mapping to individual BAC clones. Bovine chromosome 19 was found in this study to be comprised almost entirely from regions of human chromosome 17, with a small region putatively assigned to human chromosome 10. Fourteen breakpoints between the bovine and human chromosomes were detected, with a possibility of five more based on ordering of the WGRH map.

A quantitative competitive reverse transcription-polymerase chain reaction technique to measure porcine interferon-gamma.

In swine, as in other mammals, interferon-gamma (IFN-gamma) plays an important role in the regulation of the immune response. The objective of this work was to develop a sensitive, quantitative competitive reverse transcription-polymerase chain reaction (qcRT-PCR) assay, using an internal cRNA standard, to monitor the expression of porcine IFN-gamma gene. We applied this qcRT-PCR assay to quantify the expression of IFN-gamma transcripts in peripheral blood mononuclear cells (PBMC) incubated for different times with or without concanavalin A (ConA). Results showed that the expression of the IFN-gamma transcripts in PBMC occurred within the first 2 h after mitogenic stimulation and reached a peak after 24 h of incubation with ConA. The qcRT-PCR technique provides an efficient and sensitive tool for studying the expression of porcine IFN-gamma gene under a variety of experimental conditions. This will help to understand the regulation of T cell-mediated immune responses by IFN-gamma at the gene level.

Assessment of positional candidate genes myf5 and igf1 for growth on bovine chromosome 5 in commercial lines of Bos taurus.

Quantitative trait loci for growth traits in beef cattle have been previously reported and fine-mapped in three chromosomal regions of 0 to 30 cM, 55 to 70 cM, and 70 to 80 cM of bovine chromosome 5. In this study, we further examined the association between gene-specific single nucleotide polymorphisms (SNP) of two positional candidate genes, bovine myogenic factor 5 (myf5) and insulin-like growth factor-1 (igf1), in the QTL regions and the birth weight (BWT), preweaning average daily gain (PWADG), and average daily gain on feed (ADGF) in commercial lines of Bos taurus. The QTL regions for the growth traits identified using a haplotype association analysis, which included the gene-specific SNP markers for both genes in this study, were in agreement with previous studies. The gene-specific SNP marker association analysis indicated that the SNP in myf5 had a significant additive effect on PWADG in the M1 line of Beefbooster Inc. (P < 0.10), and a significant additive effect (P < 0.05) and a significant dominance effect (P < 0.10) on ADGF in the M3 line of Beefbooster Inc. When the data from the two commercial lines were pooled, the SNP in myf5 showed a significant association with PWADG (P < 0.10) and with ADGF (P < 0.05). The association between the SNP and BWT, however, did not reach a significance level in the M1 line, the M3 line, or across the lines. For igf1, no significant association between the SNP and the growth traits was detected in either the M1 line or the M3 line, whereas there was only a significant dominance effect (P < 0.10) on BWT detected for the SNP in igfl when the data from the two commercial lines were pooled. These results suggest that myf5 is a strong candidate gene that influences PWADG and ADGF in beef cattle. The SNP of igf1 may not be a causative or close to the causative mutation that affects the three growth traits in the populations of beef cattle examined in this study. Other SNP of igf1 and myf5 or other genes in their respective chromosomal regions, however, should also be studied.

The identification of common haplotypes on bovine chromosome 5 within commercial lines of Bos taurus and their associations with growth traits.

The cosegregation between a genetic marker and the QTL in a well-designed mapping population is the basis for successful QTL mapping. Linkage disequilibria are, however, also expected among individuals that descended from the same breeding line, and some common haplotypes should carry on and segregate among individuals of the line. These identical by descent haplotypes make it possible to identify and locate the QTL segregating in the line. We report the identification of common haplotypes within commercial lines of Bos taurus and their associations with growth traits. One hundred and seventy six male calves and their 12 sires (9 to 30 male calves of each sire) of the Beefbooster, Inc., M1 line selected for maternal traits over 30 yr were genotyped using 16 microsatellite markers chosen from bovine chromosome 5 for the initial haplotype and growth association analysis. In order to verify the results from the M1 line, another 170 male calves and their 14 sires from the Beefbooster M3 line were genotyped using nine microsatellite markers chosen from bovine chromosome 5. The alleles of each male calf contributed by the sire and by the dam were identified, and haplotypes in the M1 line were established along 93% of bovine chromosome 5. The haplotypes in the M3 line were established along the chosen regions of bovine chromosome 5. Regression analysis detected 10 haplotypes in three chromosomal regions (0 to 30 cM, 55 to 70 cM, and 70 to 80 cM) that showed significant associations with birth weight, preweaning average daily gain, and average daily gain on feed in M1 line and 9 haplotypes associated with the growth traits in the same chromosomal regions in the M3 line at the comparisonwise threshold level. On average, the 19 haplotypes have an effect of 0.68 SD on the growth traits, ranging from 0.41 SD to 1.02 SD The results provide a useful reference for further positional candidate gene research and marker-assisted selection.

Identification and fine mapping of quantitative trait loci for backfat on bovine chromosomes 2, 5, 6, 19, 21, and 23 in a commercial line of Bos taurus.

Backfat thickness is one of the major quantitative traits that affects carcass quality in beef cattle. In this study, we identified and fine-mapped QTL for backfat EBV on bovine chromosomes 2, 5, 6, 19, 21, and 23 using an identical-by-descent haplotype-sharing analysis in a commercial line of Bos taurus. Eleven haplotypes were found to have significant associations with backfat EBV at the comparison-wise P-value threshold, and one at the chromosome-wise P-value threshold on bovine chromosomes 5, 6, 19, 21, and 23. On average, the 12 significant haplotypes had an effect of 0.62 SD on backfat EBV, ranging from 0.38 SD to 1.33 SD. The 12 significant haplotypes spanned nine chromosomal regions, one on chromosome 5 (65.4 to 70.0 cM), three on 6 (8.2 to 11.8 cM, 63.6 to 68.1 cM, and 81.5 to 83.0 cM), three on 19 (4.8 to 15.9 cM, 39.4 to 46.5 cM, and 65.7 to 99.5 cM), one on 21 (46.1 to 53.1 cM), and one on 23 (45.1 to 50.9 cM). Among the nine chromosomal regions, six were new QTL regions and three showed remarkable agreement with QTL regions that were previously reported. Eight of the nine QTL regions were localized to less than or close to 10 cM in genetic distance. The results provide a useful reference for further positional candidate gene research and marker-assisted selection for backfat.

Fine mapping of quantitative trait loci and assessment of positional candidate genes for backfat on bovine chromosome 14 in a commercial line of Bos taurus.

Backfat thickness is one of the major quantitative traits that affect carcass quality in beef cattle. In this study, we have fine mapped a QTL for backfat EBV on bovine chromosome 14, using an identical-by-descent haplotype-sharing analysis, in a commercial line of Bos taurus. We also examined the association between gene-specific single nucleotide polymorphism (SNP) markers of the genes diacylgcerol acyltransferase 1 (DGAT1) and thyroglobulin (TG) and the backfat EBV. The results indicate that the QTL region for backfat identified on chromosome 14 is in agreement with previous studies. However, neither of the two polymorphisms of candidate genes tested, DGAT1 nor TG, showed a significant (P > 0.10) association with the backfat EBV in the cattle populations examined. However, a strong association (P = 0.0058) was detected between a microsatellite marker (CSSM66) lying approximately mid-way between the two candidate genes and the backfat EBV. These results suggest that other SNP of DGAT1, TG, or other gene(s) in the chromosomal region should be examined to test whether they have a significant effect on lipid metabolism.

Different measures of energetic efficiency and their phenotypic relationships with growth, feed intake, and ultrasound and carcass merit in hybrid cattle.

Residual feed intake (RFI) has been proposed as an index for determining beef cattle energetic efficiency. Although the relationship of RFI with feed conversion ratio (FCR) is well established, little is known about how RFI compares to other measures of efficiency. This study examined the phenotypic relationships among different measures of energetic efficiency with growth, feed intake, and ultrasound and carcass merit of hybrid cattle (n = 150). Dry matter intake, ME intake (MEI), ADG, metabolic weight (MWT), and FCR during the test averaged 10.29 kg/d (SD = 1.62), 1,185.45 kJ/(kg0.75 x d) (SD = 114.69), 1.42 kg/d (SD = 0.25), 86.67 kg0.75 (SD = 10.21), and 7.27 kg of DM/kg of gain (SD = 1.00), respectively. Residual feed intake averaged 0.00 kg/d and ranged from -2.25 kg/d (most efficient) to 2.61 kg/d (least efficient). Dry matter intake (r = 0.75), MEI (r = 0.83), and FCR (r = 0.62) were correlated with RFI (P < 0.001) and were higher for animals with high (>0.5 SD) RFI vs. those with medium (+/-0.5 SD) or low (<0.5 SD) RFI (P < 0.001). Partial efficiency of growth (PEG; energetic efficiency for ADG) was correlated with RFI (r = -0.89, P < 0.001) and was lower (P < 0.001) for high- vs. medium- or low-RFI animals. However, RFI was not related to ADG (r = -0.03), MWT (r = -0.02), relative growth rate (RGR; growth relative to instantaneous body size; r = -0.04), or Kleiber ratio (KR; ADG per unit of MWT; r = -0.004). Also, DMI was correlated (P < 0.01) with ADG (r = 0.66), MWT (r = 0.49), FCR (r = 0.49), PEG (r = -0.52), RGR (r = 0.18), and KR (r = 0.36). Additionally, FCR was correlated (P < 0.001) with ADG (r = -0.63), PEG (r = -0.83), RGR (r = -0.75), and KR (r = -0.73), but not with MWT (r = 0.07). Correlations of measures of efficiency with ultrasound or carcass traits generally were not different from zero except for correlations of RFI, FCR, and PEG, respectively, with backfat gain (r = 0.30, 0.20, and -0.30), ultrasound backfat (r = 0.19, 0.21, and -0.25), grade fat (r = 0.25, 0.19, and -0.27), lean meat yield (r = -0.22, -0.18, and 0.24), and yield grade (r = 0.28, 0.24, and -0.25). These phenotypic relationships indicate that, compared with other measures of energetic efficiency, RFI should have a greater potential to improve overall production efficiency and PEG above maintenance, and lead to minimal correlated changes in carcass merit without altering the growth and body size of different animals.

Cloning and characterization of a bovine genomic fragment homologous to epidermal growth factor genes.

Epidermal growth factor (EGF) is a potent mitogen for a variety of cell types. The 53-amino acid mature EGF protein is encoded by sequences in exons 20 and 21 of a gene spanning over 110 kb. In this study, we report the cloning and characterization of 7.5 kb of bovine genomic sequence homologous to exon 19 through 21 from EGF genes from other mammalian species. The cloned gene fragment had an unusual sequence composition in the form of an in-frame TGA codon in the coding sequence. The sequence was expressed at low levels in kidney tissue and the corresponding cDNA contained the TGA codon. The level of similarity between the bovine exonic sequence and the human, porcine, murine, feline, and canine corresponding sequences varied from 64% to 73%; however, when only sequences encoding the mature EGF protein were compared, the level of similarity between the bovine sequence and the sequence from these species was 59% to 66%. The sequence similarity of the deduced mature protein was lower (34% to 39%) than the sequence similarity of the deduced propeptide. Although the cloned sequences could originate from a bovine EGF pseudogene, the possibility exists that they originate from the functional EGF gene. An as yet unidentified mechanism to by-pass the stop codon would allow the synthesis of a functional EGF protein. Alternatively, the cloned sequence could originate from an EGF-like gene.