RESUMO
Haematobia irritans, commonly known as the horn fly, is a globally distributed blood-feeding pest of cattle that is responsible for significant economic losses to cattle producers. Chemical insecticides are the primary means for controlling this pest but problems with insecticide resistance have become common in the horn fly. To provide a foundation for identification of genomic loci for insecticide resistance and for discovery of new control technology, we report the sequencing, assembly, and annotation of the horn fly genome. The assembled genome is 1.14 Gb, comprising 76,616 scaffolds with N50 scaffold length of 23 Kb. Using RNA-Seq data, we have predicted 34,413 gene models of which 19,185 have been assigned functional annotations. Comparative genomics analysis with the Dipteran flies Musca domestica L., Drosophila melanogaster, and Lucilia cuprina, show that the horn fly is most closely related to M. domestica, sharing 8,748 orthologous clusters followed by D. melanogaster and L. cuprina, sharing 7,582 and 7,490 orthologous clusters respectively. We also identified a gene locus for the sodium channel protein in which mutations have been previously reported that confers target site resistance to the most common class of pesticides used in fly control. Additionally, we identified 276 genomic loci encoding members of metabolic enzyme gene families such as cytochrome P450s, esterases and glutathione S-transferases, and several genes orthologous to sex determination pathway genes in other Dipteran species.
Assuntos
Genes de Insetos , Muscidae/genética , Muscidae/metabolismo , Processos de Determinação Sexual/genética , Sequenciamento Completo do Genoma , Animais , Análise por Conglomerados , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Ontologia Genética , Inativação Metabólica/genética , Resistência a Inseticidas/genética , Masculino , Modelos Genéticos , Anotação de Sequência Molecular , Família Multigênica , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Canais de Sódio/genética , Canais de Sódio/metabolismoRESUMO
Alternative splicing and alternative polyadenylation (APA) of pre-mRNAs greatly contribute to transcriptome diversity, coding capacity of a genome and gene regulatory mechanisms in eukaryotes. Second-generation sequencing technologies have been extensively used to analyse transcriptomes. However, a major limitation of short-read data is that it is difficult to accurately predict full-length splice isoforms. Here we sequenced the sorghum transcriptome using Pacific Biosciences single-molecule real-time long-read isoform sequencing and developed a pipeline called TAPIS (Transcriptome Analysis Pipeline for Isoform Sequencing) to identify full-length splice isoforms and APA sites. Our analysis reveals transcriptome-wide full-length isoforms at an unprecedented scale with over 11,000 novel splice isoforms. Additionally, we uncover APA of â¼11,000 expressed genes and more than 2,100 novel genes. These results greatly enhance sorghum gene annotations and aid in studying gene regulation in this important bioenergy crop. The TAPIS pipeline will serve as a useful tool to analyse Iso-Seq data from any organism.
Assuntos
Perfilação da Expressão Gênica/métodos , Proteínas de Plantas/metabolismo , Sorghum/genética , Sorghum/metabolismo , Transcriptoma , Regulação da Expressão Gênica de Plantas , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Proteínas de Plantas/genética , Reação em Cadeia da Polimerase , Isoformas de Proteínas , Splicing de RNA , RNA de Plantas/genética , RNA não Traduzido , Análise de Sequência de RNARESUMO
BACKGROUND: Polyclonal serum consists of vast collections of antibodies, products of differentiated B-cells. The spectrum of antibody specificities is dynamic and varies with age, physiology, and exposure to pathological insults. The complete repertoire of antibody specificities in blood, the IgOme, is therefore an extraordinarily rich source of information-a molecular record of previous encounters as well as a status report of current immune activity. The ability to profile antibody specificities of polyclonal serum at exceptionally high resolution has been an important and serious challenge which can now be overcome. METHODOLOGY/PRINCIPAL FINDINGS: Here we illustrate the application of Deep Panning, a method that combines the flexibility of combinatorial phage display of random peptides with the power of high-throughput deep sequencing. Deep Panning is first applied to evaluate the quality and diversity of naïve random peptide libraries. The production of very large data sets, hundreds of thousands of peptides, has revealed unexpected properties of combinatorial random peptide libraries and indicates correctives to ensure the quality of the libraries generated. Next, Deep Panning is used to analyze a model monoclonal antibody in addition to allowing one to follow the dynamics of biopanning and peptide selection. Finally Deep Panning is applied to profile polyclonal sera derived from HIV infected individuals. CONCLUSIONS/SIGNIFICANCE: The ability to generate and characterize hundreds of thousands of affinity-selected peptides creates an effective means towards the interrogation of the IgOme and understanding of the humoral response to disease. Deep Panning should open the door to new possibilities for serological diagnostics, vaccine design and the discovery of the correlates of immunity to emerging infectious agents.
