Clc genomics workbench 8.5.112/2/2023 Dead ones were picked out and stored in a refrigerator. The inoculated eggs were further incubated for 3 days, and checked twice each day during the incubation period. The SPF embryonated eggs were purchased from Shandong Healthtec Laboratory Animal Breeding Company (Jinan, China). The swab sample was clarified by centrifugation at 10,000× g for 5 min, and the supernatants were inoculated in 10-day-old SPF embryonated chicken eggs via the allantoic sac route. The sample was negative for Avian influenza virus (AIV) detection, but caused death to specific-pathogen-free (SPF) embryonated chicken eggs in 72 h. The swab sample was collected through taking smears at both cloacal and oropharyngeal tracts, and stored in 1.5 ml phosphate buffered saline (PBS, pH 7.2) containing 10% glycerol. Sample collectionĪ swab sample was collected from a duck in a live bird market from Guizhou province, China, in October of 2013. Swab samples were collected by gently taking smears from the trachea and cloacae of domestic fowl in China and then placed in a transport medium. Fecal samples were collected from fresh feces in the ground of poultry farms in China. The fecal and swab samples were all collected with permission given by the multiple relevant parties, including the Ministry of Agricultural of China, China Animal Health and Epidemiology Center, the relevant veterinary sections in the provincial and county government. ![]() This study was conducted according to the animal welfare guidelines of the World Organization for Animal Health, and approved by the Animal Welfare Committee of China Animal Health and Epidemiology Center. The effectiveness and practicality to identify viruses and sequence their genomes using this method are discussed in this study. In order to establish the NGS platform in diagnosis and surveillance of viral infection, we developed a NGS library preparation method based on RT-PCR random primers. A suitable pipeline of library construction is very essential for virus genome sequencing by NGS. Each NGS platform has its own sequencing library preparation procedure. Among these platforms, Ion Torrent PGM is competitive for detection of viruses and bacteria with respect to instrumental price, sequencing cost, and simplicity of operation, although its sequencing throughput is lower than MiSeq and Proton. This facilitates research in virus ecology, novel virus discovery, and the development of larger datasets of complete virus genomes for studies on virus evolution and pandemic prediction.įour popular second-generation sequencing platforms have been released: Illumina HiSeq, MiSeq and NovaSeq, Ion Torrent PGM, Proton and S5, BGISeq-500, have been commercially available. ![]() ![]() Unlike insensitive traditional virological methods and highly specific reverse transcription-polymerase chain reaction (RT-PCR), NGS methods have the advantage of being able to sequence total or targeted DNA and RNA from samples in an unbiased way, without a priori knowledge of the possible viral agent(s) present, thus making them the ideal tool for novel and divergent viral genome discovery. Numerous viruses and variant strains have been identified using NGS approaches. ![]() Since the development of next generation sequencing (NGS) technologies, great progress has been made in the rapid identification and characterization of RNA viruses. Identification and analysis of RNA viruses are important for the diagnosis, treatment, control, and prevention of human and animal infectious diseases. RNA viruses are the agents of many human, animal, and plant infectious diseases, including influenza, severe acute respiratory syndrome (SARS), and so on.
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