Abstract
Peste-des-petits-ruminants virus (PPRV) is currently the focus of a control and eradication program. Full genome sequencing has the opportunity to become a powerful tool in the eradication program by improving molecular epidemiology and the study of viral evolution. PPRV is prevalent in many resource-constrained areas, with long distances to laboratory facilities, which can lack the correct equipment for high-throughput sequencing. Here we present a protocol for near full or full genome sequencing of PPRV. The use of a portable miniPCR and MinION brings the laboratory to the field and in addition makes the production of a full genome possible within 24 h of sampling. The protocol has been successfully used on virus isolates from cell cultures and field isolates from tissue samples of naturally infected goats.
Highlights
With the development of new and portable sequencing equipment, it is possible to perform—in very basic laboratories—sequencing that was previously limited to well-equipped laboratories [1,2,3,4]
With a small thermocycler such as the miniPCR (Amplyus, Cambridge, United States), the hand-held MinION sequencer (Oxford Nanopore Technologies, Oxford, United Kingdom), and portable computational resources, full genome sequencing and advanced molecular epidemiology can be performed in almost any setting [1,2,3,4]
The cause is the peste-des-petits-ruminants virus (PPRV), a single-stranded negative-sense RNA virus belonging to the genus Morbillivirus [6]
Summary
With the development of new and portable sequencing equipment, it is possible to perform—in very basic laboratories—sequencing that was previously limited to well-equipped laboratories [1,2,3,4]. With a small thermocycler such as the miniPCR (Amplyus, Cambridge, United States), the hand-held MinION sequencer (Oxford Nanopore Technologies, Oxford, United Kingdom), and portable computational resources, full genome sequencing and advanced molecular epidemiology can be performed in almost any setting [1,2,3,4]. This is highly advantageous for the diagnosis and control of viral diseases. The cause is the peste-des-petits-ruminants virus (PPRV), a single-stranded negative-sense RNA virus belonging to the genus Morbillivirus [6]. Other morbilliviruses include canine distemper virus, measles virus, feline morbillivirus, marine morbilliviruses, and the eradicated rinderpest virus (RPV) [7]
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