[Standardization of rabies virus genome amplification for its use in molecular epidemiology studies].
In order to improve the diagnosis and typification of rabies viruses at the Instituto Nacional de Salud National Reference Laboratory for rabies, we standardized techniques for the amplification of a 902 nucleotide DNA fragment, complementary to a selected region of the rabies virus genomic RNA. This region codes for a segment of both the glycoprotein and protein L, and contains the G-L intergenic noncoding region known as Pseudogen Psi. The standardized techniques included: 1) biological amplification of rabies viruses by intracerebral mouse inoculation; 2) total RNA extraction from the brains of infected mice, 3) RT-PCR amplification of a 902 nucleotide DNA fragment complementary to the selected RNA region. The study sample consisted of 30 rabies virus strains isolated from dogs and selected from the virus bank of the Virology Laboratory. Due to their simplicity the methods described have several advantages when compared to methods reported in previous papers. The technology proposed is a precise complement to rabies diagnosis techniques, and can be applied to the identification of phylogenetic relations among rabies isolates, and, therefore, it is also used to identify rabies transmission dynamics and geographical distribution.
Páez
Andrés
A
Laboratorio de Virología, Instituto Nacional de Salud, Bogotá, D.C., Colombia. apaezm@hemagogus.ins.gov.co
García
Clemencia
C
Boshell
Jorge
J
D000818:Animals; D001921:Brain / Q000821:virology; D016679:Genome, Viral; D051379:Mice; D017720:Molecular Epidemiology / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000592:standards*; D012367:RNA, Viral / Q000235:genetics* / Q000302:isolation & purification; D011820:Rabies virus / Q000235:genetics* / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012367:RNA, Viral