Direct sequencing of large flavivirus PCR products for analysis of genome variation and molecular epidemiological investigations

Molecular epidemiology of dengue 3 viruses and genetic relatedness among dengue 3 strains isolated from patients with mild or severe form of dengue fever in French Polynesia

“…Only limited genetic information on dengue 3 viruses is available (Osatomi & Sumiyoshi, 1990;Lewis et al, 1992). Our results on a short fragment of the E protein gene provided data on the variation among geographically distinct isolates thus allowing their genotypic grouping.…”

Identification of a major determinant of mouse neurovirulence of dengue virus type 2 using stably cloned genomic-length cDNA.

“…Reverse transcription (RT)-PCR was used to synthesize cDNA which was either used in the production of gene constructs or sequenced to verify the identity of recombinant viruses. First strand cDNA synthesis was performed using DEN-2 RNA extracted from 250 µl culture supernatant fluid using the acid-guanidinium isothiocyanate method of Lewis et al (1992). Onefifth of the RNA isolated was mixed with a DEN-2 specific oligonucleotide primer and RNA was reverse-transcribed using SuperScript II (Life Technologies).…”

Heterogeneity in the haemagglutinin gene and emergence of the highly pathogenic phenotype among recent H5N2 avian influenza viruses from Mexico

“…RT-PCR was performed in a single tube (Lewis et al, 1992) using a DNA Thermal Cycler (Perkin Elmer). The RT reaction was conducted at 42 °C for I h using Superscript (Life Technologies), followed by denaturation at 99 °C for 20 min.…”