Novel single-round PCR and cloning of full-length envelope genes of HIV-1 may yield new insight into biomolecular antibacterial drug development.
Nested or semi-nested polymerase chain reaction (PCR) with a 'hot start' is the preferred amplification method for full-length, in-frame envelope genes (gp160) of the human immunodeficiency virus type 1 (HIV-1). This generally follows an extensive screening process. This paper describes an effective single-round PCR method and cloning process for HIV-1 gp160 from clinical samples, and cell and tissue cultures developed during the early stages of construction of a molecular HIV-1 vaccine. The amplification method and cloning process are adaptable to full-length HIV-1, HIV-2, and other viral production processes. Also described within, is one solution to the most-often extensive screening process for inserts containing full-length, in-frame gp160. Of note, was a perceived toxicity of gp160 to bacteria during the culturing and the scaling-up process that created the extensive screening process. The toxicity association was not found with the individual gp160 genes, the gp120 or the gp41 gene, with other viral regions similar or larger in molecular weight to gp160, or with other non-gp160 full-length genes of HIV-1 such as pol and gag genes. The HIV-1 gp160 toxicity issue may provide insight towards the development of the next generation of novel biomolecular drugs against bacterial infections.
McDonald
Richard
R
Biomedical Health Sciences Division, Genovar Diagnostics, 1030 Heeley Close, Sittingbourne Research Centre, Sittingbourne ME9 8HL, Kent, UK. mcdonald@scientist.com
Burnett
Virginia
V
D016915:AIDS Vaccines; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D015752:Genes, env; D019302:HIV Envelope Protein gp160 / Q000235:genetics* / Q000633:toxicity*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000633:toxicity
D016915:AIDS Vaccines; D019302:HIV Envelope Protein gp160; D012367:RNA, Viral; D011994:Recombinant Proteins