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Virology ; 278(1): 103-10, 2000 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-11112486

RESUMO

We previously described the use of extended polymerase chain reaction (PCR) to amplify contiguous 9.2-kilobase (kb) single-long terminal repeat (LTR) proviral sequences from HIV-1 genetic subtypes A through G. We now extend these findings by describing a novel vector system to recover infectious molecular clones from long PCR amplicons. Directional ligation of 9.2-kb proviral amplicons into a recovery vector reconstitutes missing LTR sequences, providing candidate molecular clones for infectivity screening. We show that a previously characterized infectious molecular clone of HIV-1 retains its biological properties upon recovery with this strategy. Three additional infectious molecular clones generated, from primary isolates of subtype B (HIV-1(WR27)) and circulating recombinant form 01_AE (subtype E) (HIV-1(CM235)) by subtype-specific LTR reconstitution, displayed biological properties reflecting their cognate parental isolates. This represents the first report of infectious molecular clones from circulating recombinant form 01_AE (subtype E).


Assuntos
HIV-1/genética , Provírus/genética , Células Cultivadas , Clonagem Molecular , DNA Viral/análise , Proteína do Núcleo p24 do HIV/análise , Repetição Terminal Longa de HIV/genética , HIV-1/crescimento & desenvolvimento , HIV-1/imunologia , HIV-1/fisiologia , Humanos , Cinética , Reação em Cadeia da Polimerase/métodos , Provírus/fisiologia , Replicação Viral
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