Fluorescent dye terminator sequencing methods for quantitative determination of replication fitness of human immunodeficiency virus type 1 containing the codon 74 and 184 mutations in reverse transcriptase.

ABSTRACT: The fluorescent dye-labeled dideoxynucleotide automated DNA sequencing system has been routinely used for monitoring the development of resistance mutations in human immunodeficiency virus type 1 reverse transcriptase (RT) and protease genes during therapy. This system has provided information regarding the presence of mixtures of nucleotides in the clinical samples but has not previously been validated for the quantitative determination between peak heights and relative DNA concentration. We evaluated this system by using various ratios of wild-type and mutated DNA fragments and by performing sequencing reactions at actual melting temperatures of specific primers. Several different ratios of purified DNA fragments containing mixtures of L74/V74 and M184/V184 were sequenced, and peak heigh

SUBMITTER: Nurpeisov V 

PROVIDER: S-EPMC165373 | biostudies-literature | 2003 Jul

REPOSITORIES: biostudies-literature