Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Current HIV-1 genotyping assays were developed using subtype B viruses prevalent in Western countries. It is not clear whether these assays are appropriate for use among African patients, who are likely to be infected with non-B subtypes. We evaluated the Bayer TRUGENE HIV-1 genotyping (TG) assay using prospectively collected samples from HIV-1-infected individuals who acquired infection in either sub-Saharan Africa or the West (Europe, North America, and Australia). Plasma samples from 208 individuals with an HIV-1 viral load of >1,000 copies/ml were tested using version 1 primers supplied with the TG assay. If these failed, an alternative primer set version 1.5 was used. Of the 208 individuals, the likely origin of infection was Africa (n = 104), Western (n = 87) and "Others" (i.e., all other geographic locations or origin not certain; n = 17). Among the three groups, the version 1 primers were successful in 85 (82%), 77 (89%), and 13 (76%) individuals, respectively (P = 0.1). Of the remaining 32 samples, 30 were successfully amplified by using the version 1.5 primers. HIV-1 subtypes deduced from the reverse transcriptase sequences correlated with the likely origin of infection: Africa (28A, 3B, 33C, 13D, 6G, 4J, 2K, 5CRF01_AE, and 10CRF02_AG), Western (86B and 1K), and Others (1A and 16B). The success of the version 1 primers correlated with viral load (P < 0.014) and not with HIV-1 subtypes. A protocol based on version 1 primers, followed by 1.5 primers, was successful in sequencing 99% of the samples in this cohort.
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PMID:Genotyping of B and non-B subtypes of human immunodeficiency virus type 1. 1614 17

Nucleic acid amplification testing for hepatitis C virus (HCV) RNA has become an essential tool for the prevention and clinical management of hepatitis C. We describe the development, validation and evaluation of a homogenous reverse transcriptase-initiated HCV-PCR assay with competitive internal control that is applicable to both the quantitative detection of HCV genomes in single patient samples and the screening of blood donations by mini-pool testing. For the implementation of a positive run control, a HCV RNA-positive plasma sample was calibrated against an international HCV RNA standard preparation. For quantification purposes, an in vitro-transcribed RNA calibrator sequence was used. The detection limit of the assay (95% positive cut-off) was determined by probit analysis and was calculated as 114 IU/mL. Comparable sensitivity to different HCV template sequences was verified for HCV genotypes 1-5. Quantitative test results correlated well with viral loads that had been previously determined by the Bayer VERSANT HCV RNA 3.0 bDNA assay (n=53, R=0.943, p<0.001). During more than 5 years of blood donation testing, the specificity of the assay was found to be 99.51%. All assay components showed constant performance during this time period. In conclusion, we introduce a well-proven method that allows fast and reliable quantification of HCV genomes.
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PMID:Development, validation and evaluation of a homogenous one-step reverse transcriptase-initiated PCR assay with competitive internal control for the detection of hepatitis C virus RNA. 1620 92

Human immunodeficiency virus type 1 (HIV-1) protease (PR) and reverse transcriptase (RT) gene sequences obtained during antiretroviral resistance testing with a commercial genotyping assay (Tru Gene; Bayer Corp.) were analyzed to assess the utility of these data for detecting and characterizing non-subtype-B HIV-1 strains. A total of 125 viral sequences obtained from patients believed to have acquired their HIV-1 infection in Africa were analyzed, of which 121 were determined to belong to non-B subtypes. Utilizing Tru Gene sequence data alone, 92 (76%) of these viruses could be subtyped by conventional phylogenetic analysis. The addition of supplemental RT sequence data enabled a further 28 (23.1%) viruses to be classified, while one (0.9%) sample could not be classified conclusively. Two internet-accessible databases that generate HIV-1 subtypes from PR and RT sequences (HIV-SEQ and Geno 2 Pheno) were also evaluated, and both achieved 88% concordance (106/120) with phylogenetic analysis. Non-subtype-B and B-subtype HIV-1 sequences could be readily discriminated by tallying silent polymorphisms listed on the Tru Gene research report. The mean number of silent polymorphisms in the non-B HIV-1 sequences identified in this study was 58.3 (95% confidence interval [CI], 41.1 to 75.5), compared with 20.7 (95% CI, 9.9 to 31.5) for the four subtype B viruses in the study cohort and 118 case-matched B-subtype controls. Sequence data generated in the Tru Gene HIV-1 genotyping assay could, therefore, provide a ready means of tracking the prevalence and identity of non-B subtypes in HIV-1-infected populations undergoing routine antiretroviral resistance testing.
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PMID:Use of sequence data generated in the Bayer Tru Gene genotyping assay to recognize and characterize non-subtype-b human immunodeficiency virus type 1 strains. 1620 93

The TruGene human immunodeficiency virus type 1 (HIV-1) genotyping kit/OpenGene DNA sequencing system (Bayer HealthCare, Tarrytown, NY) reliably produced clinically acceptable resistance profiles for reverse transcriptase and protease inhibitors on patient samples diluted to approximately 100 copies/ml following extraction with the QIAamp viral RNA minikit (QIAGEN Inc., Valencia, CA). One modification of the standard protocol was made to guarantee PCR amplification: a centrifugation step to concentrate virus was added before RNA extraction. For genotypic antiretroviral resistance testing, no significant differences in the identification and sensitivity of detection for codon mutations, base mutations, and multibase sites were found between the original and diluted samples.
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PMID:Performance of the TruGene human immunodeficiency virus type 1 genotyping kit and OpenGene DNA sequencing system on clinical samples diluted to approximately 100 copies per milliliter. 1646 31

The high genetic diversity of HIV-1 has a major impact on the quantification of plasma HIV-1 RNA, representing an increasingly difficult challenge. A total of 898 plasma specimens positive for HIV-1 RNA by commercial assays (Amplicor v1.5; Roche Diagnostic Systems, Alameda, CA or Versant v3.0; Bayer Diagnostics, Emeryville, CA) were tested using the Agence Nationale de Recherches sur le SIDA second-generation (G2) real-time reverse transcriptase polymerase chain reaction (RT-PCR) test: 518 samples containing HIV-1 of known subtype, including 88 from 2 subtype panels and 430 harboring B (n = 266) and non-B (n = 164) group M HIV-1 subtypes from patients followed up in 2002 through 2005 at Necker Hospital (Paris, France), and 380 samples from 10 different countries (Argentina, Cambodia, Cameroon, Central African Republic, France, Ivory Coast, Madagascar, Morocco, Thailand, and Zimbabwe). HIV-1 RNA values obtained by G2 real-time PCR were highly correlated with those obtained by the Amplicor v1.5 for B and non-B subtypes (R = 0.892 and 0.892, respectively) and for samples from diverse countries (R = 0.867 and 0.893 for real-time PCR vs. Amplicor v1.5 and real-time PCR vs. Versant v3.0, respectively). Approximately 30% of specimens harboring non-B subtypes were underquantified by at least -0.51 log10 in Amplicor v1.5 versus 5% underquantified in G2 real-time PCR. Discrepant results were also obtained with subtype B samples (14% underquantified by Amplicor v1.5 vs. 7% by G2 real-time PCR). Similar percentages were observed when comparing results obtained with the G2 real-time PCR assay with those obtained using the Versant assay. Addressing HIV-1 diversity, continual monitoring of HIV-1 RNA assays, together with molecular epidemiology studies, is required to improve the accuracy of all HIV RNA assays.
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PMID:Impact of HIV-1 genetic diversity on plasma HIV-1 RNA Quantification: usefulness of the Agence Nationale de Recherches sur le SIDA second-generation long terminal repeat-based real-time reverse transcriptase polymerase chain reaction test. 1746 66


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