Transcript analysis of 1003 novel yeast genes using high-throughput northern hybridizations

The expression of 1008 open reading frames (ORFs) from the yeast Saccharomyces cerevisiae has been examined under eight different physiological conditions, using classical northern analysis. These northern data have been compared with publicly available data from a microarray analysis of the diauxic transition in S.cerevisiae. The results demonstrate the importance of comparing biologically equivalent situations and of the standardization of data normalization procedures. We have also used our northern data to identify co-regulated gene clusters and define the putative target sites of transcriptional activators responsible for their control. Clusters containing genes of known function identify target sites of known activators. In contrast, clusters comprised solely of genes of unknown function usually define novel putative target sites. Finally, we have examined possible global controls on gene expression. It was discovered that ORFs that are highly expressed following a nutritional upshift tend to employ favoured codons, whereas those overexpressed in starvation conditions do not. These results are interpreted in terms of a model in which competition between mRNA molecules for translational capacity selects for codons translated by abundant tRNAs

Sensitive testing of plasma HIV-1 RNA and Sanger sequencing of cellular HIV-1 DNA for the detection of drug resistance prior to starting first-line antiretroviral therapy with etravirine or efavirenz

OBJECTIVES: This study investigated strategies that may increase the yield of drug resistance testing prior to starting antiretroviral therapy (ART), and whether transmitted and polymorphic resistance-associated mutations (RAMs) correlated with virological outcomes. METHODS: We carried out retrospective testing of baseline samples from patients entering the SENSE trial of first-line ART in Europe, Russia and Israel. Prior to randomization to etravirine or efavirenz plus two nucleos(t)ide reverse transcriptase inhibitors (NRTIs), plasma samples underwent routine Sanger sequencing of HIV-1 RT and protease ((plasma)SS) in order to exclude patients with transmitted RAMs. Retrospectively, Sanger sequencing was repeated with HIV-1 DNA from baseline peripheral blood mononuclear cells (PBMCSS); baseline plasma samples were retested by allele-specific PCR targeting seven RT RAMs (AS-PCR) and ultra-deep RT sequencing (UDS). RESULTS: By (plasma)SS, 16/193 (8.3%) patients showed ≥ 1 transmitted RAM affecting the NRTIs (10/193, 5.2%), non-nucleoside reverse transcriptase inhibitors (4/193, 2.1%) or protease inhibitors (2/193, 1.0%). No additional RAMs were detected by AS-PCR (n = 152) and UDS (n = 24); PBMCSS (n =  91) yielded two additional samples with one RAM each. Over 48 weeks, 4/79 (5.1%) patients on etravirine and 7/78 (9.0%) on efavirenz experienced virological failure; none had baseline RAMs. Conversely, 11/79 (13.9%) patients randomized to etravirine had one polymorphic RAM from the etravirine score in baseline plasma (V90I, V106I or E138A), without any impact on virological outcomes. CONCLUSIONS: The detection of resistance increased marginally with PBMC testing but did not increase with sensitive plasma testing. A careful consideration is required of the cost-effectiveness of different strategies for baseline HIV drug resistance testing