Cloning and characterization of the groE locus from Actinobacillus pleuropneumoniae

The aims of the present investigation were to develop and test a sensitive and reproducible method for the study of gene expression in the porcine lung pathogen Actinobacillus pleuropneumoniae by real-time quantitative reverse transcription (RT)-PCR and to evaluate a number of suitable internal controls, as such controls have not been defined yet for this bacterium. Bacterial gene expression was studied during in vitro exponential and early stationary growth in medium with and without sufficient iron, respectively. First, the stability of expression of five genes, the glyA, tpiA, pykA, recF, and rhoAP genes involved in basic housekeeping, was evaluated on the basis of the mean pairwise variation. All the housekeeping genes included were stably expressed under the conditions investigated and consequently were included in the normalization procedure. Next, the geometric mean of the internal control genes was used to correct five genes of interest. These genes were three genes involved in iron acquisition (tbpA, exbB, and fhuD), the heat shock protein gene groEL, and a putative quorum-sensing gene (luxS). The level of tbpA, exbB, and fhuD expression in A. pleuropneumoniae showed significant up-regulation under iron-restricted conditions compared to bacteria grown in medium with sufficient iron. The observed expression patterns of the genes of interest were consistent with previous observations. This study therefore lends further support to the use of real-time quantitative RT-PCR, with the glyA, tpiA, pykA, recF, and rhoAP genes as internal controls, for future similar gene expression studies in A. pleuropneumoniae.

show abstract

Real-Time Quantitative Reverse Transcription-PCR Analysis of Expression Stability ofActinobacillus pleuropneumoniaeHousekeeping Genes during In Vitro Growth under Iron-Depleted Conditions

Klitgaard

Boye

2005

Appl Environ Microbiol

View full text Add to dashboard Cite

show abstract

Cloning of the Lactococcus lactis adhE Gene, Encoding a Multifunctional Alcohol Dehydrogenase, by Complementation of a Fermentative Mutant of Escherichia coli

Arnau¹,

Jørgensen²,

Madsen³

et al. 1998

J Bacteriol

View full text Add to dashboard Cite

The Lactococcus lactis adhE gene, which encodes a multifunctional alcohol dehydrogenase, has been cloned and characterized. A DNA fragment encoding the putative alcohol dehydrogenase domain of the AdhE protein was cloned by screening anL. lactis genomic library in a fermentative mutant ofEscherichia coli and selecting for the ability to grow anaerobically. Further analysis of the clone obtained allowed the cloning of the entire adhE gene sequence. Analysis ofadhE expression in L. lactis during anaerobiosis showed induction at the transcriptional level, especially in medium containing glucose. Constructed mutant strains produced reduced amounts of ethanol under anaerobic conditions. With theL. lactis gene as a probe, adhE homologs were found in other industrially relevant lactic acid bacteria.

show abstract

scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.

Contact Info

customersupport@researchsolutions.com

10624 S. Eastern Ave., Ste. A-614

Henderson, NV 89052, USA

This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.

Blog Terms and Conditions API Terms Privacy Policy Contact Cookie Preferences Do Not Sell or Share My Personal Information

Made with 💙 for researchers

Part of the Research Solutions Family.

Cloning and characterization of the groE locus from Actinobacillus pleuropneumoniae

Cited by 2 publications

References 13 publications

Real-Time Quantitative Reverse Transcription-PCR Analysis of Expression Stability ofActinobacillus pleuropneumoniaeHousekeeping Genes during In Vitro Growth under Iron-Depleted Conditions

Real-Time Quantitative Reverse Transcription-PCR Analysis of Expression Stability ofActinobacillus pleuropneumoniaeHousekeeping Genes during In Vitro Growth under Iron-Depleted Conditions

Cloning of the Lactococcus lactis adhE Gene, Encoding a Multifunctional Alcohol Dehydrogenase, by Complementation of a Fermentative Mutant of Escherichia coli

Contact Info

Product

Resources

About