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Introduction
Lactobacilli are members of the lactic acid bacteria, a functional group related by formation of lactic acid as the primary product of carbohydrate metabolism. Lactobacilli have long been considered beneficial, occupying important niches in the gastrointestinal (GI) tracts of humans and animals. Select lactobacilli are increasingly recognized as modulators of health, gaining interest as microbes used as health‐promoting functional food ingredients, and as delivery vectors for vaccines and biotherapeutics (Wells et al., 1996).
There continues to be great interest in the development of genetic tools for production of proteins and enzymes from lactic acid bacteria. Several gene expression systems for lactobaclli have been developed. The widely used nisin‐controlled expression (NICE) system, originally developed for use in Lactococcus lactis (de Ruyter et al., 1996), has been adapted for use in lactobacilli (Kleerebezem et al., 1997; Wu et al., 2006). Expression systems based on control by other bacteriocins (Axelsson et al., 2003; Mathiesen et al., 2004) and lactose (Gosalbes et al., 2001) are also available.
Lactobacillus acidophilus NCFM is a probiotic culture widely used in nutritional supplements, dairy products and infant formulas. The availability of the L. acidophilus genome sequence (Altermann et al., 2005), gene expression profiling (Azcarate‐Peril et al., 2005; 2006; Barrangou et al., 2006) and functional genomic studies (Russell and Klaenhammer, 2001a;Barrangou et al., 2003; Duong et al., 2006) has provided considerable insight into the physiology of this organism and established a technical basis that can be used to improve both the fermentation and probiotic functionalities of this organism.
In this study, we exploit the genome sequence, gene expression profiling and functional genomic data to construct a series of expression vectors and analyse their properties using a β‐glucuronidase (GusA3) reporter protein. Additionally, one vector encoding a strong constitutive promoter was employed for overexpression of the L. acidophilus oxalate‐degradation operon and complementation of a deletion mutation therein (Azcarate‐Peril et al., 2005).
Results
Promoter selection
Previous global gene expression analysis found a number of genes involved in carbohydrate transport and metabolism to be differentially expressed during growth on various carbohydrates (Barrangou et al., 2006). Also, a number of genes were...