pmdy23 plasmid vector長雙歧桿菌穿梭表達質粒載體

the versatile plasmid pMDY23 based on an Escherichia coli origin of replication, a small cryptic B. longum plasmid, and the Escherichia coli β-glucuronidase gene, gusA (5), that has been successfully used as a reporter gene in a wide range of organisms (6, 7, 15, 20). pMDY23 has been designed to function (i) as a reporter for promoter strength or regulation analysis and (ii) for the identification of genomic fragments containing active promoters (promoter probe). Here we demonstrate the use of pMDY23 as a reporter plasmid with the analysis of the activity of three B. longum promoters. Preliminary microarray-based gene expression data were used to identify probable expressed genes, and the regions directly preceding two of them were tested for promoter activity in plasmid pMDY23. These were the potential promoters pr-BL1363 (promoter of gene BL1363) preceding BL1363, encoding the enzyme glyceraldehyde 3-phosphate dehydrogenase, and pr-BL1613, preceding BL1613, encoding a member of the LacI family of bacterial regulator proteins (Pfam no. PF00356). Secondly, a predicted sugar-regulated promoter, pr-BL1518, preceding the gene BL1518, encoding the enzyme α-galactosidase, was also tested.

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