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Figure 2 | Microbial Cell Factories

Figure 2

From: Regulation of mtl operon promoter of Bacillus subtilis: requirements of its use in expression vectors

Figure 2

Promoter of mtl operon fused to lacZ (A) and β-galactosidase activity of the constructs (B). (A) Upstream sequence of lacZ in plasmid pKAM1 and deletion derivatives are shown. The intervening sequence between stop codon of ycnL (bold capital letter) and mtlA start codon was obtained from B. subtilis and was inserted between Nhe I and Afl II site of the vector. It includes the promoter elements of mtl operon (P mtlA ). The promoter -10 and -35 putative boxes are enclosed by rectangles, while the single C residue (bold capital letter) is the transcription start site. The determined cre site (underline) has an overlap with the -10 box. Ribosomal binding sites of the promoter (W) and vector (V) are underlined, and the lacZ start codon is marked in bold capital letters. The first bases of the shortened promoters are shown by boxes and the arrows. (B) Activity of different constructs of P mtlA in B. subtilis 3NA containing the wild type promoter (pKAM1), the 5' shortened P mtlA , i.e., pKAM9, pKAM43, pKAM48, pKAM49, pKAM57, pKAM58, and pKAM59, as well as 3' shortened P mtlA , i.e. pKAM12 and pKAM44, were induced by 0.2% of mannitol at OD600 of 0.4. β-galactosidase activity of the cells was measured after 1 h of induction. Plasmids pKAM27, pKAM45, pKAM52, pKAM84, and pKAM92 contain the complementary base pairs in comparison to a wild type promoter in the discriminated region between the reported MtlR binding site and -35 box.

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