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. 2003 Mar;185(6):1783–1795. doi: 10.1128/JB.185.6.1783-1795.2003

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Copyright © 2003, American Society for Microbiology

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FIG. 2. — Organization of the GntI and GntII genes, and clones for disruptants and lacZ operon fusions. (A) The GntII genes, gntV, idnD, idnO, gntW, and gntH, located on the E. coli W3110 genome are represented by open boxes. Parts of the Kohara clones, E4D8 and 5C4, are shown at the top. The direction of their transcription from promoters, gntVp, idnDp, and gntHp, are shown by arrows. Plasmid pGNT2 with the 8.4-kb BglII-PstI fragment from E4D8 bears all five genes. pGNTH and pGNTH119 with the 1.7-kb DraI fragment bear the gntH gene. pGNTH18 containing the 1.7-kb PstI-BamHI fragment from pGNTH119 bears the gntH gene. (B) The open boxes indicate the GntI and GntII genes. pGNTR18 containing the 1.0-kb PCR fragment bears gntR. pGNTR177-CM, pGNTH-DIS2-CM, and pBRIDNO-CM bear the gntR, gntH, and idnO genes, respectively, which have insertion of the cml gene (open boxes) and are used for gene disruption. (C and D) The gntRKU and gntT genes (open boxes) are shown at the top. The regions inserted in the gnt-lacZ operon fusions and the promoterless lacZ genes are shown by hatched and dotted boxes, respectively. The promoter-operator regions of gntKU (C) and gntT (D), based on the previous data (12, 13, 25), are schematically represented at the middle and bottom, respectively. Arrows indicate the initiation sites and direction of the gntKU and gntT mRNAs. The corresponding promoter sequences with the −10 and −35 sequences are shown by brackets. The GntR- and cAMP-CRP-binding sites are represented by boxes, and the ribosome-binding sites (SD) and the start codons of gntK and gntT are also shown. Three mutants (MR1, MR2, and MR12) and their mutation sites (boxed) in the GntR-binding elements of the gntK promoter-operator region are shown at the bottom of panel C. The procedures for cloning and mutant construction are described in Materials and Methods. Hatched boxes represent the DNA fragments inserted into vectors, which were derived from the genomic DNA, and arrowheads P1 to P18 represent the position and direction of primers corresponding to those in Table 1.

FIG. 2. — Organization of the GntI and GntII genes, and clones for disruptants and lacZ operon fusions. (A) The GntII genes, gntV, idnD, idnO, gntW, and gntH, located on the E. coli W3110 genome are represented by open boxes. Parts of the Kohara clones, E4D8 and 5C4, are shown at the top. The direction of their transcription from promoters, gntVp, idnDp, and gntHp, are shown by arrows. Plasmid pGNT2 with the 8.4-kb BglII-PstI fragment from E4D8 bears all five genes. pGNTH and pGNTH119 with the 1.7-kb DraI fragment bear the gntH gene. pGNTH18 containing the 1.7-kb PstI-BamHI fragment from pGNTH119 bears the gntH gene. (B) The open boxes indicate the GntI and GntII genes. pGNTR18 containing the 1.0-kb PCR fragment bears gntR. pGNTR177-CM, pGNTH-DIS2-CM, and pBRIDNO-CM bear the gntR, gntH, and idnO genes, respectively, which have insertion of the cml gene (open boxes) and are used for gene disruption. (C and D) The gntRKU and gntT genes (open boxes) are shown at the top. The regions inserted in the gnt-lacZ operon fusions and the promoterless lacZ genes are shown by hatched and dotted boxes, respectively. The promoter-operator regions of gntKU (C) and gntT (D), based on the previous data (12, 13, 25), are schematically represented at the middle and bottom, respectively. Arrows indicate the initiation sites and direction of the gntKU and gntT mRNAs. The corresponding promoter sequences with the −10 and −35 sequences are shown by brackets. The GntR- and cAMP-CRP-binding sites are represented by boxes, and the ribosome-binding sites (SD) and the start codons of gntK and gntT are also shown. Three mutants (MR1, MR2, and MR12) and their mutation sites (boxed) in the GntR-binding elements of the gntK promoter-operator region are shown at the bottom of panel C. The procedures for cloning and mutant construction are described in Materials and Methods. Hatched boxes represent the DNA fragments inserted into vectors, which were derived from the genomic DNA, and arrowheads P1 to P18 represent the position and direction of primers corresponding to those in Table 1.