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Figure 5 | BMC Microbiology

Figure 5

From: A gonococcal homologue of meningococcal γ-glutamyl transpeptidase gene is a new type of bacterial pseudogene that is transcriptionally active but phenotypically silent

Figure 5

Transcriptional expression of the ggh genes. A. Dot blot analysis using the meningococcal ggt gene as a probe. One to 16 micrograms of RNAs isolated from H44/76, HT1089 (H44/76 Δggt::spc) [39], ATCC49226, NIID54, NIID103 and NIID106 were subjected to this analysis. B. RT-PCR to detect the transcripts of the gonococcal ggh genes. The schematic figure in the box depicts the position of the primers used in this experiment (see Table 1). RT-PCR was performed without reverse transcriptase (RTase) (lanes 1 to 5) or with RTase (lanes 6 to 10). Lanes 1 and 6, H44/76 (N. meningitidis); lanes 2 and 7, ATCC49226 (N. gonorrhoeae); lanes 3 and 8, NIID54 (N. gonorrhoeae); lanes 4 and 9, NIID103 (N. gonorrhoeae); lanes 5 and 10, NIID106 (N. gonorrhoeae). The marker in the left-most lane is φ X174 DNA digested with Hae III. Primer sets used for RT-PCR are shown on the left side, and the corresponding PCR products are indicated by arrows on the right side. C. Primer extension analysis to detect the transcriptional start point of the ggt and ggh genes. Total RNA extracted from H44/76 (N. meningitidis), ATCC49226 and NIID106 (N. gonorrhoeae) was used for the primer extension with AMV reverse transcriptase XL and biotin-labeled oligonucleotide ggt-ext-2. The arrow on the right side indicates the transcriptional start site. D. Alignment of the nucleotide sequences of the upstream regions of the ggt and ggh genes. The sequence data have been deposited in the DDBJ/EMBL/GenBank Databases under the following Accession Numbers:N. meningitidis strains H44/76 [DDBJ:AB193252], H114/90 [DDBJ:AB193253], N. gonorrhoeae strains ATCC49226 [DDBJ:AB193254], NIID54 [DDBJ:AB193255], NIID103 [DDBJ:AB193296], NIID106 [DDBJ:AB193256]. An identical nucleotide is represented as *. The transcriptional start site is shown in bold as +1. The putative -35, -10 elements and Shine-Dalgarno sequence (SD) are depicted in the box, and the ideal -35 and -10 nucleotide sequences are shown above the boxes. The previously predicted start codon (ATG) [25] and newly predicted start codon (GTG) of the meningococcal {\it ggt} gene are underlined. The amino acid sequence deduced from the putative start codon GTG (shown in bold) in the meningococcal {\it ggt} gene is also shown under the corresponding nucleotide sequences.

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