Figure 1.
Construction of expression variants in pGL3 vector using oligonucleotides for the proximal AGT promoter.
The oligonucleotides containing the proximal promoter region of the AGT gene from position −290 to +35 were ligated in the pGL3 vector to produce the reporter construct. luc+ = cDNA encoding the modified firefly luciferase; Ampr = gene conferring ampicillin resistance; f1 ori = origin of replication derived from filamentous phage; E = exon.
Table 1.
General characteristics of patients included in the study.
Figure 2.
AGT G-6A polymorphism genotyping by direct sequencing.
The arrows indicate the polymorphic site of GG, AA and AG genotypes.
Figure 3.
Linkage disequilibrium plot of AGT promoter polymorphisms.
Pairwise linkage disequilibrium analysis shows r2 (×100) values. The intensity of gray is proportional to r2.
Table 2.
Haplotype frequency estimates of AGT gene in patients with sick sinus syndrome and controls.
Table 3.
Distribution of genotypes and alleles of Cx40 and AGT in patients with and without sick sinus syndrome.
Figure 4.
Comparison of the transcriptional activities of reporter constructs containing AGT proximal promoter polymorphisms in HepG2 cells.
Transcriptional activities are presented as a ratio of the activity of the p(-6G) promoter-luciferase construct. pGL3 represents the blank vector, which does not contain any AGT promoter sequence. Values are presented as means±SE.
Figure 5.
EMSA results of comparison of the binding affinities of biotinylated oligonucleotides.
(A) Lanes 1 and 2 indicating the mobilities of labeled, biotinylated oligonucleotides (A33 and G33) with nuclear extracts. A stronger shift was observed in the G33 in comparison to the A33 oligonucleotide. Lanes 3 and 4 showing the competition experiments. The unlabeled oligonucleotides completely inhibited the specific binding complex of biotin-labeled A33 and G33 probes to nuclear extract. (B) Lanes 1 and 2 indicating the mobilities of labeled oligonucleotides (G23 and A23) with nuclear extracts. A stronger binding complex was observed in G23 in comparison with the A23 oligonucleotide. The unlabeled oligonucleotides completely inhibited the specific binding complex of biotin-labeled G23 and A23 probes to nuclear extract in lane 3 and 4. The arrow points to the specific nuclear complex, which binds with the labeled, biotinylated oligonucleotides. C = competitor.
Table 4.
Effects of AGT G-6A polymorphism on heart rate and PR interval measured by ECG recordings in control patients without SSS.