**Abstract**

In our study, we used PCR to clone *papA*, *papEF*, *papG* and *F17G* genes of *Escherichia coli* isolated from faecal samples of dogs with diarrhoea. Annealing temperature of 55°C was used in the PCR. Nucleotide sequence analysis of 26 cloned PCR products showed that in PCRs with *papA* primers, six out of eight obtained PCR products were false due to non-specific binding of the forward primer on both DNA strands; in PCRs with the *papEF* primers, all seven obtained PCR products originated from specific binding of the forward primer on the 3′ → 5′ DNA strand and non-specific binding of the forward primed on the 5′ → 3′ DNA strand; and in PCRs with the *F17G* primers, four out of eight obtained PCR products were false due to non-specific binding of forward and reverse primer. The anticipated annealing sites for non-specific primer binding in analysed nucleotide sequences are presented. In the case of PCR products obtained with *papG*-specific primers, all PCR products were amplifications of the *papG* sequence. When the annealing temperature of *papA* PCRs was raised to 60°C, all obtained PCR products were amplifications of the correct DNA sequences.

**Keywords:** primer binding, annealing temperature, sequence analysis, *Escherichia coli*, adhesin, P-fimbriae, F17-fimbriae
