Annealing temperature is one of the most important
parameters in multiplex PCR assay. Optimal temperatures
are often lower than the melting temperature of the primers.
Annealing temperature given by Dorch9
for a primer specific
for 16SrRNA gene of A. hydrophila was 62°C,
Wannet et al.10 for a primer specific for 16SrRNA gene of
Y. enterocolitica was 62°C and by Blanco et al.8 for a
primer specific for uid R gene of E. coli as 57°C.