COMPARISON OF DIFFERENT METHODS TO PRODUCE SINGLE-STRAND DNA FOR IDENTIFICATION OF CANNED TUNA BY SINGLE-STRAND CONFORMATION POLYMORPHISM ANALYSIS

By using single-strand conformation polymorphism (SSCP) analysis of th ree amplicons of the cytochrome b gene obtained by the polymerase chai n reaction (PCR) it was possible to differentiate between various spec ies of tunas and bonitos processed as canned fish. Four different tech niques were used to produce single-strand DNA (ssDNA): (i) Denaturatio n of double-strand DNA (dsDNA) by formamide and alkali, (ii) two-step asymmetrical PCR, (iii) onestep asymmetrical PCR, and (iv) exonuclease digestion of the phosphorylated strand of dsDNA. The technique render ing optimal results depended on the type of amplicon (i.e. the sequenc e).