A heteroduplex-preferential T sub(m) depressor for the specificity-enhanced DNA polymerase chain reactions

A macrocyclic tetraamine zinc( II) complex appended with two quinoline groups, Zn super(2+)-1, 7-bis(4-quinolylmethyl)-1, 4, 7, 10-tetraazacyclododecane (Zn super(2+)-Q sub(2)-cyclen), was successfully used as a novel additive to suppress nonspecific products in DNA polymerase chain reaction (PCR). In the presence of Zn super(2+)-Q sub(2)-cyclen, the T sub(m) drop of 20-bp heteroduplexes containing a noncomplementary basepair was greater than that of the corresponding homoduplex (i.e. primer DNA). Here, we applied such preferential DNA melting to a specificity-enhanced PCR using micromolar concentrations of Zn super(2+)-Q sub(2)- cyclen. We demonstrated the selective amplification of target DNA fragments (i.e. the human heart sodium channel Na sub(v)1.5 gene) from genomic DNA or a cDNA library. The optimum condition for the specificity-enhanced PCR could be determined in the concentration range of 1-50 mu M of Zn super(2+)-Q sub(2)-cyclen.