Abstract: Methods are provided here for an easy and reliable non-fluorescent approach to detecting and monitoring PCR amplification. This includes monitoring PCR amplification by using magnesium-sensitive colorimetric dye. The dye is visually detectable color dye. Compositions are described here that comprise a magnesium-sensitive colorimetric dye, a buffer, dNTPs, magnesium ion and DNA polymerase.

Abstract: Site-specific mutation in methylated circular stranded DNA molecules is conferred by mutagenic primer pairs and methylase deficient Escherichia coli. The mutagenic primer pairs are complementary at 5? end or 3? end, or completely complementary to each other. Firstly, mutagenic primer pair is annealed to opposite strands of the methylated circular double-stranded parent DNA molecules. Then, polymerase chain reaction is performed by using unmethylated dNTPs to create unmethylated mutagenized double-stranded daughter DNA molecules. Finally, the reaction mixture of the methylated parent DNA molecules and unmethylated mutagenized daughter DNA molecules is transformed into a methylase deficient E. coli. The replication of methylated parent DNA is inhibited in methylase deficient host cell. In contrast, the unmethylated daughter DNA, which contains the desired mutation, are efficiently replicated in methylase deficient host cell and recovered thereafter.