A NOVEL FLUOROGENIC SUBSTRATE FOR THE USE OF NUCLEIC-ACID HYBRIDIZATION

The phosphates derived from newly synthesized 3'-O-aralkyl-5-(4-biphen ylcarboxamido)fluorescein derivatives were examined for the alkaline p hosphatase-linked fluorescence assay of the membrane-bound DNA. lambda DNA was detectable to the amount of 5 fg in the assay using phosphory lated nylcarboxamido)-3'-O-(1-naphthyl)methylfluorescein (BNFP). The s pots gave distinguishably clear fluorescence without diffusion and non -specific adsorption. In the Southern blot hybridization, 0.1 pg of DN A could be detected. Detection of two different DNAs on a single blot was successfully performed by using two different fluorogenic phosphat es, Phosphrorilated N-(2-biphenyl)-3-hydroxy-2-naphthalenecarboxamide (HNPP) and BNFP. The fluorescein derivatives were also examined as sub strates for horse radish peroxidase.