SYNTHESIS OF FLUORESCENCE-LABELLED NUCLEIC ACIDS

摘要

The present invention provides a method for synthesizing nucleic acids of at least 200 or 500 bases length, including strands of up to several kb in length, wherein at least one of the four NTPs is completely substituted by a fluorescent-labelled ribo- or deoxyribonucleotide, r- or dNTP-X. The method includes the use of dNTP-X compounds having the nucleotide base moiety covalently joined to the fluorophore by a linker chain of 8 to 12 atoms length, the use of a processive DNA polymerase, and the use of single-strand DNA binding protein in the reaction mixture. Novel compounds suitable for use in synthesizing fully labelled fluorescent DNA and a kit for carrying out the method are also disclosed. The method of the present invention is useful to provide fully labelled fluorescent RNA or DNA for sequence analysis, for histochemical fluorescent labelling and for micro-analytical techniques where highly fluorescent RNA or DNA of specified sequence is desired.