[Use of phage display for detecting single-nucleotide differences in genes].
Experimental strategy has been developed for selection of mismatched DNA binding phages from library of E. coli f1 filamentous phages carrying random peptide inserts on the surface of bacteriophage particles. The strategy is based on the use of phage display technique, DNA heteroduplexes (with single nucleotide variations), and paramagnetic beads. DNA heteroduplexes have been obtained from biotin-labeled PCR product. During the first stage the phage particles were incubated with DNA heteroduplexes possessing mismatched nucleotides. The next step after elimination of free phages and separation of bound phages from DNA heteroduplexes was subtraction of phages binding with DNA heteroduplexes (without mismatched nucleotides). Phages selected by this method were capable of discriminating DNA heteroduplexes with single nucleotide variations from DNA homoduplexes. Phages immobilized on solid base retain their activity and specificity, and therefore can be used for developing a new screening automated method for detecting point mutations and gene polymorphism.
Denisov
S G
SG
Institute of Cytology and Genetics, Siberian Division, Russian Academy of Sciences, Novosibirsk.
Beliavskaia
V A
VA
Voevoda
M I
MI
D001483:Base Sequence; D003001:Cloning, Molecular; D003090:Coliphages / Q000235:genetics*; D004274:DNA, Recombinant / Q000235:genetics; D004926:Escherichia coli / Q000821:virology; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic
D004274:DNA, Recombinant; D009692:Nucleic Acid Heteroduplexes