Gene alteration using TFOs
Triple helix forming oligonucleotides (TFOs) recognize and bind sequence in duplex DNA and have received attention due to their potential for targeting specific genomic sites. Three different mechanisms of triplex mediated interference with DNA information processing are shown in. DNA cross-linking reagent psoralen has been used for linking to prepare modified TEOs, that may be directed to a specific site of a gene. For instance, in Chinese Hamster hypoxanthine phosphoribosyl transferase (Hprt) gene, in exon E4 and immediately adjacent to the smallest exon E5, there is a polypyrimidine:

It has been shown that replacement of a base in a TFO with an intercalator (e.g. pyrene or acridine) at Ta inversion site stabilizes triplexes. When TFOs carrying intercalators at the inversion site were used with cells that were electroporated and UV-A treated, large number (77% clones) of deletions (4-50 bases long) and point mutations (4%) were deteched. Point mutations were located mainly at the cross-linking site.

The TEOs thus are capable of delivering psoralen at the target sequence and enhance the frequency of mutagenesis few hundred fold relative to spontaneous mutations. No increase in mutation frequency was observed, when either TFOs without psoralen were introduced in the cell or when psoralen-
TFO were not protected against nucleases.