Stable stem-loop and cruciform DNA stuctures: isolation of mutants with rearrangements of the palindromic sequence at the Simian Virus 40 replication origin

With the objective of generating DNA molecules that form stable stem-loop structures or cruciform structures in solution, we have altered the palindromic sequence at the Simian Virus 40 (SV40) replication origin. These alterations include: deletion of 18 of the 27 base pairs (bp) in the 13-bp inverted repeat; deletion of 26 of the 27 bp; substitution of the entire 27 bp with a totally different 26-bp sequence containing a 13-bp inverted repeat; and substitution of the 27 bp with an 8-bp sequence containing a 4-bp inverted repeat. The DNA from these mutants was purified. Mutant DNAs were hybridized to wild-type SV40 DNA or to each other, and the heteroduplexes were purified. The predicted structures were verified by S1 and restriction endonuclease digestion. The mutants - the heteroduplexes have not been tested - are capable of complementing tsA mutants and/or transforming mouse cells.