Annealing Properties of Oligoribonucleotides Derived from T5 DNA-directed RNA Compared to Those from T2 and T7 DNA-directed RNAs

Abstract

Abstract RNA made in vitro on a T5 phage DNA template was partially digested with ribonuclease T1 and fractionated according to chain length. The oligoribonucleotides of various chain lengths were tested for their annealing at different temperatures to denatured DNAs immobilized on membrane filters. As observed before for the T-even and other T-odd phage DNAs, a chain length of 10 or more ribonucleotides is required to form a ribonuclease-resistant complex with denatured T5 DNA in the absence of Mg2+. The shortest oligoribonucleotides capable of forming a complex with denatured T5 DNA display a high degree of specificity for species: these oligoribonucleotides do not anneal to either T2 or T7 DNA. As expected, the temperature for optimum complex formation and the melting temperature of the oligoribonucleotide-denatured DNA complex increase with the chain length of the oligoribonucleotide. With increasing chain length, the average base composition increasingly reflects the base composition of the parent RNAs. For the same chain length, T5-produced oligoribonucleotides having a G+C content intermediate between those of T2- and T7-produced oligoribonucleotides form complexes having stabilities intermediate between those of the latter. There are linear relationships between the melting temperatures and the G+C contents for oligoribonucleotides of a defined chain length. This makes it possible to characterize oligoribonucleotide-denatured DNA complexes as a function of both the chain length and the base composition of the oligoribonucleotide.