For many of us, Doty's major contribution to molecular biology was his study of DNA renaturation with his long-time post-doc and collaborator, Julius Marmur (1926 - 1996)1, a graduate of McGill University in Montréal, Canada. The paper that most of us remember is Marmur and Doty 1962: "Thermal Renaturation of Deoxyribonucleic Acids." This was the first time that the renaturation of complex DNA had been studied in detail and the results have led to many of the common techniques in use today.
Deoxyribonucleic Acid (DNA)DNA is normally double-stranded and the two strands are held together by weak interactions, chiefly stacking interactions and hydrogen bonds [DNA Denaturation and Renaturation and the Role of Hydrogen Bonds and Stacking Interactions].
The strands can be separated by heating the DNA to the "melting temperature." What Marmur and Doty showed is that the two strands could, under the appropriate conditions, come back together to form a fully base-paired double helix. This is called renaturation.
Single-stranded DNA absorbs much more UV light than double-stranded DNA so you can follow denaturation and renaturation by simply measuring UV light absorbance in a spectrophotometer. The figure shows that the DNA molecules "melt," or denature, as the temperature is raised. When the temperature was lowered to 67°C they could follow the formation of double-standed DNA over time. For virus and bacteriophage DNA (solid triangles and circles) most of the DNA seemed to renature in about one hour. In the case of bacterial DNA (D. pneumoniae, open circles) the process was much slower and for cow DNA (open triangles) hardly any renaturation was seen in 100 minutes.
Marmur and Doty showed that the optimum temperature for renaturation was about 25°C below the melting temperature. This was the temperature that prevented inappropriate base pairing with mismatches while still allowing true complementary strands to come together. Renaturation was more efficient in the presence of high concentrations of salt (0.3M NaCl) because the negatively charged phosphate groups had to be shielded.
Did the renaturated DNA truly represent re-formation of intact DNA molecules? Marmur and Doty proved this by showing that the pneumoniae DNA regained the ability to transform bacteria.
DNA Denaturation and Renaturation and the Role of Hydrogen Bonds and Stacking Interactions].
The second variable is DNA complexity as shown in the first figure. Cow DNA is much more complex than bacterial DNA so it takes much longer for the complementary strands to find each other in a solution of cow DNA than in a solution of bacterial DNA at the same concentration. By studying the rate of DNA renaturation as a function of time and initial concentration (Co) you can determine the genome size and how much of the DNA is single copy and multi-copy. This is the basis of Cot curves and it led to the discovery of repetitive DNA and junk DNA in eukaryotic genomes.
There are many other common techniques that rely on the pioneering work of Marmur and Doty. They include in situ hybridization, Southern blots, and the priming of PCR reactions.
On a personal note, Paul Doty is my scientific grandfather. I got my Ph.D. in 1974 with Bruce Alberts and he got his Ph.D. in 1965 with Paul Doty.
1. See the obituary in Nature by Paul Doty: Julius Marmur (1926 - 1996).
Marmur, J. and Doty, P. (1962) Thermal Renaturation of Deoxyribonucleic Acids. J. Mol. Biol. 3:585-594. [PubMed] [doi: 10.1016/S0022-2836(61)80023-5]