Experiment

An experiment has been performed at the Institute of Physics of the UNAM to test the proposed thermodynamic dissipation theory for the origin of life. For more details than can be presented here, please consult the article;

Michaelian, K., Santillán Padilla, N. DNA Denaturing through UV-C Photon Dissipation: A Possible Route to Archean Non-enzymatic Replication, bioRxiv [Biophysics] (2014). doi: http://dx.doi.org/10.1101/009126

The experiment consists of a deuterium lamp which emits in the UV with emission from 220 nm to 500 nm, an aqueous solution of RNA or DNA in a quartz container  (cuvette) in a temperature controlled holder connected by means of a fiber optic to a spectrometer.

ExperimentFoto

One of the biggest problems in solving the riddle of the origin of life is how denaturing (the first step in replication) could have occurred during the Archean without the complex enzymes life uses today to denature.

The purpose of the experiment, therefore, was to determine if UVC light itself could denature DNA and, if so, measure quantitatively the amount of denaturing as a function of solution temperature.

The method was to shine UV light on the DNA solution at a fixed temperature and with the spectrometer measure any hyperchromic effect in the absorption (when DNA denatures it absorbs more UVC light since the bases are no longer tightly stacked and thus present a larger cross section to the light).

We can compare double strand with single strand DNA. Light-on periods were 1 hour, followed by light-off periods of also one hour. The results are given below for different temperature runs.

Fig1

Absorption spectrum of 25 bp synthetic DNA showing the hyperchromic effect of increased absorption as the temperature is raised due to denaturing. The lower curves are the different spectra obtained for 3 °C bins centered at the specified temperature.

Fig3

Average of the extinction integrated between 255 and 265 nm for double strand (left panel) and single strand (right panel) at the fixed different temperatures. The increase in extinction can be seen for the double strand DNA during the light-on periods, especially at the higher temperatures. This we attribute to UVC-light induced denaturing of DNA. For the single strand, a slight decrease in extinction can be seen over the light-on periods which we attribute to cyclobutane pyrimidine dimer formation.

Fig4

The difference spectra for UVC light-induced denaturing during the one hour light-on periods at the given fixed temperatures. For the double strand DNA (solid curves) there is an increase in extinction (due to light-induced denaturing) while for the single strand DNA, there is a decrease in extinction which we attribute to CPD dimer formation.

Clearly we do see UVC light-induced denaturing in our data. We have quantified it in the following graph as a function of temperature.

Fig7

Light induced denaturing per hour for double strand (solid line) and single strand (dashed line). For the single strand DNA the decrease is due to CPD formation.

Fig8

Effective complete denaturing rate for 25 bp synthetic DNA as a function of temperature.

More details of the experiment and its analysis can be found in the following article;

Michaelian, K., Santillán Padilla, N. DNA Denaturing through UV-C Photon Dissipation: A Possible Route to Archean Non-enzymatic Replication, bioRxiv [Biophysics] (2014). doi: http://dx.doi.org/10.1101/009126

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