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The assessmentof the effect of acrylamide concentration on the diffusion of biomolecules in a gel
Zubik A. N.
1, Bulyanitsa A.L.
1, Rudnitskaya G.E.
1, Evstrapov A.A.
1
1Institute for Analytical Instrumentation of the Russian Academy of Sciences, Saint Petersburg, Russia
Email: tunix@yandex.ru
A highly accurate method for quantifying target DNA molecules is the implementation of an amplification reaction in a thin layer of polyacrylamide gel.The gel impedes the diffusion of DNA molecules, which leads to the concentration of amplification products (amplicons) around the original target molecule and the formation of molecular colonies. Increasing the concentration of acrylamide produces denser gels, which reduces colony size. Reducing the size of colonies while maintaining the effective gel area allows you to record a larger number of colonies, increasing the dynamic range of the method. To study the effect of acrylamide concentration on the diffusion of amplicons (DNA fragments obtained as a result of the analysis) in the gel, a simple method was used to estimate the diffusion coefficient based on recording the moving boundary of fluorescently labeled biomolecules. An experimental assessment of the effect of acrylamide concentration on the diffusion of synthetic oligonucleotides and amplicons in the gel shows that when the acrylamide concentration increases from 7 to 10%, the estimated diffusion values change by approximately 2 times. Keywords: microfluidic chip, polyacrylamide gel, diffusion coefficient, amplicon.
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