Biophysical Society Thematic Meeting| Santa Cruz 2018

Genome Biophysics: Integrating Genomics and Biophysics to Understand Structural and Functional Aspects of Genomes

Poster Abstracts

22-POS Board 22 Quantitative Gel Electrophoresis for Fragment Library Preparation Riccardo Ziraldo 1 , Stephen D. Levene 1 , Massa J. Shoura 2 , Andrew Z. Fire 2 . 1 The University of Texas at Dallas, Richardson, TX, USA, 2 Stanford University, Stanford, CA, USA. Gel electrophoresis has been a convenient and cost-effective tool for the characterization of biopolymers. However, many details of the underlying physical mechanisms that govern electrophoretic mobility as a function of molecular size, shape, and mechanical flexibility remain poorly understood. In the case of species that migrate as distinct, well-resolved bands, the band’s integrated intensity is both a qualitative and quantitative measure of the mass of a particular molecular species in the sample. For gel-electrophoretic patterns that approach that of a continuum, however, extracting useful information about size distributions requires careful assumptions regarding the underlying sample composition. We have developed a versatile and tunable software tool to fit univariate functions to gel-mobility profiles using minimal information from duplex-DNA standards having both simple and complex size distributions. We apply the tool to quantitative characterization of fragment libraries for next-generation DNA sequencing (NGS), optimization of which depends on knowledge of the size distribution of randomly fragmented DNA samples. We show that the tool provides reasonable average values for fragment size when selecting a portion of the continuous profile for library preparation. In addition to characterizing fragment distributions for library preparation based on a line-profile continuum, the tool is applicable to quantifying complex topologies of circular DNA.

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