Capillary electrophoresis as a method to study DNA reassociation

Biotechnology Progress
Y LiM J Sepaniak

Abstract

To develop analytical methodology to assess the genetic complexity of a DNA sample, capillary electrophoresis with laser-induced fluorescence detection is used to monitor the annealing process of DNA samples. Coated columns are filled with an entangled polymer solution shown to optimally separate DNA through size-selective capillary electrophoresis. DNA samples are denatured by heating in a boiling water bath for approximately 10 min and then cooled to approximately 25 degrees C below the melting point of the DNA sample to initiate the reassociation process. The DNA is detected by means of the laser-induced fluorescence of intercalated ethidium bromide, which produces a substantially greater signal for double- versus single-stranded DNA. The rate of reassociation is dependent upon the rate at which complimentary strands of DNA encounter each other and the degree of repeating base sequences in the sample (hence, the diversity of the DNA). Experimental parameters also influence the reassociation rate. The effects of salt concentration and incubation temperature are presented. Traditional plots of C(o)t (C(o) = DNA concentration and t = reassociation time) versus % recovery of double-stranded DNA signal are generated for PhiX 174 ...Continue Reading

References

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Citations

Sep 19, 2002·Journal of Chromatography. a·Phyllis R BrownSusan E Geldart
Jul 9, 2008·DNA and Cell Biology·Soheila KashanianFereshteh Abasi Tarighat
Dec 15, 2010·DNA and Cell Biology·Hao-Yu ShenJia Li
Jan 14, 2010·Tissue Engineering. Part a·Frank Xue JiangNoshir A Langrana
Jul 14, 2010·Biochemistry. Biokhimii︠a︡·Cheng-Yong ZhouPin Yang
Feb 21, 2007·Biochemistry. Biokhimii︠a︡·Zhou Cheng-YongYang Pin

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