Effects of Polyols, pH and Electrolyte Concentrations in TBE Buffer o Separation of Double Strand DNA Fragments by Capillary Electrophoresis

Ren, Jicun; Fang, Nenghu; Wu, Dan

doi:10.2116/analsci.18.469

Cited by 2 publications

(1 citation statement)

References 14 publications

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“…Tris is commonly used as a biological buffer. 29–31 The effect of adding TBE in the silk/DNA mixture on DNA preservation was evaluated. The samples were stored at different temperature for 24 hours before being analyzed.…”

Section: Resultsmentioning

confidence: 99%

DNA preservation in silk

et al. 2017

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The structure of DNA is susceptible to alterations at high temperature, pH, irradiation and exposure to DNase. Options to protect and preserve DNA during storage are important for applications in genetic diagnosis, identity authentication, drug development and bioresearch. In the present study, the stability of total DNA purified from human dermal fibroblast cells as well as plasmid DNA was studied in silk protein materials. The DNA/silk mixtures were stabilized on filter paper (silk/DNA+filter) or filter paper pre-coated with silk and treated with methanol (silk/DNA+PT-filter) as a route to practical utility. After air-drying and water extraction, 50–70% of the DNA and silk could be retrieved and showed a single band on electrophoretic gels. The 6% silk/DNA+PT-filter samples provided improved stability in comparison to 3% silk/DNA+filter and DNA+filter for DNA preservation, with ~40% of the band intensity remaining at 37 °C after 40 days and ~10% after exposure to the UV light for 10 hours. Quantitative analysis using the PicoGreen assay confirmed the results. The use of Tris/Borate/EDTA (TBE) buffer enhanced the preservation and/or extraction of the DNA. The DNA extracted after storage maintained integrity and function based on serving as a functional template for PCR amplification of the gene for zinc finger protein 750 (ZNF750) and for transgene expression of red fluoresence protein (dsRed) in HEK293 cells. The high molecular weight and high content of crystalline beta-sheet structure formed on the coated surfaces likely accounted for the preservation effects observed for the silk/DNA+PT-filter samples. Although similar preservation effects were also obtained for lyophilized silk/DNA samples, the rapid and simple processing available with the silk-DNA-filter membrane system makes it appealing for future applications.

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Section: Resultsmentioning

confidence: 99%