Authors: S.M. Azimi, G. Nixon, J. Ahern, W. Balachandran
Affilation: Brunel University, United Kingdom
Pages: 46 - 49
Keywords: DNA extraction, magnetic mixer, magnetic bead
This paper introduces a new DNA extraction and purification method from whole blood. Mixing and separation steps are performed using functionalised superparamagnetic beads suspended in cell lysis buffer in a circular chamber that is sandwiched between two electromagnets. Non-uniform nature of magnetic field causes temporal and spatial distribution of beads within the chamber. This process efficiently mixes the lysis buffer and whole blood in order to extract DNA from white blood cells. Functionalized surface of the magnetic beads then attract the exposed DNA molecules. Finally, DNA-attached magnetic beads are attracted to the bottom of the chamber by activating the bottom electrode. DNA molecules are extracted from magnetic beads by washing and re-suspension processes. In this work a 25µl PMMA chamber, 500µm in depth and 8mm in diameter was fabricated to purify DNA from spiked bacteria cell culture into the whole blood sample using Promega Magazorb DNA purification kit. Spiked bacterial cell culture was prepared from a gram positive; B. Subtilis and a gram negative, E. coli cell culture in different concentrations. The lysis was performed in 5min incubation at 56°C followed by 5min incubation for binding process. Thermal stability was generated and controlled by the same external coils which were used for continuous mixing and on-chip clamping of the magnetic beads in the chamber. The yield and purity of extracted DNA was measured using real-time PCR and spectrophotometer respectively. The preliminary result indicated successful lysis of E. coli cell and a weak lysis of B. subtilis (~18%) in adopted protocol. The efficiency of lysis on B. subtilis was improved significantly by applying the two-step protocol.
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