DNA Processing

DNA is nothing but a deoxyribonucleic acid. This blog will explain the DNA extraction from blood. A phlebotomist should always perform the blood sample collection with familiarization of protocol.

Below are the steps and pre-requisite you should follow while extracting the DNA samples from the Whole Blood.

• The place where you are performing DNA extraction should be clean and has been decontaminated. This can be done with freshly made 10% bleach solution allowed to sit for fifteen to thirty minutes and then wipe away any residual bleach, followed by rinsing with clean water and finally spraying with 70% ethanol.
• Make sure you wear gloves throughout the extraction process and change if coming into contact with other surfaces that might be contaminated.
• Check all equipment and consumables you are going to use are in place and that the buffers in the kit have been prepared.
• The heating block must have been set at 56 degrees centigrade and ready to use before commencing with DNA extraction.

• Mix the blood by inverting the tube approximately ten times to ensure you have a homogenous sample vortex of the protein ASX, followed by a quick centrifugation step about 10 seconds.
• The heating block’s microtubes have been set at 56 degrees centigrade and incubate the samples for 10 minutes at the end of the incubation period.
• Remove the samples from the heating block and centrifuge to remove any droplets from the lids. Add 200 microliters of absolute ethanol to each microtube and mix each sample by pulse vortexing for about 15 seconds.
• Centrifuge the microtubes prepare a mini spin column for each sample — spin columns ensure that each spin column corresponds to the sample ID.
• Transfer the mixture to the spin column taking care not to wet the spin column’s room. Close the spin column and centrifuge at 6000 G that’s about 8,000 rpm for one minute at the end of the centrifugation period. All the filtrate should be in the collection tube transfer the spin columns now contain the DNA to clean collection tubes and discard the collection tubes containing the lysate.
• Open the spin column and add 500 microliters of aw1 buffer, taking care not to wet the room. Remember to change your tip between each sample, close the caps and return the samples to the centrifuge and centrifuge for one minute again at 6000 G at is 8,000 rpm.
• At the end of the centrifugation period, check that all the filtrate is in the collection tube. If there is still some in the spin column, repeat that centrifugation step discards the collection tub,e, and transfer the spin columns to clean collection tubes.
• Open the lid and add 500 microliters of a w-2 without wetting the rim of these spin columns. Close the cap and centrifuge at full speed that’s 20,000 G around 14000 rpm for three minutes. Discard the filtrate from the collection tube into a waste container as the collection tube will be reused. Dry the rim of each collection tube on a paper towel and replace the spin column centrifuge at full speed for one minute. This ensures that all residual wash popper and ethanol are removed, which could affect downstream processes.
• Label clean 1.5 ml microtubes one for each sample and transfer the spin columns to each corresponding micro tube the DNA that was on the spin column will be eluted from the column to the AE buffer add 200 microliters of AE buffer to each spin column remembering to change tips between samples incubate for one minute at room temperature centrifuge at 6000 G that’s 8,000 rpm the filtrate transferring this back onto the spin column. Repeat the incubation and centrifugation steps; this repeat dilution step optimizes a DNA yield.

References:

DNA extraction from Blood

Difference Between DNA and RNA

DNA TESTING PROCEDURES