A buccal swab collects skin cells (stratified squamous epithelial cells) from the inside of a cheek. Scissors are used to cut off the end of the swab and place it into an eppendorf tube. Micro pipette is used to add lysis solution, which is a combination of detergent, which breaks apart the cell and nuclear membranes, and proteinase k, which cuts away histones. This tube with lysis is added to a warm water bath, facilitating the action of the lysis. We then use a micro pipette to add concentrated salt solution. This solution causes proteins and other cellular debris to clump together. Then, the tube is added to a micro centrifuge, along with a "balancing" water test tube. The fast spinning in the centrifuge causes the clumped debris to fall to the bottom of the tube, and a micro pipette can then be used to remove the top level of liquid and place it in a different tube. Isopropyl alcohol is added to this solution, which separates out clumped DNA into visible form. This tube goes back into the micro centrifuge, and, this time, the DNA falls to the bottom. A pipette is used to remove the top liquid, and the DNA on the bottom is then ready for use or storage.
Electrophoresis
A gel is created by heating agarose along with a salt water solution/liquid buffer in a microwave. This is then added to a mold, and a comb in inserted to create small wells for the DNA samples at one end. It is allowed to cool, and the comb is removed. A buffer is added to an electrophoresis box to allow for conduction of current and to prevent drying out of the mold. The mold is placed into this box. Loading buffer, which allows DNA to be seen, is added to a DNA sample, which is then placed into one of the wells. Standard size DNA (a sample in which DNA length is already known) is added to another well for comparison. A current is then run through the box-the negative charge is at the end with the wells, and a positive charge is on the opposite end. DNA is repelled from the negative charge and begins to make its way to the positive charge. The gel itself is filled with small holes that the DNA travels through to get to the positive charge. Smaller strands are able to move more quickly. Eventually, strands of the same size end up together and clump, which makes them ultimately more visible. Once the process is complete, the mold is removed from the electrophoresis box and placed in an ethidium bromide solution, which stains DNA and makes it visible when it is placed under UV light (accomplished with a UV light box). The clumps appear as bands and their location can be compared with the bands of the known lengths of the standard DNA sample. This gives a rough estimate of the number of base pairs present at each band site.
Electrophoresis can be used to determine paternity. Because each person's DNA pattern is very unique, it shows up in electrophoresis as a unique set of bands, unlikely to be repeated by any but a close relative. Thus, parent and child can be identified by a close match of an electrophoresis reading. This is accomplished by taking the DNA of mother, child, and two possible fathers and putting them through gel electrophorisis. Every band of the child MUST MATCH at least one of the parents. Whatever bands don't match the mother must be found in the father's reading. Therefore, by process of elimination, paternity is determined.
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