Title: DNA Extraction Abstract: DNA extraction is a procedure to collect DNA For the purpose of genetic testing, body

identification, and forensic analysis. It provides student to gain experience and knowledge of DNA. Simple protocol and visualization was devised. DNA is the largest known molecule. A single unbroken strand can contain millions of atom. When DNA is released from a cell, it breaks up into tiny strands. These tiny fragments are negative charge which causes it to adhere to one another. Criteria were followed to evaluate the protocol. In this experiment human DNA is collected from the cells of buccal mucosa due to its availability and can be collected in a non invasive manner. Breaking the cells open, removing membranes, removing proteins, removing RNA and precipitating the DNA with alcohol.

Objective: To follow the protocol provided and evaluate by following the procedures.

Related Reading Literature

Article 1: Mitochondrial DNA analysis of 114 hairs measuring less than 1 cm from a 19year-old homicide.

Mitochondrial DNA analysis is typically applied to degraded skeletal remains and telogen or rootless hairs. application of the method to very small hairs less than 0.5 cm from an age-matched and -challenged sample set are lacking. Degraded samples were amplified with mini-primers, Crime scene hairs that are both aged and small are often judged to be unsuitable for either hair microscopy or DNA analysis. This study of age-matched challenged small hairs indicates that even the smallest probative crime scene hairs are suitable for mitochondrial DNA analysis and can provide useful data. Even it is said to be not an absolute basis for forensics and in molecular study to prove. It has property that allow biologists to elucidate evolutionary relationship among species can prove the relatedness during examination.

Article 2: Long-term room temperature preservation of corpse soft tissue: an

approach for tissue sample storage.

Disaster victim identification is the most difficult challenges in forensics to uncover. Collected DNA samples from human for identification are frequently store at room temperature to stop the early degradation of the remains. Scientist had tried to experiment the longevity of samples if exposed to different nature. First, by preserving the sample with salt; Second, by preserving the sample with no additional substances; third, by preserving the samples with garden soil. Fortunately, all worked. Therefore, these procedures are additional storage samples when preservation of DNA samples are required for DNA analysis and PCR.

Article 3: Differential DNA extraction of challenging simulated sexual-assault samples: a Swiss collaborative study.

In cases of sexual assault, collecting

gynecological evidence is challenging. Quantity of male DNA is usually prevented due to the large quantity of female material. The solution for this problem is differential DNA extraction. But this does not guarantee reliable results. Samples of epithelial cells from buccal swabs and semen are mixed. They are processed in the laboratory without differential lysis. We used qPCR to assess the quantities of male and female DNA recovered in the sperm and epithelialcell fractions after differential DNA extraction. DNA profiling was used to determine whether DNA characteristics of the male contributor could be retrieved. an initial lysis that is too strong can break not only the epithelial-cell membranes, but also some of the spermatozoa, leading to the displacement of a significant amount of male DNA into the epithelial-cell fraction. On the other hand, an initial lysis that is too weak may not break all the epithelial-cell membranes leading to a contamination of epithelial cells in the sperm fraction. Though the reliability is sometimes undertaken, but the effectiveness of these methods are helpful.

Materials and Methods

Warm water bath concentrated salt solution Centrifuge resuspension buffer buccal swabs ethanol micropipettors isoprophyl alcohol sample tubes solution lysis

Collect cheek cells

Drag the swab over to the inside of the test subjects mouth to collect a sample.

Place the swab into the Eppendorf tube

Burst cells to release DNA Cut the buccal swab stick so the collected sample will remain in the tube and to be able to close the tube. Using the micropipettor, add some lysis solution to the tube. Place the tube into the warm water bath.

Seperate DNA from protein and debris After the cells have stayed in the warm

water bath, remove the swab from the tube. Add some concentrated salt solution from the tube Place the tube in the centrifuge. Balance the centrifuge by placing another tube containing water at the opposite to the tube with DNA solution. Remove the tube in the centrifuge. Use the micropipettor to carefully remove the top liquid containing DNA and place it in the clean tube

Isolate concentrated DNA Add some isoprophyl alcohol to the tube. Invert the tube several times to mix isoprophyl alcohol and DNA. Place the tube in the centrifuge

Remove the liquid for the DNA to dry. Dried DNA can be redissolved at any solution of your choice. *Freeze the DNA for future use.

Reading and Discussion

In DNA extraction you need to isolate some DNA from human test subject for genetic testing, body identification and forensic analysis. DNA is typically the first step in longer laboratory process. The DNA needs to be purified away from proteins and other cellular contaminants. We need cells because that is where DNA is. Inside every cell in our body has a nucleus and inside its nucleus has two meters of DNA. Collection of cells from the test subject is the initial procedure. We can either collect a blood sample or a check swab. In our case we chose the check samples. The skin on the inside of our mouths loses millions of cells every day. Different steps are followed to in order to purify DNA from check swab. Proper equipment and right amount of materials are very important to obtain accurate results. First off, drag the buccal swab in the inside check of the subject. Buccal swab needs to be sterile to avoid contamination. When the buccal swab in laced inside the Eppendorf tube, youll notice that the swab is covered with hundreds of tiny check cells. Inside the check cells is the nucleus and inside the nucleus in the DNA. Second, burst the cells open to release the DNA. End of the swab must be cut so we can close the tube. Lysis solution is added. Lysis means to separate. The lysis solution added contains two important ingredient: detergent and an enzyme protienase K. The detergent disrupts the cell membrane and nuclear envelope, causing the cell to burst open and release DNA. The DNA is still wrapped very tightly around proteins called histones and the protienase K cuts apart the histones to free the DNA. There are different type of lysis buffer that can be applied depending on what analysis the cell lysate will be used. Generally, lysis buffers contain trisHCl, EDTA, EGTA, SDS,deoxycholate, tritonX and/or NP-40. In some cases the buffer may also contain NaCl. In studies like DNA fingerprinting the lysis buffer is used for DNA isolation. Dish soap can be used in a pinch to break down the cell and nuclear membranes, allowing the DNA to be released. The enzymes in the soap are breaking down the lipid molecules of the cell and nuclear membranes, releasing the contents of the cell, including the DNA. These enzymes in the soap are what break down grease while washing dishes.

Third is the separation of DNA from protein and debris. The cells have stayed long enough in the water bath for the DNA to be freed from the cells and we have removed the cells from the tube. Concentrated salt is now added. The salt causes proteins and other cellular debris to clump together. Salt provides DNA with a favorable environment. It contributes positively charged atoms that neutralize negative charge of DNA. By placing the tube inside the centrifuge, heavy clumps of proteins and cellular debris sink to the bottom of the tube, while the strands of DNA remain distributed through the liquid. At home, we can use the blender. It still exhibits the same function like what the centrifuge does. It breaks down the cell wall. In order to remove the top liquid, the micropipettor is used. It contains DNA and transfers it in the clean tube, while the proteins and other cellular debris are left behind. Fourth is the isolation of concentrated DNA. Addition of isopropyl alcohol is used. The tube is inverted upside down for several times. DNA is not dissolved in alcohol so it comes out from the solution or it forms as precipitates. It is less dense than water or cell scum which is what settles to the bottom of the glass, so it floats up into the alcohol layer, where you see it as a snotty, stringlike substance, with small bubbles formed on it. The tube is place in the centrifuge again for the purpose of sinking the DNA at the bottom of the tube. Once the liquid is removed, the DNA is allowed to dry. It can be re dissolved at any solution of choice. It can be stored in the cryogenic freezer for future use.

Conclusion & Research DNA extraction is unique that can determine analysis on human and animal [Link] is because our DNA is so unique that DNA testing can yield some very accurate results. DNA testing can identify criminals who were present at a crime, or it can prove the guilt or the innocence of individuals suspected of committing a [Link] DNA testing determine human and animal

DNA, it tests samples of DNA extracted from the elements for healthpurposes, it determine steroid or alcohol and drug use and so much more. Legal DNA testing is permissible as evidence in courts. DNA samples can be taken from anywhere on the body; skin cells, hair follicles, semen and more. DNA samples are important to answer many of the questions we need answered in our everyday lives. Parental DNA testing can determine if missing children who are found many years later are related to a couple, while forensic DNA testing is used in rape cases, and for paternity issues.

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