Pre Lab Report DNA Fingerprinting PDF

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Introduction Deoxyribonucleic acid, commonly known as DNA is a molecule that contains the genetic material (instructions) utilized in the development, reproduction and functioning of the living organisms. DNA carries the biological instructions which make every species unique. This also makes DNA fingerprinting possible in the field of science as it is used by various experts to identify the biological parents or the crime suspect. The identification in the criminal investigations is done by collecting the tissue evidence to determine the genetic clues which can lead the investigator to the suspect. The results are obtained by cutting the lengthy strand of DNA into shorter segment by using the chemicals or restriction enzymes (enzymes which cut double or single stranded DNA at a restriction site) which cut the segments at the beginning and at the end of the repeating string of nucleotides. These repeating strings are then separated based on the length using the process known as gel electrophoresis followed by introducing a set of the markers (radioactive) to the sample. The markers introduced are the DNA fragments of identified length, with bases that match the code, and attach to the identical length sample fragments. To determine the site of the samples that are marked, photographic film is used which gets dark when exposed to the markers that are radioactive. The photographic film becomes the DNA fingerprint which is used in various areas of study, investigation, research, science, forensic, etc. When comparing two samples, the more segments match or are in common between the sample, the more it is the sample that came from the same person. In this experiment DNA fingerprinting process was used to identify the potential suspect in a certain situation. Six DNA samples were obtained from a crime scene out of which five represented potential suspect. The samples were digested using the restriction enzyme and placed on the agarose gel to check the pattern of the restriction. Results for each sample were observed and analyzed to determine the suspect. Procedure Restriction Digestion A tube containing restriction enzyme mix (labeled ENZ) was stored on the ice. Micro test tubes were labeled as CS (crime scene), S1 (suspect 1), S2, S3, S4, and S5. Thereafter, 10μL of DNA sample was transferred and then mixed from the micro test tubes to the corresponding colored micro tubes. To gather all the liquid on the bottom, the sample was centrifuged and the tubes were incubated (in the micro foam tube holder) at 37 degrees Celsius for 45 minutes. The tubes were then placed in the refrigerator. Agarose Gel Electrophoresis After removing the digested DNA sample from the refrigerator, it was centrifuged. 5μL of loading dye LD was added into each tube using a fresh tip each time and centrifuged. Agarose gel was placed in the electrophoresis apparatus. 275mL 1x TAE buffer was obtained to cover the gel in the electrophoresis chamber. The agarose gel wells were situated such that the wells were near the – (black) electrode and the bottom edge near the + (red) electrode. Thereafter, certain volume per sample was added into the 7 wells of the gel. Lane 1 (M, DNA size marker)- 10μL,

Lane 2 (CS, green) to Lane 7 (S5, yellow)- 20μL. The lid was closed and plugged into the power supply by matching the red and black jacks on the lib by the red and black jacks on the base. The sample was electrophoresed for 30 minutes at 100V and thereafter, the gel was removed from the box. Quick staining method was used to stain the gel to observe better results. 120 ml of 100x Fast Blast stain was added to a staining tray. The gels were stained for 2 minutes and transferred into a washing container to be rinsed with 40-55 degrees’ Celsius water for about 10 seconds and the process was repeated twice for 5 minutes with a light shake each time. Results were then recorded and analyzed. Results DNA fingerprinting process was used to identify the potential suspect in a certain situation. The results are attained by cutting the lengthy sequence of DNA into shorter segment using the restriction enzymes or chemicals. Lane L is the DNA size ladder; CS lane is the crime suspect lane. S1 lane does not have any bands visible, S2-3 lanes consists of few thick and thin bands and S5 lane consists of few fuzzy and fainted bands at the bottom of the gel. Figure 1 DNA Fingerprinting Gel S5 S4 S3 S2 S1 CS

L

Figure 1. DNA Fingerprinting Gel. DNA sample of 20 μL was loaded on the gel for the electrophoresis at 100V for 30 minutes. The lane CS on the gel contains DNA from the crime scene, lanes 1-5 contains the DNA from each of the suspects. Lane L on the gel contains the ladder. Discussion The method of DNA fingerprinting was used to determine the suspect for the crime scene. The DNA sample was run on the gel electrophoresis at 100V for 30 minutes. The lane L on the gel consists of the DNA ladder, the lane CS consists of the crime scene DNA whereas, the lanes from S1-S5 consists of the potential suspects for the criminal scene. In the gel picture used for the result analyses, various bands appeared on each of the columns. Few bands were observed to be brighter and thicker on the gel because a certain gene was targeted (more) and amplified. Bands

that displayed thinner on the gel was due to the less amplification of a certain gene. Lane S1 on the gel displayed no bands which could be due to a human error during the experiment. Lane S2 displayed a thick band and a distinct thin band in the column which matches the with the bands on the crime suspect lane making S2 close match for the crime scene suspect. Lane S3 showed a thick band which matches with one of the crime scenes suspect bands but the other two bands in the lane does not match. Thus, S3 is not a crime suspect for this scene. Lane S4 displayed two thick bands and a fuzzy (fainted) band at the bottom of the gel. Thus, S4 is not the right suspect for this scene. Lane S5 consists of few fuzzy and fainted bands at the bottom of the gel which state that certain gene did not go through a lot of amplification. Thus, S5 is not the suspect for this scene. Hence, DNA fingerprint which is used in various areas of study, investigation, research, science, forensic, etc....