A sample of DNA found in a crime scene was provided along with five suspects. Their DNA was then processed using restriction enzymes and Agarose Gel Electrophoresis. The objective of this lab was to match a criminals DNA to a crime scene using restriction enzymes EcoRI and Pstl with Agarose gel electrophoresis. Restriction enzymes cut DNA at a specific base pair site recognized by the enzyme, which then turns one single strand of DNA into many fragmented strands of DNA. EcoRI recognizes and cuts the palindromic base pair sequence GATTC while Pstl recognizes and cuts the palindromic base pair sequence CTGCAG. Agarose gel electrophoreses separates these fragmented DNA by their size. The negatively charged DNA moves through the Agarose gel to the positively charged end of the gel. The smaller fragments move through the gel more quickly allowing a linear view of the fragmented DNA when the process is complete. Since each individuals DNA will be cut into different size fragments when restriction enzymes are applied we can match one of the suspects to the crime scene DNA sample. This process enables an individual’s DNA to be matched, much like a fingerprint, to a sample of unknown DNA. Methods
An enzyme mix of EcoRl and Pstl was added 10 microliters at a time to the crime scene sample and suspect samples one through five each containing 20 microliters of DNA. A new pipet was used for each transfer of the enzyme mix to ensure that there was no cross contamination of the suspects. To guarantee the enzyme reacts with the DNA the six samples mixed with enzyme were then centrifuged. The samples were incubated at 37 ° C for 45 minutes, after incubation 5 microliters of dye were added to each sample. During this time an Agarose gel was cast using an 8 well comb. The Agarose gel was placed in the electrophoresis chamber with the wells at the cathode end and 275mL of electrophoresis buffer was added. In the first well 10 microliters of Hindlll DNA...
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