Analysis of Lambda DNA using Restriction Enzymes and Agarose Gel Electrophoresis Sage Hill School, 20402 Newport Coast Drive, Newport Coast, CA 92657 1. Introduction DNA, also known as deoxyribonucleic acid, consists of nitrogenous base molecules held together by weak hydrogen bonds. DNA is necessary to encode genetic instructions for the development and functioning of all living organisms. If the DNA is changed or adjusted, the structure and function of the organism will change as well. Selectively replacing certain portions of a DNA strand may change the original characteristics of the organism. DNA splicing allows us to remove a functional fragment of DNA from one organism and replace that fragment with a substitute DNA sequence another …show more content…
These micro test tubes were then placed on a Styrofoam test tube holder and put onto a beaker containing an ice water solution. We obtained four more micro test tubes and labeled them L, P, E, and H. The tube labeled L contained lambda DNA, the tube labeled P contained PstI lambda digest, the test tube labeled E contained EcoRI lambda digest, and the test tube labeled H contained HindIII lambda digest. (Labels and contents are shown in Figure 2.1). Using a micropipette, 4 μl (four microliters) of lambda DNA were added to each micro test tube. Then, 5 μl of restriction buffer were added to each micro test tube, using a fresh pipette tip to avoid mixing the contents of the test tubes. Test tube L, containing lambda DNA, would be our control reference in the experiment, therefore remaining uncut throughout the experiment. We added one type of enzyme to each of the other micro test tubes using a micropipette (once again using fresh tips). We added one μl of PstI restriction enzyme in test tube P, one μl of of EcoRI restriction enzyme in test tube E, and one of μl of HindIII restriction enzyme in test tube H (as shown in Figure 2.1). Next, the micro test tubes were tightly capped, and the contents inside were gently mixed by flicking the tube. To collect all the liquid to the bottom, we placed the test tubes in a centrifuge for about 30 …show more content…
However, the smaller fragments of DNA will travel faster than the bigger fragments, allowing us to determine the sizes of the fragments. The effectiveness of the restriction enzymes are observed by the number of fragments the strand is broken into. When analyzing the results, one can determine the number of restriction sites by the number of bands seen. Our results showed the lambda digest of PstI (P) traveled the farthest and possess the greatest number of bands (the most restriction sites) and the smallest DNA fragments. In addition, our results showed that lambda DNA digest for restrictive enzyme HindIII (H), which traveled the second farthest, has the second most amount of bands, indicating this restrictive enzyme has the second greatest number of restriction sites, but still larger DNA fragments than restrictive enzyme PstI digest. The restriction enzyme EcoRI has the fewest restriction sites, therefore the longest DNA fragments. Finally, as we expected the results for the undigested lambda DNA traveled the least because it had not been cut by a restriction