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Unformatted text preview: Experiment 6 (Lab periods 5 and 6) Restriction Enzymes and Restriction Fragments Restriction enzymes (or restriction endonucleases) are bacterial proteins that act as defense mechanisms in these organisms. Restriction endonucleases cleave double stranded DNA internally, acting essentially like a pair of scissors to cut both strands of the molecule. When foreign DNA (such as viral DNA) is introduced into a bacterium it is the job of the restriction endonucleases to cut the foreign DNA so that they act like a simple immune system. The bacteria protect their own DNA by chemically modifying it a job performed by a special DNA methylase enzyme. In molecular biology we take advantage of the natural function of restriction enzymes by using them to perform a number of useful procedures. Two very common uses are 1) to generate DNA molecules that can be used to create recombinant DNA and 2) to get an estimate of the similarity in nucleotide sequence of different pieces of DNA. In this lab we will use restriction enzymes for the second purpose. Different species of bacteria have different restriction enzymes. The enzymes are referred to by a designation of the species and strain from which they are isolated. For example, the enzyme EcoRI is isolated from Escherichia coli strain R and SacI is isolated from Streptomyces achromogenes . All restriction enzymes act in the same general manner but differ in an important detail. A restriction enzyme does not cut a DNA molecule randomly but, instead, each restriction enzyme recognizes a specific nucleotide sequence in the DNA molecule. This sequence is referred to as the recognition sequence for the enzyme. The difference between different restriction enzymes is that each has its own recognition sequence. For example the following are the recognition sequences for EcoRI and SacI: sequence....
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