Identifying an Unknown Plasmid Through the Process of Gel Electrophoresis
Introduction:
Biotechnology requires certain techniques and methods that help identify plasmids, which can be used for forensics, DNA fingerprinting, etc. In this class, each lab focused on teaching the process of using the correct techniques used to identify a plasmid. Plasmids are pieces of DNA that are circular and relatively smaller than chromosomes. They aren’t important in the sense that they don’t carry out critical functions required for growth and life. Also, they are located in the cytoplasm, outside of the chromosome and nucleus of a cell. They hold what are considered to be “accessory” genes. This key characteristic allows scientists to conduct studies and manipulate bacterium. A plasmid has certain restriction sites in which enzymes cut the DNA in order to insert a desired DNA sequence. These restriction enzymes help identify the plasmids which is crucial in many areas of science. Plasmids are used for gene expression …show more content…
In planning, it is important to make sure the enzymes are compatible with each other in the double digest. In the single digest we selected the enzyme Pst1, which uses a 3.1 buffer. Then for the double digest, using the provided resources from New England Biolabs, such as the poster in classroom, we selected two enzymes that were compatible with each other and that required the same buffer. HindIII and Ndel1 required a 2.1 buffer. Now that the enzymes had been prepared for the digest, locate the NEBcutter online to virtually digest them to see what the gel should look like. Use dnalc.org to copy and paste the sequences for the 3 possible types of plasmids the unknown plasmid would be. Which are pAMP, pBLU, or pKAN. When completing digestion of the single and double digests with the appropriate enzymes, print them out in order to compare when the results have been produced in the
Introduction:
Biotechnology requires certain techniques and methods that help identify plasmids, which can be used for forensics, DNA fingerprinting, etc. In this class, each lab focused on teaching the process of using the correct techniques used to identify a plasmid. Plasmids are pieces of DNA that are circular and relatively smaller than chromosomes. They aren’t important in the sense that they don’t carry out critical functions required for growth and life. Also, they are located in the cytoplasm, outside of the chromosome and nucleus of a cell. They hold what are considered to be “accessory” genes. This key characteristic allows scientists to conduct studies and manipulate bacterium. A plasmid has certain restriction sites in which enzymes cut the DNA in order to insert a desired DNA sequence. These restriction enzymes help identify the plasmids which is crucial in many areas of science. Plasmids are used for gene expression …show more content…
In planning, it is important to make sure the enzymes are compatible with each other in the double digest. In the single digest we selected the enzyme Pst1, which uses a 3.1 buffer. Then for the double digest, using the provided resources from New England Biolabs, such as the poster in classroom, we selected two enzymes that were compatible with each other and that required the same buffer. HindIII and Ndel1 required a 2.1 buffer. Now that the enzymes had been prepared for the digest, locate the NEBcutter online to virtually digest them to see what the gel should look like. Use dnalc.org to copy and paste the sequences for the 3 possible types of plasmids the unknown plasmid would be. Which are pAMP, pBLU, or pKAN. When completing digestion of the single and double digests with the appropriate enzymes, print them out in order to compare when the results have been produced in the