Bacterial Transformation Lab Report

Bacterial Transformation: Effect of pGreen on the Amount of Bacterial Colonies on an Ampicillin Based Agar Plate Introduction: Bacterial transformation is the result that occurs when exogenous DNA is infused into a prokaryotic cell. The new genotypic makeup translates into a new phenotypic expression. In the case of the lab, the plasmid, pGreen was introduced to the experimental group. In both the control and the experimental group, the existing bacteria was E. Coli MM294. Plasmids are parts of DNA that carry genes. The particular plasmid that was used in this lab was pGreen, which carries the genes GFP and ampR. GFP codes for a protein that makes the bacteria a fluorescent green color and ampR codes for a protein that makes the bacteria

The purpose of this lab was to ascertain the effect that the adoption of the plasmid pGreen has on the survival of bacterial colonies. Through the results, bacterial transformation is proven and the relationship between DNA to RNA to a protein through transcription and translation is demonstrated. In this experiment there were two test tubes with a solution of calcium chloride (CaCl2). The E. Coli MM294 was exposed to the solution at a cold temperature. Then a inoculating loop was used to add plasmid DNA to the experimental group. The test tubes were then incubated in ice for 15 minutes. The plates were subsequently immersed into a 42°C water bath for 90 seconds for a heat shock. Then luria broth was added to both tubes to allow for an increased rate of bacteria growth. After a recovery period, the cells were transferred onto the corresponding ampicillin based agar plates. The data collected was the number of colonies that had adopted the plasmid on the experimental plate and the number of colonies on the control plate. In the experimental group, some of the small colonies that were surrounding other colonies were not fluorescent green. These colonies were only surviving due to an antibiotic shadow and were not counted in the data. If the

In order for the bacteria to survive on the ampicillin plate, the bacteria would have to be resistant to ampicillin. For a protein to be produced the existing DNA of the E. Coli MM294 should be converted into RNA. This process is called transcription which contains the processes of initiation and elongation in prokaryotic cell. In the initiation phase, RNA polymerase binds to the promoter region of the DNA and then breaks the hydrogen bonds between the two strands of DNA. In the elongation phase, RNA polymerase adds the corresponding nucleotides to the DNA strand, but instead of thymine there is uracil as a nitrogenous base and the sugar is ribose instead of deoxyribose. There are three types of RNA, mRNA, tRNA, and rRNA. Once the RNA strand is produced, the next step of gene expression is translation. First, in translation, the ribosome assembles on the mRNA. The ribosome contains two parts, with the smaller part containing 3 “sites.” There is the amino acid (A), polypeptide (P), and exit (E) regions. The tRNA carries the amino acids and each tRNA also carries an anticodon. The codon AUG (UAC anticodon) is the start codon. The tRNA with the corresponding anticodon will temporarily bind to the mRNA codon. The tRNA shifts from the A to P and a new tRNA takes the spot of the A. As the process continues, the tRNA exits through the E. The amino acids bind with each other