Rapid Colony Transformation of E -Coli with Plasmid DNA Lab Report

Rapid Colony Transformation of E -Coli with Plasmid DNA - Lab Report Example The paper "Rapid Colony Transformation of E -Coli with Plasmid DNA" analyzes E-Coli transformed by plasmid DNA using a rapid method. The transformation method can be categorized into four stages. The first is Pre-incubation stage. The low temperature gels the cell membrane, thereby stabilizing the distribution of charged phosphates and allowing them a more effective shield from the cat ions. The second stage is Incubation. The DNA is added and the cell suspension is kept at 0Â ºC.the cat ions are thought to neutralize negatively charged phosphates in the DNA and the cell membrane. The third stage is Heat Shock. The cell +DNA suspension is briefly incubated at 42Â ºC and then returned to 0Â ºC. The rapid temperature change creates a heat imbalance on either side of the E-Coli membrane and is supposed to create a wave that sweeps plasmids into the cell. The fourth stage is Recovery. LB broth is added to the DNA/cell suspension and incubated at 37Â ºC before being put on plates with different selected antibiotic resistant markers. Transformed cells recover from the treatment, amplify the transformed plasmid and begin to express the antibiotic resistant strain. Samples of E-coli cells are taken from a nutrient agar plate (LB agar) and suspended in two tubes containing a solution of calcium chloride. Plasmid pAMP is added to one cell suspensionBothe the tubes are then incubated at 0Â ºC for 15 minutes. After this a brief heat shock is administered at 42Â º .the samples are cooled and LB broth is added.