In this protocol, you will be using chemically competent E coli that have been placed in a calcium chloride solution prior to freezing. The solution acts to neutralize the cells so that they don’t repel each other. The bacteria and plasmid mixture will be chilled on an ice for 30 minutes. Placing the mixture in a 42 degrees Celsius water bath for 30 seconds will heat shock the mixture, which will cause the transformation. Once the LB is added and mixed with the transformed bacteria, it can be plated on the LB plate with an antibiotic. Since a lot of the bacteria will not be transformed after the heat shock, plating E. coli with an antibiotic will ensure only the transformed E. coli survive. Before doing this lab, please watch this video: https://vimeo.com/25201947. Note that we do not have a control for our experiment, and we are not using a thermomixer. Separate reagents from materials. Basic Terminology and concepts Transformation – The process in which the genetic makeup of a cell is changed by introduction of DNA from the surrounding environment. Competent E. coli – E. coli in a state where it can allow the uptake of DNA Heat Shock – A sudden increase in temperature used to propel a plasmid into a bacterium Safety precautions This is a benign lab protocol, but don’t forget that PPE is necessary at all times. The E. coli you will be working with will be non-pathogenic but it should still be handled properly, which means it should not come into contact with your skin or gloves. If you are removing materials from the -80 degrees Celsius freezer, do not use your bare hands or regular lab gloves, use gloves designated specifically for the freezer.