E. Coli Lab Report

The purpose of this lab is to successfully infiltrate E. coli bacterial cells with a pARA-R plasmid that is antibiotic resistant and has the rfp gene, or red fluorescent protein. This can be verified if the E. coli obtains the characteristics of the plasmid when it enters. To start, three Petri plates containing agar are needed. On each plate there is a control group and a treatment group; the treatment group being the one with the plasmid. Before the plasmid is put with the E. coli, first the bacteria are “stressed out” by warming them up in a hot water bath and cooling them down very rapidly in ice. The first plate consists of Luria Broth (LB), the second plate consists of LB and the antibiotic ampicillin (amp), and the last one contains LB, amp, and the sugar arabinose (ara). The bacterial cells are subjected to a heat shock and then are placed onto the three plates. The plasmid is spread on to only half of the first two plates, on the sides of the treatment group. Half of the E. coli get the plasmids and the other half do not (that side being the control group). On the third plate the plasmids are spread on the whole plate. The bacteria are left in an
Once the recombinant plasmid has entered the bacterial cell, DNA polymerase initiates replication at the ori site, or the DNA sequence that signals for the origin of replication. The plasmid replicates using the bacterial DNA replication enzymes. The multiple copies of plasmids now can produce the red fluorescent protein in great quantities. If the simple sugar arabinose is present in the bacteria then an activator protein made by the araC (arabinose activator protein) gene turns on the promoter (pBAD), which then binds binds RNA polymerase. Consequently, transcription of the rfp gene occurs. After the processes of transcription and translation the protein alters the observable traits of the organism, specifically making it