INTRODUCTION

Bacterial transformation occurs if a bacterium uptakes a piece of DNA and

integrates it into its genome. The phenotype of any organism, including that of a human

being, is not solely determined by its genotype. In nearly all cases, it is the complex

interaction of genotype and environment that determines the overall phenotype of an

individual (McGee, 1997). Exactly how the environment affects phenotype is an area of

ongoing investigation. Not surprisingly, this topic has been studied thoroughly in

prokaryotes. Bacterial cells exposed to changing environmental conditions often

respond by altering their shape, form, and/or cellular metabolism (White, 2000).

Bacterial transformation is done in two different ways in the lab: electroporation and

through calcium chloride/heat-shock. Electroporation is when electrical shock is used to

make the cell membrane permeable to DNA. Calcium chloride/heat-shock is when the

plasmids are incorporated into the chemically-competent cells made permeable by the

heat shock and calcium chloride solution.

Escherichia coli or E. coli is a gram negative, bacillus, bacteria. E. coli makes a

great bacterial candidate for the bacterial transformation because it is made up of one

cell, it reproduces every twenty minutes, it does not harm people, and it cannot survive

outside of the lab. Ampicillin is an antibiotic that has a beta-lactam structure. It has the

ability to kill E. coli cells.

Plasmids are small circular autonomously replicating pieces of DNA, found inside

prokaryotic bacterial cells. They replicate their own DNA by borrowing the cells

polymerase. Due to their size, plasmid DNA is easy to extract and purify from bacterial
cells. Plasmids may express antibiotic resistant genes or be modified to express

proteins of interest, and are useful for cloning foreign genes.

The pGLO plasmid contains the gene for green florescent protein (GFP) and a

gene for ampicillin resistances called beta-lactamase. The pGLO plasmid also contains

a special gene regulation system or operon,that can be used to control expression of

the florescent protein in the transformed cells called araC regulator protein.

Green fluorescent protein, or GFP, is found naturally in the luminescent

jellyfish Aequorea Victoria. GFP glows bright green when exposed to ultraviolet light due

to resonation which the light causes. GFP is mainly used in biotechnology as a

biological marker or indicator. It has also been used to produce luminescent plants and

animals. GFP production is monitored by a modified arabinose operon located

on the pGLO plasmid. In the operon, the protein araC blocks RNA polymerase from

binding to the Pbadpromoter. When arabinose is present it changes the shape of

the araC protein causing it to promote, instead of prevent, RNA polymerase binding.

Once RNA polymerase has attached to the promoter, transcription of the GFP gene

begins and continues until arabinose runs out. However, AraC only binds to the

appropriate activator sequence in the presence of the sugar arabinose (Snyder and

Champness, 1997).

Ampicillin is an antibiotic that falls into the penicillin group of drugs. Ampicillin is

often used to fight bacteria in the human body, specifically various types of infections.

Such infections may include those caused by bacteria, like ear infections, bladder

infections, pneumonia, gonorrhea, and E. coli or salmonella infection. Ampicillins ability

to exterminate bacteria makes it ideal for PGLO transformation.

REFERENCE
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gene expression in Escherichia coli. Bioscene, 28(3), 17-23.
2. Sha, J., Wang, Y., Wang, J., Liu, W., Tu, Q., Liu, A., & Wang, J. (2011). Heat-shock
transformation of Escherichia coli in nanolitre droplets formed in a capillary-composited
microfluidic device. Analytical Methods, 3(9), 1988-1994.
3. Wright, L. K., & Newman, D. L. (2013). Using PCR to target misconceptions about gene
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