Author: Vy Nguyen

Group members: Shelby Baughn,

Katie Holland, Vy Nguyen

To: Dr. Alice Jernigan

Date of experiment conducted: December 2, 2014

Date of report submitted: December 9, 2014

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 Abstract

This experiment is about gel electrophoresis.Gel electrophoresis is one of the most

common technique for separating macromolecules by size or charge, and is widely used in
biochemical research and pharmacology. The gel experiment was proceeded by loading dye
samples into the prepared gel and electrophoresed by the power supply. The results were
ponceau G and xylene cyanol for the unknown 1, bromphenol blue and ponceau G for unknown
number 2, and the mixture of xylene cyanol and pyronin Y for unknown number 3. Besides, the
farthest dye sample was pyronin Y. Overall, the experiment went well because all the data were
achieved properly. In addition, there was no sign of errors in the gel electrophoresis experiment.
 Table of Contents
Page
Abstract ..........................................................................................................0
Table of Contents............................................................................................ii
List of Table....................................................................................................iii
List of Figures.................................................................................................iv
Introduction....................................................................................................1
Materials and Methods...................................................................................2
Results, Discussion and Conclusion...............................................................3
References......................................................................................................4
Appendix A....................................................................................................5

ii
 List of Tables
Page
Table 1: Results of the Gel Electrophoresis…………………………………3

iii
 List of Figures
Page
Figure 1: Example of band of DNA...........................................................1
Figure 2: Picture of dye samples from group B.........................................2

iv
 1

Introduction

Gel electrophoresis is one of the most common technique for separating macromolecules
by size or charge, and is widely used in biochemical research and pharmacology.
Macromolecules, such as DNA, is often used for CSI or criminal cases to identify fingerprints.
To examine, a sample of DNA is placed in an agarose gel, and an electrical charge is sent
through by the power supply, based on size. The negatively charged molecules of DNA moves
toward to another the positively charged molecules. After separating, the bands are stained with
dyes, such as methylene blue or ethidium bromide in order to be seen. The result of DNA is a
series of bands with different sizes. The farthest band from the start of gel is the smallest band of
DNA. Meanwhile, the closet band to the start of gel is the largest band of DNA (1).

Figure 1: Example of band of DNA (2).

Agarose is a popular gel that often used in chemistry labs and other science fields.
Agarose gel is formed by mixing agarose with the buffer, and allows time to become a gel
matrix. By adjusting the concentration of agarose used in the gel, the sizes can be determined.
Specifically, the more concentrated the gel, the longer DNA takes to move. Conversely, the less
concentrated of the gel, the fast the DNA moves (3). There are many advantages of using agarose
gel. One of the main advantages is that agarose is a nontoxic gel, that does not harm to human.
Besides, agarose can recovers samples by melting the gel, digesting with enzyme agarose or
treating with salts (4).
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Materials and Methods

The main materials used in the gel electrophoresis lab were 1.6 percent of agarose
mixture, 1X TBE, unknowns, dye samples and power supply. The experiment began by casting
the agarose gel with 1X TBE and dissolved in a hot bath. Then, the mixture was filled, and
covered about half of the height of the comb teeth of the casting tray. The process to condense
took around 15 minutes. Once the agarose was solidified, the comb was removed and ready to
use.
Before loading the dye samples into the gel, the gel was filled with 1X TBE buffer that
just covered the surface of the gel. The given dye samples, including three unknowns, were
loaded into the gel by using the digital pipettor. To load the first dye sample, the pipette tip was
used to draw 10 micro liter of the dye. Using the dominant hand, the pipette was steadily poured
into the well above the buffer layer from left to right. Besides, the non-dominant hand was used
to guide the pipet through the surface of the buffer and positioned directly over the well. The dye
was sunk to the bottom of the well because the dye was already mixed with surcrose to increase
the density. The experiment continued processing for the rest of the samples.
Once all of the dye samples were loaded, the lid was placed on the electrophoresis
chamber, and the electrical cords were connected to the power supply. 100 Volts was used for
the whole process. When the power supply was turned on, the gel ran slowly until the dye in lane
3 was 0.5 cm from the end of the gel. Later, the result was all recorded and analyzed.

Figure 2: Picture of dye samples from group B (5)

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Results, Discussion, and Conclusions

Table 1: Results of the Gel Electrophoresis

Lane Sample Number of bands (+) or (-) Migration

distance (cm)
1 Bromphenol blue 1 (-) 1.5
2 Methyl orange 1 (-) 0.8
3 Pronceau G 1 (-) 2
4 Xylene cyanol 1 (-) 0.5
5 Pyronin Y 1 (+) 1.7
6 Unknown #1 2 (-) 0.5/2.2
7 Unknown #2 2 (-) 1.5/2.2
8 Unknown #3 2 (-)/(+) 1/0.5

After the results, the unknown number 1 was identified as a mixture of xylene cyanol and
ponceau G; bromphenol blue and ponceau G for unknown number 2; and the mixture of xylene
cyanol and pyronin Y for unknown number 3. From observing, the farthest dye molecule through
the gel was pyronin Y because this type of gel was the smallest from total of eight dye samples.
The smaller the gel is, the faster the sample goes. Conversely, the bigger the gel is, the slower the
sample goes. Besides, the charges of the dye samples were also determined. Negative charge was
found for bromphenol blue, methyl orange, pronceau G, xylene cyanol, and unknown number 2.
Positive charge was found for pyronin Y. The only mixture of negative charge and positive
charge was unknown number 3. When doing electrophoresis, the mixture of unknown 3 went
two ways, including xylene cyanol and pyronin Y. In the aspect, the result seemed to be correct
because pyronin Y was positive, and xylene cyanol was negative; therefore, the mixture had to
go two different directions.
Overall, the gel electrophoresis experiment went really well. There were no source of
errors since the steps were done properly.
4

References
1. "Gel Electrophoresis." Gel Electrophoresis. N.p., n.d. Web. 06 Dec. 2014.
2. T., and Introduction to Agarose Gel Electrophoresis: A Precursor to Cornell Institute for
Biology Teacher’s Lab (n.d.): n. pag. Web. 6 Dec. 2014.
3. "Agarose Gel Electrophoresis." Life Technologies. N.p., n.d. Web. 06 Dec. 2014.
4. Barril, Patricia, and Silvia Nates. Introduction to Agarose and Polyacrylamide Gel
Electrophoresis Matrices with Respect to Their Detection Sensitivities. N.p.: INTECH
Open Access, 2012. Web. 6 Dec. 2014.
5. Nguyen, Vy, Katie Holland, and Shelby Baughn. Picture of Gel Electrophoresis. Digital
image. N.p., n.d. Web. 6 Dec. 2014.
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Appendix A
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