Department of Technology and Biotechnology of Drugs – Biologics 2020

Gel electrophoresis: Self-evaluation:

1) 10microlitre of ethidium bromide(10mg/ml)stock solution is made up to 100ml. 1ml of the diluted
solution is taken; find the amount of ethidium bromide in it ?

a) 10 microgram
b) 1 microgram
c) 100 microgram
d) 1 milligram

2) Agarose is a:

a) Polysaccharide obtained from red algae

b) Monosaccharide obtained from red algae
c) Monosaccharide obtained from brown algae
d) Polysaccharide obtained from brown algae

3) If the electrodes are 10cm apart then the gel is run at:

a) 100V
b) 500V
c) 10V
d) 50V

4) TAE buffer contains (i) Tris base (ii) Glacial acetic acid (iii) Ethidium bromide (iv) EDTA

a) i,ii,iii
b) i,ii,iv
c) ii,iii,iv
d) i,iii,iv

DNA Restriction Analysis

Besides their own large circular chromosome, bacteria contain several small circular pieces of
DNA called plasmids. Restriction enzymes can be used to cut open these plasmids. The following
plasmid was cut with two different restriction enzymes, EcoR1 and HindIII. Begin by determining
the number and size of the fragments produced with each enzyme. “Kb” stands for kilobases or
thousands of base pairs.
 Department of Technology and Biotechnology of Drugs – Biologics 2020

Answer the following questions by looking at the gel below.

 Department of Technology and Biotechnology of Drugs – Biologics 2020

a.Which lane shows a digest of EcoR1 only?

b.Which lane shows a digest of HindIII only?

c.Which lane shows a double digest of BamHI and EcoRI?

d.Which lane shows a double digest of BamHI and HindIII

e.On each band in lanes I, II and III and IV write the actual length of each fragment.

1. Which option BEST describes sticky ends.

a) Sticky ends are DNA fragments that carry a higher charge than normal after they have
been cleaved by restriction enzymes.
b) Sticky ends are DNA fragments cleaved by a restriction enzyme so that one strand is
longer than the other.
c) Sticky ends are DNA fragments cleaved by a restriction enzyme so that both strands
are the same length.
d) Sticky ends are DNA fragments that attract a carbohydrate molecule to one end after
being cleaved by a restriction enzyme.

2. In nature, the purpose of restriction enzymes is to:

 Department of Technology and Biotechnology of Drugs – Biologics 2020

a) Protect the bacterium from virus attack by not allowing the virus to attach to the cell
wall.
b) Protect the bacterium from the DNA of other organisms the bacterium infects.
c) Protect the bacterium from replicating its DNA at the wrong time.
d) Protect the bacterium from virus attack by cutting up foreign DNA.

3. Fill:
Cutting DNA with a particular restriction enzyme produces ___________
____________ that can be separated by gel electrophoresis.

4. Fill:
________________ ends are required to create a recombinant DNA molecule.

Use the following information to answer questions 5–7.

Below is a plasmid with restriction sites for BamHI and EcoRI. Several restriction digests
were done using these two enzymes either alone or in combination.
Hint: Begin by determining the number and size of the fragments produced with each
enzyme. "kb" stands for kilobases, or thousands of base pairs.

5. Which lane shows a digest with BamHI only?

a.I
 Department of Technology and Biotechnology of Drugs – Biologics 2020

b.II
c.III
d.IV
e.V
6. Which lane shows a digest with EcoRI only?
a.I
b.II
c.III
d.IV
e.V
7. Which lane shows the fragments produced when the plasmid was incubated with
both EcoRI and BamH1?
a.I
b.II
c.III
d.IV
e.V