The document describes the process of inserting a human insulin gene into bacterial DNA through genetic engineering. First, the insulin gene is cut from human DNA using a restriction enzyme, leaving "sticky ends." A plasmid from bacteria is also cut with the same enzyme. The insulin gene and plasmid are mixed and joined together with the enzyme DNA ligase. The combined DNA is then inserted into E. coli bacteria through applying heat or electric shock to open pores in the bacterial membrane, allowing the plasmid to enter. The engineered bacteria now contains the human insulin gene and can produce human insulin protein.

1. Chapter 20
Molecular
Genetics

2. Deoxyribonucleic acid (DNA)
• Deoxyribonucleic acid (DNA) is a
molecule that carries genetic
information.
• These genetic information is important
for all cellular functions, such as cell
division and cell differentiation.
• Almost all cells in our body contain
DNA inside their nuclei.

3. DNA
Each DNA molecule
consists of two parallel
strands twisted around
each other to form a
double helix.
A molecule of DNA
is wrapped around
proteins to form a
single chromatin
thread.
During cell
division, the
chromatin threads
coil more tightly to
form chromosomes
inside the cell
nucleus.
proteins
nuclear pore
nucleus
nuclear envelope

4. What is DNA made of?
Double helix ‘untwisted’
One strand of nucleotides
Components of a single nucleotide
DNA molecule: a long double helix
basedeoxyribose
sugar
phosphate
group
Sugar phosphate ‘backbone’

5. Basic units of DNA
adenine cytosine guanine thymine deoxyribose
sugar
phosphate
group
Bases
Nucleotides
base joins with the
phosphate group and
deoxyribose sugar group

6. Basic units of DNA
• The basic unit of DNA is a
nucleotide.
• Each nucleotide is made
of
- a sugar called
deoxyribose;
- a phosphate group; and
- a nitrogen-containing
base, all joined together
• The four bases of
nitrogen-containing bases
are
- adenine (A)
- thymine (T)
- cytosine (C)
adenine cytosine guanine thymine deoxyribose
sugar
phosphate
group
Bases
Nucleotides
adenine nucleotide thymine nucleotide
guanine nucleotide cytosine nucleotide
base joins with the
phosphate and deoxyribose
sugar group

7. The building blocks of DNA
• Nucleotides are joined together to form long chains
called polynucleotides.
• Each gene is made up of a sequence of nucleotides. This
sequence can vary.
bases
sugar-phosphate backbone
polynucleotide

8. Rule of base pairing
• The bases of one strand form bonds with bases of the
other strand according to the rule of base pairing.
• Adenine (A) bonds with thymine (T), while
cytosine (C) always bonds with guanine (G).
• Bases that bond with each other are known as
complementary base pairs.
base pair

10. Guide to be a God!
1) Colour each of the individual structures on
the worksheet with a different colour:
Example:
adenine = red
thymine = green
guanine = blue
cytosine = yellow
phosphate = brown
deoxyribose = purple

11. Guide to be a God!
2) Cut out each structure.
3) Using the small symbols (squares, circles and
stars) on the structures as guides, line up the
bases, phosphates and sugars.
4) Glue the appropriate pairs together to form
nucleotides.

12. Example:

13. Guide to be a God!
5) Construct the right side of your DNA molecule
by putting together in sequence a
cytosine, thymine, guanine and adenine
nucleotide.
6) Complete the left side of the DNA ladder by
adding complementary nucleotides or
nucleotides that fit. Your finished model should
resemble a ladder.

14. Guide to be a God!
7) To show replication of your model, separate
the left side from the right side on your desk,
leaving a space of about 15 to 20 cm.
8) Using the remaining nucleotides, add to the
left side of the model to build a new DNA
molecule. Do the same with the separated right
side.

15. Questions?!?!
1) When constructing the DNA molecule, what
did you notice about the orientation of the
two strands?
2) What DNA strand would bond opposite?
3) What is a similarity and a difference in DNA
between Homo sapien and Blattaria?

16. The DNA double helix
The DNA molecule has a spiral structure known as the
double helix. Both strands of DNA that run in opposite
directions are twisted to form this double helix.
a base pair
sugar-phosphate backbone
coiling of DNA
double helix
structure of DNA

17. Genes
• A DNA molecule contains many genes along its
length.
• A gene is a small segment of DNA which controls
the formation of a protein, such as an enzyme.
gene
DNA
molecule

18. Genes
• Each gene stores a message that determines how a
protein should be made in a cell.
• The message stored by a gene is known as the
genetic code.
• Proteins are responsible for the development of
certain characteristics in the body.
gene
DNA molecule
part of a DNA molecule unzipped to show a gene
M E S S A G E
a gene is a segment of DNA
protein coded
by the gene

19. Structure of a gene
• Each gene consists of two polynucleotide chains. One of the
chains determines the type of protein made. This chain is
called the template.
• The template is made up of a sequence of nucleotide bases.
• Three sequential bases code for one amino acid. This is
known as the triplet code or codon.
DNA template
Process of decoding and protein synthesis
polypeptide made of five
amino acids
triplet code/ codon

20. How are proteins made?
• Proteins in the cell are made through a two-step
process — transcription and translation.
• Transcription occurs when the message in the template
has to be copied into an RNA molecule called messenger
RNA (mRNA).
• Transcription occurs in the nucleus.
• Three bases in the mRNA made up a codon.
DNA template
TranscriptionmRNA
- RNA contains U
(uracil) instead of T
(thymine)
codon

21. How are proteins made?
• The mRNA moves out of the nucleus and carries the
message to the cytoplasm.
• A ribosome helps to translate the sequence of codons on
the mRNA into a protein molecule.
mRNA
polypeptide
Translation

22. Comparing DNA and RNA
DNA (double helix) RNA
Sugar unit is deoxyribose. Sugar unit is ribose.
Nitrogen-containing bases
are adenine (A), thymine
(T), cytosine (C) and
guanine (G).
Nitrogen-containing bases
are adenine (A), uracil (U),
cytosine (C) and guanine
(G).
Permanent molecule in
the nucleus
Temporary molecule that
is made when needed
Found only in nucleus Found in nucleus and
cytoplasm

23. 1 part of a gene
Transcription and Translation

24. First, the gene unzips.
1 part of a gene
Transcription and Translation

25. template
mRNA molecule is
made
One of the strands in the gene is
used as the template to make
mRNA. This is transcription. The
mRNA molecule copies the genetic
code in the DNA template,
following the rule of base pairing.
1
Note that mRNA does not contain
T (thymine). A (adenine) in DNA
pairs with U (uracil) in mRNA.
Transcription and Translation

26. mRNA molecule is
made
ribosome
mRNA
nuclear
envelope
The mRNA leaves the nucleus
and attaches to a ribosome in
the cytoplasm.
2
nuclear pore
Transcription and Translation

27. tRNA • In the cytoplasm are amino acids
and transfer RNA (tRNA). Transfer
RNA or tRNA is another RNA
molecule also needed for protein
synthesis.
• tRNA molecules have amino acids
attached to one end of their
structure.Each tRNA is very specific
and attaches only to its own amino
acid For example, a tRNA with the
anticodon UAC always attaches to
the amino acid M.
• Each tRNA also has three bases at
one end. This is an anticodon that
can bind to complementary codons
on mRNA.
cytoplasm
amino
acids
3
anticodon
Transcription and Translation

28. codon
The anticodons on tRNA
bind with their respective
codons on mRNA.
tRNA
amino acid
attached to
tRNA
peptide bond
ribosome
4 • Translation starts with mRNA attaching
to a ribosome.
• The first two tRNAs together with their
amino acids also fit into the ribosome.
They attach to the codons on the mRNA
according to the rule of base pairing.
• A peptide bond is formed between the
two amino acids.
Transcription and Translation

29. 5
peptide bond between
amino acids
first tRNA
is released
a new tRNA
fits into the
ribosome
• Once the peptide bond is formed
between the first two amino
acids, the ribosome moves along
one codon to the right of the
mRNA.
• As the ribosome moves to this
position, the first tRNA is released.
• At the same time, the third tRNA
and its amino acid slots into the
ribosome.
codon ribosome moves along
the mRNA strand
Transcription and Translation

30. 6
another amino
acid is attached to
the chain
direction of movement
of ribosome
• Another amino acid is attached
to the chain.
Transcription and Translation

31. polypeptide formed
• The process continues as
the ribosome moves along
the mRNA.
• At the end of the mRNA
is a stop codon such as
UGA, UAA or UAG. A
stop codon does not have
any tRNA with
complementary codons.
This means that
anticodons ACU, AUU or
AUC do not exist.
• Eventually, the whole
chain of polypeptide is
produced. The ribosome
leaves the mRNA.
7
Transcription and Translation

32. Transcription and Translation
polypeptide formed
• The ribosome may
attach to the same
mRNA for another
round of translation.
8

33. insulin gene
• Obtain the human chromosome
containing the insulin gene.
1
How the human insulin
gene is inserted into
bacterial DNA
Genetic Engineering

34. insulin gene
• Obtain the human chromosome
containing the insulin gene.
• Cut the gene using a restriction
enzyme. This enzyme cuts the
two ends of the gene to produce
‘sticky ends’.
1
cut by restriction
enzyme
How the human insulin
gene is inserted into
bacterial DNA
fragment of DNA containing
the insulin gene
sticky end
Genetic Engineering

35. insulin gene
• Obtain the human chromosome
containing the insulin gene.
• Cut the gene using a restriction
enzyme. This enzyme cuts the
two ends of the gene to produce
‘sticky ends’.
• Each ‘sticky end’ is a single
strand sequence of DNA bases.
These bases can pair with
complementary bases to form a
double strand.
1
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
How the human insulin
gene is inserted into
bacterial DNA
Genetic Engineering

36. insulin gene
• Obtain a plasmid from a
bacterium.
2
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
plasmid
How the human insulin
gene is inserted into
bacterial DNA
Genetic Engineering

37. insulin gene
• Obtain a plasmid from a
bacterium.
• Cut the plasmid with the same
restriction enzyme. This produces
complementary sticky ends.
2
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
plasmid
cut by same
restriction enzyme
sticky ends
How the human insulin
gene is inserted into
bacterial DNA
Genetic Engineering

38. insulin gene
• Mix the plasmid with the DNA
fragment containing the insulin
gene.
3
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
plasmid
cut by same
restriction enzyme
sticky ends
How the human insulin
gene is inserted into
bacterial DNA
Genetic Engineering

39. insulin gene
• Mix the plasmid with the DNA
fragment containing the insulin
gene.
• Add the enzyme DNA ligase to
join the insulin gene to the
plasmid.
3
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
plasmid
cut by same
restriction enzyme
sticky ends
insulin gene
inserted into
plasmid
How the human insulin
gene is inserted into
bacterial DNA
DNA
ligase
Genetic Engineering

40. insulin gene
• Mix the plasmid with E. coli
bacteria.
4
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
plasmid
cut by same
restriction enzyme
sticky ends
insulin gene
inserted into
plasmid
E. coli
bacterial DNA
How the human insulin
gene is inserted into
bacterial DNA
DNA
ligase
Genetic Engineering

41. Genetic Engineering
insulin gene
• Mix the plasmid with E. coli
bacteria.
• Apply temporary heat or electric
shock. This opens up pores in the
cell surface membrane of each
bacterium for the plasmid to
enter.
4
cut by restriction
enzyme
fragment of DNA containing
the insulin gene
sticky end
plasmid
cut by same
restriction enzyme
sticky ends
insulin gene
inserted into
plasmid
plasmid
bacterial
DNA
plasmid enters
the bacterium
trangenic bacterium
E. coli
bacterial DNA
How the human insulin
gene is inserted into
bacterial DNA
DNA
ligase