College Board AP® Biology Your notes

Biotechnology: Genetic Engineering

Techniques
Contents
Gel Electrophoresis
The Polymerase Chain Reaction (PCR)
Bacterial Transformation
DNA Sequencing

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 Gel Electrophoresis
Your notes
Gel electrophoresis
Gel electrophoresis separates molecules according to their relative size and electrical
charge
This technique can also be used with DNA, RNA and proteins
During gel electrophoresis molecules are placed into a gel, and the different rates at
which they move through the gel causes them to separate
This separation of molecules occurs due to:
their electrical charge
There is an electrical charge across the gel, with a positive electrode at one end
and a negative electrode at the other
DNA is negatively charged so DNA molecules move towards the positive
electrode
their different sizes
Molecules move through the gel at different rates depending on their size
The tiny pores in the gel allow smaller molecules to move more quickly, while
larger molecules move more slowly
It is possible to identify the size of fragments of unknown length by comparing
the distance travelled with fragments of known size
Gel electrophoresis can be used to produce DNA profiles, which can be useful for, e.g.
determining how closely related individuals or species are
forensic analysis, e.g. identifying DNA found at a crime scene
aiding in the process of DNA sequencing

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 Your notes

Gel electrophoresis separates molecules by size and charge

Examiner Tips and Tricks

The full details of the process of gel electrophoresis are beyond the scope of the AP
Exam. You should focus on understanding application of the technique.

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 The Polymerase Chain Reaction (PCR)
Your notes
The polymerase chain reaction (PCR)
The polymerase chain reaction (PCR) is a common technique used in most applications
of gene technology, e.g. to produce enough DNA for:
DNA profiling when identifying criminals or determining paternity
phylogenetic analysis
transforming bacteria during genetic modification
PCR is used to produce many identical copies of a DNA fragment from a small initial
sample
The DNA is said to be amplified
The PCR process involves three key stages per cycle:
denaturation
The double-stranded DNA is heated to break the hydrogen bonds that bond the
two DNA strands together
annealing
The temperature is decreased so that DNA primers can anneal to the ends of the
single strands of DNA
elongation/extension
The temperature is increased to the optimum temperature for DNA polymerase
to build the complementary strands of DNA
This produces the new identical double-stranded DNA molecules
The three stages are undertaken in a PCR machine, or thermal cycler, which
automatically provides the optimal temperature for each stage and controls the length
of time spent at each stage
In each cycle the mass of DNA is doubled, so amplification occurs very quickly

Examiner Tips and Tricks

The details of the PCR technique is beyond the scope of the AP Exam, although you
should familiarize yourself with the concept of PCR and how it is used.

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 Bacterial Transformation
Your notes
Bacterial transformation
Bacterial transformation is a technique that modifies the genome of bacterial cells, e.g.
in order to produce a useful protein that is not bacterial in origin
It involves introducing new DNA into bacterial cells
Bacteria that contain new DNA are said to have been transformed
Bacterial transformation occurs during:
the insertion of human genes into bacterial cells, e.g. the gene for human insulin
1. The insulin gene is inserted into bacterial plasmids, which act as DNA vectors
2. Bacteria take up the plasmids and are transformed
3. The gene can then undergo transcription and translation to produce human
insulin
in vivo gene cloning; this uses living cells to propagate DNA fragments for scientific
study
1. Vectors are used to carry a DNA fragment into bacterial cells, which are
transformed
2. The bacteria reproduce, resulting in many identical offspring that all contain
copies of the DNA fragment
Bacteria are extremely useful for DNA manipulation because:
The genetic code is universal across all types of organisms, meaning that
transformed bacteria can easily produce proteins from other species
There are no ethical concerns over the manipulation of bacterial DNA
The presence of plasmids, which are separate from the main bacterial
chromosome, means that they can act as convenient DNA vectors

Examiner Tips and Tricks

Details of the process of bacterial transformation are beyond the scope of the AP
Exam.

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DNA Sequencing
Your notes
DNA sequencing
DNA sequencing allows the nucleotide base sequence of an organism's genetic
material to be identified
Advances in technology have enabled the development of rapid sequencing methods
which allow scientists to sequence the genomes of organisms very quickly
Most sequencing methods are now automated rather than requiring manual
interpretation
The data obtained from sequencing can be entered into computers with specialized
programmes that compare the base sequences of different organisms

DNA sequencing allows the nucleotide base sequences in different species to be

determined

Uses of DNA sequence data include:

forensic identification
phylogenetic analysis
designing personalised medical care

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 Your notes

DNA sequencing can be used to perform phylogenetic analyses to compare relatedness

of organisms

Examiner Tips and Tricks

The details of DNA sequencing techniques are not required for the AP Exam.

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