Genetic engineering is the process of inserting a gene from
one organism into the DNA of another. This makes the modified cell capable of
producing a new and useful protein—something it could not do before. This
technique forms the foundation of modern biotechnology.
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| Formation of recombinant DNA |
Understanding Recombinant DNA (rDNA)
Recombinant DNA, often abbreviated as rDNA, is a strand of
DNA formed by combining genetic material from two or more sources. This new DNA
can be inserted into a host cell, allowing the host to express a gene that
wasn’t originally part of its genome.
What Are Vectors?
Vectors act as carriers for transferring genes. A common
vector is a plasmid, a small circular DNA molecule found in bacteria
that replicates independently of the bacterial chromosome. Viruses, such as phages,
can also serve as vectors. They inject their DNA into host cells, and this DNA
can be engineered to include foreign genes.
When a plasmid is combined with foreign DNA using genetic
tools, it becomes recombinant DNA. Once inserted into a bacterium, this plasmid
replicates as the bacterium divides—cloning the inserted gene along the way.
The Five Key Steps of Genetic
Engineering
- Isolate
the Gene: Extract the desired gene from
the donor organism.
- Insert
into a Vector: Introduce this gene into a
plasmid or virus.
- Transfer
to Host Cell: Move the vector into a bacterial
host.
- Identify
Modified Cells: Screen for host cells that
accepted the foreign DNA.
- Clone
the Gene: Allow the modified cells to
reproduce, copying the gene.
Restriction Enzymes: Nature’s Molecular
Scissors
Genetic engineers rely on restriction enzymes—proteins that
naturally occur in bacteria—to cut DNA at specific sequences known as recognition
sites. These sites are often palindromic sequences, meaning the
sequence reads the same forward and backward.
One commonly used enzyme is EcoRI, which recognizes
and cuts at the sequence GAATTC. The cuts produce sticky ends—single-stranded
overhangs that help link DNA from different sources.
How DNA Fragments Are Joined
Once the sticky ends of different DNA strands find their
match, the enzyme DNA ligase seals them together, forming strong
covalent bonds. This final product is recombinant DNA, a hybrid molecule
with genes from multiple sources.
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| Steps showing formation of rDNA using restriction enzyme and DNA ligase |
How Recombinant DNA is Cloned
To clone a gene:
- Isolate
the desired gene and plasmid vector.
- Cut
both with the same restriction enzyme to create matching sticky ends.
- Mix
them together so the sticky ends pair up.
- Use
DNA ligase to permanently join the fragments.
- Insert
the recombinant plasmid into bacteria via transformation.
- Allow
the bacteria to multiply, copying the foreign gene during cell division.
Gene Libraries and cDNA
Plasmid and Bacteriophage Libraries
- Plasmid
libraries: Collections of bacteria carrying
different DNA fragments.
- Phage
libraries: Collections of viruses
engineered with DNA fragments.
These libraries hold an entire organism’s genome broken into
smaller segments, ready for analysis or future use.
Using cDNA Instead of Genomic DNA
Bacteria cannot process introns—non-coding regions in DNA.
To solve this, scientists use reverse transcriptase to convert mature
mRNA into complementary DNA (cDNA), which contains only the coding
regions of a gene. This cDNA can then be cloned and expressed in bacterial
cells.
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| Cloning a Gene in a Bacterial Plasmid |
Finding a Gene with a Probe
If scientists already know part of a gene’s sequence, they
can create a probe—a short piece of RNA or DNA that matches the
sequence. These probes are labeled with radioactive or fluorescent tags. When
added to a sample, they bind to the target gene through base pairing, allowing
researchers to locate and isolate it for further study.
The Polymerase Chain Reaction (PCR)
What Is PCR?
Developed by Kary Mullis in 1983, PCR is a method that
allows scientists to make millions of copies of a specific DNA segment in a
test tube. This is done without needing to clone the gene in a living cell.
How PCR Works
PCR uses:
- Primers:
Short sequences that match the start and end of the target DNA.
- Taq
polymerase: A heat-resistant enzyme from hot
spring bacteria.
- Thermocycler:
A machine that rapidly heats and cools samples to carry out DNA
replication.
This method is highly specific and efficient, allowing
scientists to study very small amounts of DNA.
DNA Fingerprinting and RFLPs
When DNA is treated with restriction enzymes, it breaks into
fragments of different lengths. These fragments vary from person to person due
to differences in DNA sequences, known as restriction fragment length
polymorphisms (RFLPs).
Using gel electrophoresis, the fragments are
separated by size, creating a unique pattern—similar to a barcode—that can be
used for:
- Criminal
identification
- Paternity
testing
- Diagnosing
genetic diseases
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| Preparation of a genomic library |
Sequencing DNA with Gel Electrophoresis
In this process, DNA fragments move through a gel under
electric current. Smaller fragments move faster and farther. This allows
scientists to identify and analyze the DNA based on the pattern of separated
fragments. The gel is later stained to visualize the results.
Real-World Applications
Genetic engineering is not just a scientific marvel—it’s a
tool with real-life impact. Its uses include:
- Creating
insulin and other medications
- Identifying
criminals and victims through DNA evidence
- Tracking
genetic diseases and mutations
- Mapping
evolutionary relationships
- Developing
genetically modified crops and organisms
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| Reverse Transcriptase helps make DNA for cloning |
Key Takeaways for Curious Minds
- Genetic
engineering makes it possible to modify life
at the molecular level.
- Recombinant
DNA technology is the foundation of modern
biotechnology and medicine.
- PCR
revolutionized how we copy and study DNA—quickly, accurately, and in tiny
amounts.
- Restriction
enzymes and DNA ligase act like
scissors and glue to build new DNA.
- Probes
and DNA fingerprinting help solve crimes, track diseases, and study
evolution.
With these technologies, we're not just studying life—we’re
reshaping it for better health, safer environments, and a deeper understanding
of who we are.
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| Identification of a cloned gene |
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| Polymerase chain reaction (PCR) |
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| Gel electrophoresis |
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