Synthetic biology and DNA assembly

1.

DATE 03-06-2025
synthetic biology
and DNA assembly
By Ana and Maki

2.

SYNTHETIC BIOLOGY
• What is it? Engineering biological systems for new functions and
applications.
• Key Uses: Medicine (gene therapy, synthetic vaccines), agriculture,
biofuels, and environmental solutions.
• Future Potential: Synthetic
sustainable materials.
organisms,
bio-computing,
and

3.

DNA ASSEMBLY
• Purpose: Constructing custom DNA
engineering and synthetic biology.
sequences
for
genetic
• Methods: Golden Gate Assembly, Gibson Assembly, and Bio-bricks.
• Advancements: Automation, AI-driven design, and improved
accuracy.

4.

Costs
COSTS & ACCESSIBILITY
INNOVATIONS
• DNA synthesis costs dropping,
enabling whole genome
projects.
• Advances in automation & AI
lower labor expenses.
• Gene synthesis optimized via
polymerase cycling &
microarray techniques.
Easier Access Innovations
• In vivo assembly in yeast & bacteria for
large-scale DNA construction.
• Combinatorial techniques (Golden
Gate, Gibson, SLIC) streamline parallel
DNA assembly.
• Standardized DNA parts (BioBrick™,
BglBrick) enable modular plug-andplay engineering.

5.

TRADITIONAL MULTIPLE CLONING VS.
MODERN ASSEMBLY METHODS
Traditional Multiple Cloning
Modern DNA Assembly Methods
• Uses restriction enzymes to cut DNA • Gibson Assembly: Seamless joining using
exonuclease, polymerase, and ligase.
and ligase to join fragments.
• Golden Gate Assembly: Modular Type IIs
• Requires specific restriction sites,
enzyme-based assembly, scarless and
limiting flexibility.
efficient.
• Leaves scar sequences, which may
Yeast
Homologous
Recombination:
In
affect gene function.
vivo assembly of large constructs and
synthetic genomes.

6.

EXAMPLE OF CIRCUIT

7.

BIOBRICK ASSEMBLY MECHANISM
• Standardized DNA Parts
BioBricks are modular genetic components, each with specific restriction enzyme sites (EcoRI,
XbaI, SpeI, PstI).

8.

BIOBRICKS LIMITATIONS
• Scar sequences – Leaves an 8-base pair scar, which can interfere with gene
function.
• Labor-intensive – Requires multiple restriction digestions and ligations, slowing
down assembly.
• Limited flexibility – Depends on specific restriction sites, restricting design
choices.
• Not ideal for large constructs – Sequential assembly makes whole-genome projects
difficult.
• Burden on host cells – Reduces bacterial growth rates, leading to escape mutants.

9.

GIBSON ASSEMBLY
• Gibson Assembly enables seamless DNA assembly by using
exonuclease, polymerase, and ligase in a single reaction.

10.

GIBSON ASSEMBLY LIMITATION
• Fragment Number Limit – Efficiency decreases when assembling more than
~8 fragments due to complexity.
• Sequence Constraints – Requires precise overlapping sequences, limiting
flexibility
• Error Risk – Can introduce mutations at fragment junctions, affecting accuracy.
• Not Ideal for Short Fragments – Standard Gibson Assembly struggles with
single-stranded or very short DNA inserts.
• Optimization Challenges – Requires careful primer design and reaction
conditions for high efficiency.

11.

GOLDEN GATE ASSEMBLY
• Golden Gate Assembly enables efficient one-step DNA assembly using
Type IIs restriction enzymes and ligase.

12.

GOLDEN GATE ASSEMBLY
• Sequence Constraints – Requires specific restriction sites, limiting flexibility
• Efficiency Issues – Assembly success decreases with increasing fragment
number, especially beyond 10 fragments.
• Scarless but Error-Prone – While it avoids unwanted sequences, ligation errors
can occur at junctions.
• Optimization Challenges – Requires careful design of overhangs to ensure
correct assembly.
• Not Ideal for Large Constructs – Works best for modular cloning, but
struggles with whole-genome assembly.

13.

IMPORTANCE
• Enables precise DNA editing and manipulation.
• Facilitates modular cloning, allowing scalable genetic
engineering.
• Supports synthetic biology, advancing medicine, biotechnology,
and bioengineering.
• Improves efficiency in assembling complex genetic constructs for
research and applications.

14.

AREAS OF USE
• Biomedical Research – Gene therapy, vaccine development, and
synthetic genome projects.
• Industrial Biotechnology – Engineering microorganisms for
biofuel production and pharmaceutical synthesis.
• Agriculture – Developing genetically modified crops with
enhanced traits.
• Environmental Science – Bioremediation using engineered
microbes to clean pollutants.

15.

FUTURE IMPLICATIONS
• Scalable genome assembly – Enabling synthetic organisms for
tailored medical and industrial applications.
• Automated DNA design – AI-assisted genetic engineering for
faster and more precise DNA synthesis.
• Advanced gene therapies – Personalized medicine using
engineered gene circuits for targeted disease treatment.
• Synthetic biosensors – Using engineered DNA constructs for realtime monitoring of environmental and biological conditions

16.

Thank You