Bioengineering Flashcards
What is synthetic biology?
The advent of engineering in biology
or
What synthetic chemistry is to chemistry
What is synthetic chemistry?
Wohler and creating synthetic urea allowed the understanding of chemistry and the making of new things e.g. petroleum made the modern world
What biological techniques were used in the past?
Manipulating biological systems and their characteristics using selective breeding
or
From the 1970s onward, access to the genetic code allowed genetic engineering to occur (cut and paste technology -> DNA sequencing, PCR, restriction enzymes/ligation)
When did synthetic biology start?
When engineers worked in molecular biology labs and found it to be slow, complex and lacking standardisation which meant that there were ill-defined factors and hard to swap parts
Thus the idea to standardise, characterise and refine biology
What is the DBTL cycle?
Design -> define problem, design system, design experiments, host selection
Build -> assemble parts, assemble genomes, modify genomes, automation
Test -> assays, analytical chemistry
Learn -> system analysis, infer design rules
What is reverse engineering?
Back engineering
Understand how an existing system works by taking it apart and using this to modify a system to fit goals
What is forward engineering?
Start with design objectives and employ high-level concepts and models in the design process to predict and build
Why do synthetic biology now?
Can synthesis DNA quickly and cheaply now due to the HGP need for oligonucleotide primers for sequence reactions and this was adapted to synthesise complementary oligonucleotides to combine to create dsDNA
Thus there was no need to extract DNA and clone via PCR and as such no need to only use what nature provides which makes the DBTL cycle much quicker
What is abstraction?
Hide unnecessary details from the designer to allow them to focus on higher level ideas such as parts, devices or systems
Facilitates the building of models via simplification
What is modularity?
Modular parts that can be swapped out
A system is modular if it can be divided into strongly interacting parts that are autonomous
What is characterisation?
The need to know what each part of the DNA does via a process that establishes, maintains and controls certain biological characteristics in a defined system
The environmental, genetic and operator context and how they affect designs
What is standardisation?
Agreed standards for parts, methods, data sharing, model sharing and results
E.g. SBOL
Describe BioBricks RFC10
Prefix and suffix sequences flank every part and contain sites for EcoRI, XbaI, SpeI and PstI, thus the system is idempotent with EX on one side and SP on the other.
The enzymes recognise different sequences but leave compatible overhangs. When they anneal they leave a scar that can affect the functionality of the system. The scar isn’t recognised by any of the enzymes so EXSP can’t be included in the sequence thus is edited out by either mutation or in silico.
Thousands of parts are on the Registry of Standard Biological Parts.
Is modular and idempotent so it is easy to build large systems.
Doesn’t work well and can only include 3 parts, one of which is the plasmid vector.
Was the original standardised cloning method
Describe Golden Gate
Uses Type IIS restriction enzymes that recognise asymmetric sequences and cut outside the recognition site thus the overhang is decided by the designer which means that the same enzyme can be used to cut multiple fragments, all of which will have different overhangs. This allows the ligation and digestion to occur in one step.
If the fragments religate, the recognition site is remade so the enzyme can cut again.
Is scarless and efficient and rapid but not modular (though can be made modular through MoClo/EcoFlex/Phytobricks) and overhangs have to be designed everytime.
Not 100% sequence independent thus need to ensure ends are compatible but don’t include recognition sequence or ligate incorrectly
Describe GG phytobricks/MoClo
Have arbitrary sequences that must be present as the prefix/suffix to given parts thus allows for modularity.
Introduces small scar that can affect the functionality but is highly efficient and automatable
MoClo loses recognition site when constructs are made but this can be avoided by alternating between 2 or more standard vectors at different levels
