Synthetic biology Exam Flashcards
What Chssis was used in the iGEM project?
Bacillus subtilis WB800
What are the Benefits and drawbacks of using a Bacillus subtilis WB800 chassis?
Benefits: large scale fermentation, non-pathogenic, Food-grade, and Absorb foreign material which can be maintained for 100generations
Negative: Proteases that degrade foreign material
How does the final cell from the 2019 iGEM project work?
Stores the proinsulin in the cytoplasm
Loaded onto the PET membrane of the chip, promote the liver cell to synthesize glycogen from glucose in the lumen
Release of proinsulin from external lysosome which is heat controlled
How many restriction sites are in the genome for the iGEM 2019 project?
9
What are the restriction sites are in the genome for the iGEM 2019 project?
Restriction site: contains 9 restriction sites
AlwNI
ApaBI
BglII
Bsp1407I
CspI
Eco57I
PvuII
XmnI
What is the input of the synthetic cell of the 2019 iGEM project?
The input for synthetic system would be elevated blood sugar or elevated proinsulin would create a negative feedback loop.
What tool is utilised in the 2019 iGEM project?
Organ-on-a-chip: a microfluidics device that can be used invitro to load the cellular chassis with the synthetic biological circuit for diabetic treatment. used was hepatic cells.
What is the 2019 project?
The iGEM project from 2019 was the integration of biology and synthetic biology to create a “smart” bacterium.
Creating a self-regulated pro-insulin cell, which would be implanted in the gut, to compensate for the lack of pancreatic production.
What is the HPI gene created in the 2019 iGEM project?
HPI GENE: 254 bp: created for heightened expression and compatibility.
What was modified for better gene transcription in the iGEM project?
N-Terminus: Modifications made to the N terminus chassis obtained a more efficient gene expression
What Three biobricks were used in the project?
RFC 10 compatible: RFC 10 is a BioBrick®
P43 Promotor: BioBrick®
Spacer Peptide: 7 amino acids – BioBrick®
What are the advantages and disadvantages of the RFC Compatible biobric?
Advantage:
Standard
Well tested and documented
native protein start codon can be preserved while using RBS parts
large and still growing set of parts
Disadvantage: no protein fusion (stop codon in 6bp scar, frameshift and stop codon in 8bp scar)
Why was he P43 Promoter chosen for the project?
strong promoter of Bacillus subtilis expression system.
This promoter has been shown to be recognized and active during the exponential and lag phases of growth.
Synthetic improved cytidine deaminase (cdd) gene in B. subtilis used to express targeted proteins
Polymerase Per Second (PoPS) generator.
Higher Quantities: Produces higher quantities of product compared to other synthetic promoters
Why was the spacer [peptide biobrick used?
7 amino acids – BioBrick®
Increase of proinsulin: significant increase and is compatible with the N-terminus of the HPI gene
What vector was chosen?
Vector: pHY-P43 transformation vector: Human proinsulin gene (HPI) was constructed on the ends of BamHI and EcoRI of the pHY-P43 transformation vector
What are the three main areas of Ethics to be concerned with?
Biosafety
Bioethics
Biosecurity
What are the two main Biosaftey areas to be concerned with ?
Human health and environmental risks
Who did the Biosaftey experiment?
Hewit et all 2016
Biosaftey: Human Health what are the foiur concerns?
allergies
pathogens
antibiotic resistance
carcigens
Environmental risks what are the four ethical concerns?
horizontal gene transfer
toxicity
Impact on ecosystem
competition from nature
What are the 5 principle bioethics to consider?
Intelectual freedom
Public benefits
Responsible stewardship
Democratic deliberation
Justice and fairness
Who determined the 5 bioethical principles?
Presidential commission for the study of bioethical issues
What issues is there with biosecurity?
Dual use problem
Increased Hazard controls
Biosecurity: Dual use problems. What study says its ok.
Kevin SMith (2022) ethical matrix
What biosecurity increased hazard controls are there?
Intrinsic biocontainment
Auxotroph
Kill switches
Inability to replicate or pass genes to off spring
Xeno biological organisms: xeno nucleotide
Who developed the future extracellular matrix?
liu et al (2022)
Who told me about the future, with Artificial intelligence and syn-biopython?
Yeoh(2021)
Who told me about machine learning and it being 77% better then traditional random screening?
HamediRad et al. (2018)
What are the different new ways to standardise things?
Automation
biofoundaries
cloud libraries
Interchangable parts
Cello 2.0
SEGA
Curated Bricks
SEVA
Bioroboost
What is the benifit of having standard interchangeable parts?
Interchangeable parts (nearly identical)can successfully substitute the similar parts and fit into the assembly of the same type.
Mass production of identical items within the prescribed limits of sizes.
Interchangeability allows mass production, reduces the production cost.
Tell me about cello 2.0
DNA sequences for programmable circuits based on a high-level software description and a library of characterized DNA parts.
Design specification reuse, modular DNA part library curation and formalized circuit transformations based on experimental data.
Jobs may take minutes or hours to complete.
Tell me about sega: Standardised Genome Archetecture
Enables genome engineering by combining only two reagents: a DNA fragment from a commercial vendor and a stock solution of bacterial cells, incubated on agar plates.
The modular nature of SEGA allows precise multi-level control of transcriptional, translational, and post-translational regulation.
How do curated bio bricks help?
Accessible through an open database easier to understand and reuse.
Key features of Model Bricks: Standard language (SBML), rule-based specification describing species as a collection of uniquely identifiable molecules, association with model specific numerical parameters, and more common annotations.
Tell me about the SEVA?
European Vector Architecture (SEVA): plasmids
What is bioroboost?
(https://standardsinsynbio.eu), have been established with the main purpose to promote standardization in synthetic biology.