Chassis Parts jukebox : Meta-device : few strains Focus of system designs & integration...
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Transcript of Chassis Parts jukebox : Meta-device : few strains Focus of system designs & integration...
Chassis
• Parts jukebox : Meta-device : few strains
• Focus of system designs & integration
• Standardization, composability, abstraction
• Interface with environment, power supply
• Interface with computers: CAD, measurements
• Human practice: Safety & process engineering
Chassis Goals
Genome Engineering - Multiplex Automation • Accelerated evolution (GE-MASS targeting) • Decelerated evolution in production (e.g. Tn- & antimutators)• Data > model > CAD for efficient allele replacement Why genome-wide chassis engineering?• Codon changes for multi-virus resistance• Eliminate transposons (Tn-)• Change restriction sites• Orthogonal expression systems & compartments
Synergy with Thrusts & Testbeds
• Parts & Devices – chassis for surface display devices, eliminate restriction sites (CTAG); Construction Optimization Group (COG) meetings; IGEM
• Human Practices – motivation for genome isolation (via codons), ecological-testing, http://www.ia-sb.eu/
• Tumor-killing bacterium (TKB) – inherent release hence safety features – metabolic “disabling tech”
• Microbial Chemical Factory (MCF) – multi-promoter & multi-RBS tuning via GE-MASS; Tn- stability
Thrust 3: Chassis Design & Characterization 11:00-11:20 AM Tue 4-Mar-2008Carlos Bustamente, Anton Vila-Sanjurjo – mt-minigenome Drew Endy, Barry Canton – orthogonal expressionTom Knight – Mesoplasma florum Farren Isaacs, Harris Wang – MAGE & new codesDuhee Bang, Mike Jewett - in vitro & chiral engineering
Thrust 3: Chassis Design & Characterization 10:00-10:30 AM Tue 21-Sep-2008Brad Zamft, Carlos Bustamante, Natalie Kuldell– The Mitochondria as a Minimal Chassis: Expanding the Toolkit for Mitochondrial Genomic EngineeringFarren Isaacs &Harris Wang – GEMASS & new codesMike Jewett - in vitro chassis
Why Minigenomes
Tom Knight Mesoplasma florum ~750 genesCarlos Bustamante Mitochondrion ~1500 genesMike Jewett E.coli in vitro 151 genes
• Ease of synthesis• More radical designs • Existing applications -Roche Rapid Translation System (RTS) -Ambion ActivePro In Vitro Translation Kit -Novagen, Promega, Invitrogen, Qiagen Stratagene, Paragon, Amersham
113 kbp DNA 151 genes
Pure translation:Forster & Church
MSB ’05 GenomeRes.’06
Shimizu, Ueda ’01
Not minimal:High speed &
accuracy requires a few extra genes
(E.coli 20 min. doubling)
Reconstituted ribosomes:Jewett & Church
Despite early success in 1968, assembly of ribosomes under
conditions compatible with protein synthesis has been elusive
(until Aug 2008)
50S and 30S ribosomal subunits simultaneously reconstituted at
37°C and constant [Mg2+] express active luciferase
50S and 30S ribosomal subunits simultaneously reconstituted at
37°C and constant [Mg2+] express active luciferase
0
50
100
150200250
300350
Luciferase(ng/mL)
0102030405060708090
Luciferase(ng/mL)
Improving process yield & safety: trade-off
or win-win?
What do most biological systems fear?
What do all viruses have in common?
.