Understanding Simple Cells Key Engineering Ideas Removal of unnecessary complexity Simplification with standard reusable parts Hierarchical design Modular construction Standards of construction and measurement ID: 935261
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Slide1
Tom Knight
Ginkgo Bioworks
Understanding
Simple
Cells
Slide2Key Engineering Ideas
Removal of unnecessary complexity
Simplification with standard reusable parts
Hierarchical design
Modular construction
Standards of construction and measurement
Design rules
Carefully defined interfaces
Modeling at the correct level
Isolation of unrelated parts
Flexibility
Slide3Harold Morowitz 1962 & 1984
Slide44
We’ve invested > $10M in building our organism engineering factory
Slide5Design, Build & Test at Ginkgo
Slide6Unit processes
6
Slide7Pipeline for organism domestication
Genome
organism + all strains + all genus
members
Comprehensive, online bibliography
Excellent annotations
, motif finding
Knockout library
growth rate defects
RNAseq for transcript start, stop, abundance Ribosome protection assays Proteins, post-translational modifications, positions Protein complexes Metabolic maps Chip Seq for all DNA binding protein Full scale molecular level simulation Simplified cell customized for engineering
What I cannot create I do not understand
Slide8Automated sample prep & extraction methods demonstrated for many hosts
Metabolite extractionEnzyme activityProtein lysate prepsHT growth assayProduct screen
E. coli
Saccharomyces
cerevisiae
Microalgae
Mesoplasma
florum
Alpha-proteobacteriaSaccharopolyspora erythraea8
Slide9Automated metabolomics pipeline
HR-AM uHPLC-MS on the Thermo Q-Exactive Ppm mass accuracy in the full scan at ~3Hz
Data-dependent and targeted tandem MS/MS fragmentation at ~15Hz
9
Parallel column switching for increasing coverage
Routinely quantify 60-100 central and secondary metabolites
Unbiased feature searching and quantification: 2,500-4000 features detected in the full scan
Slide10Comprehensive metabolic coverage
10
Slide11Ginkgo data-store
& analysis pipeline11
Slide12Additional test pipe capabilities
12
High density culture evaluation
Strain optimization for scale up
Medium/feeding optimization
Proteomics
Enzyme expression and solubility
Global native protein expression
Metabolomics
Global central and secondary metabolism
Pathway/enzyme analysis and optimization
Transcripts and Ribosome Profiling
Organism Domestication
Pathway optimization
Slide13PPP
OH
OH
P
Bacterial transcript
Degradation Fragment
P
OH
Processed RNA
PPP
OH
App
PPP
NH2
NH2
PPP
NH2
OH
//
P
//
RNA Analysis Protocols
Select Fragment Type
Ligate
Adapter
Bind primer, reverse transcribe
Circular ligation, adapter primer PCR
Slide14Ribosome profiling
Micrococcal
nuclease
Proteinase
K
28-30
nt
mRNA transcript
Locate Translation starts, Ribosome stall locations, Frame shifts
To NGS adapter ligation and sequencing
Slide15Pipeline for organism domestication
Genome
organism + all strains + all genus
members
Comprehensive, online bibliography
Excellent annotations
, motif finding
Knockout library
growth rate defects
RNAseq for transcript start, stop, abundance Ribosome protection assays Proteins, post-translational modifications, positions Protein complexes Metabolic maps Chip Seq for all DNA binding protein Full scale molecular level simulation Simplified cell customized for engineering
What I cannot create I do not understand
Slide16Thank you for
your attention