Why synthsize genes and genomes? 10 3 10 6 10 5 10 4 -minimal life -genome rewrite -genetic circuits -metabolic pathways -vaccines -gene therapy -individual.

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Why synthsize genes and genomes? minimal life -genome rewrite -genetic circuits -metabolic pathways -vaccines -gene therapy -individual genes -assembly scaffolds DNA length (base pairs)

Error Correction with Flawed Components ACGG TCTG ACTA ACTG Winograd and Cowan (1967) “Reliable Computation in the Presence of Noise” A circuit containing N (error-free) gates can be simulated with probability of error at most , using N log(N/  ) faulty gates, which fail with probability p, so long as p<p th. von Neumann (1956) DNA “consensus vote”

x Mismatch Recognition + Nuclease Activity M N x x M N M N M N Repair Operation Complete error correction when no strand is error-free

x M N Exchange Repair Copies 1 and 2 Repair Operation Complete error correction when no strand is error-free

x M N Exchange Repair Copies 1 and 2 Repair Operation Complete error correction when no strand is error-free

Exchange Repair Copies 2 and 3 Repair Operation Complete error correction when no strand is error-free

Exchange Repair Copies 2 and 3 Current milestone: 1 error per 20,000 bp Next milestone: 1 error per 100,000 bp Repair Operation Complete error correction when no strand is error-free

Why Synthetic Genomes? Drug biosynthesis of new proteins –Nonnatural Amino Acids Optimal codons Genome stability  Safer Bio-isolation Nonnatural amino acids Genome Engineering Cellular Chassis Virus-resistant strains E. colirE.coli

Recoding E.coli: rE.coli E. Coli MG MB Remove tRNAs, tRNA synthetases - three codons “free” - Viral resistance? 2. AGR (R) > CGR (R) Remove RF1 - one codon available for unnatural amino acids - Viral resistance? 1. UAG stop > UAA stop 1. Codons: AGY Ser > UCX Ser 2. tRNAs: AGY Ser > AGY Leu 3. Codons: UUR/CUX Leu > AGY Ser 4. tRNAs: UUR Leu > UUR Ser 5. Codons: UCX Ser > UUR Ser (Leu & Ser now switched) tRNAs: AGY (S) > AGY (L) 3. AGY (S) > UCX (S) tRNAs: UUR (L) > UUR (S) 3. UUR/CUX (L) > AGY (S)