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10 Mbp of oligos / $300 chip

10 Mbp of oligos / $300 chip. ~1000X lower oligo costs. 8K Atactic/Xeotron/Invitrogen Photo-Generated Acid 12K Combimatrix Electrolytic 44K Agilent Ink-jet standard reagents 380K Nimblegen/GA Photolabile 5'protection.

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10 Mbp of oligos / $300 chip

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  1. 10 Mbp of oligos / $300 chip ~1000X lower oligo costs 8K Atactic/Xeotron/Invitrogen Photo-Generated Acid 12K Combimatrix Electrolytic 44K Agilent Ink-jet standard reagents 380K Nimblegen/GA Photolabile 5'protection Amplify pools of 50mers using flanking universal PCR primers & 3 paths to 10X error correction Tian et al. Nature. 432:1050 Carr & Jacobson 2004 NAR Smith & Modrich 1997 PNAS

  2. Improved Recombination Frequency: 10-4 0.25 (> 3 log increase!) ss-Oligonucleotide Repair DNA Replication Fork Ellis et al. PNAS 2001 Constantino & Court. PNAS 2003 Obtain 25% recombination efficiency in E. coli strains lacking mismatch repair genes (mutH, mutL, mutS, uvrD, dam)

  3. Multiplex Automated Genome Engineering (MAGE) Wash with water & DNA pool (50) Concentrate O-ring membrane Resuspend, bubble, select Concentrate, electroporate Wang, Isaacs, Terry

  4. Multiplex Automated Genome Engineering (MAGE) syringe pump computer communication / data acquisition system electrically actuated valves OD sensor electroporation cuvette w/ membrane filter Wang, Isaacs, Terry

  5. rE.coli: new in vivo genetic codes Freeing 4 tRNAs, 7 codons: UAG, UUR, AGY, AGR e.g. PEG-pAcPhe-hGH (Ambrx, Schultz) high serum stability 4 1 Isaacs Wang Church Forster Carr Jacobson 3 2

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