Accurate multiplex gene synthesis from programmable DNA microchips.
Testing the many hypotheses from genomics and systems biology experiments demands accurate and cost-effective gene and genome synthesis. Here we describe a microchip-based technology for multiplex gene synthesis. Pools of thousands of 'construction' oligonucleotides and tagged complementary 'selection' oligonucleotides are synthesized on photo-programmable microfluidic chips, released, amplified and selected by hybridization to reduce synthesis errors ninefold. A one-step polymerase assembly multiplexing reaction assembles these into multiple genes. This technology enabled us to synthesize all 21 genes that encode the proteins of the Escherichia coli 30S ribosomal subunit, and to optimize their translation efficiency in vitro through alteration of codon bias. This is a significant step towards the synthesis of ribosomes in vitro and should have utility for synthetic biology in general.
Tian
Jingdong
J
Harvard Medical School, 77 Ave Louis Pasteur, Boston, Massachusetts 02115, USA.
Gong
Hui
H
Sheng
Nijing
N
Zhou
Xiaochuan
X
Gulari
Erdogan
E
Gao
Xiaolian
X
Church
George
G
D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics; D005796:Genes / Q000235:genetics*; D023281:Genomics / Q000191:economics / Q000295:instrumentation* / Q000379:methods*; D044085:Microfluidics / Q000191:economics / Q000295:instrumentation / Q000379:methods; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000191:economics / Q000295:instrumentation*; D009841:Oligonucleotides / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D014176:Protein Biosynthesis / Q000235:genetics*; D021122:Protein Subunits / Q000235:genetics; D012107:Research Design; D012269:Ribosomal Proteins / Q000096:biosynthesis / Q000235:genetics; D012270:Ribosomes / Q000235:genetics; D012680:Sensitivity and Specificity; D012984:Software
D029968:Escherichia coli Proteins; D009841:Oligonucleotides; D021122:Protein Subunits; D012269:Ribosomal Proteins