Artificial gene synthesis, or simply gene synthesis, refers to a group of methods that are used in synthetic biology to construct and assemble genes from nucleotidesde novo. Unlike DNA synthesis in living cells, artificial gene synthesis does not require template DNA, allowing virtually any DNA sequence to be synthesized in the laboratory. It comprises two main steps, the first of which is solid-phase DNA synthesis, sometimes known as DNA printing.[1] This produces oligonucleotide fragments that are generally under 200 base pairs. The second step then involves connecting these oligonucleotide fragments using various DNA assembly methods. Because artificial gene synthesis does not require template DNA, it is theoretically possible to make a completely synthetic DNA molecule with no limits on the nucleotide sequence or size.
Synthesis of the first complete gene, a yeast tRNA, was demonstrated by Har Gobind Khorana and coworkers in 1972.[2] Synthesis of the first peptide- and protein-coding genes was performed in the laboratories of Herbert Boyer and Alexander Markham, respectively.[3][4] More recently, artificial gene synthesis methods have been developed that will allow the assembly of entire chromosomes and genomes. The first synthetic yeast chromosome was synthesised in 2014, and entire functional bacterial chromosomes have also been synthesised.[5] In addition, artificial gene synthesis could in the future make use of novel nucleobase pairs (unnatural base pairs).[6][7][8]
^Khorana HG, Agarwal KL, Büchi H, Caruthers MH, Gupta NK, Kleppe K, et al. (December 1972). "Studies on polynucleotides. 103. Total synthesis of the structural gene for an alanine transfer ribonucleic acid from yeast". Journal of Molecular Biology. 72 (2): 209–17. doi:10.1016/0022-2836(72)90146-5. PMID4571075.
^Itakura K, Hirose T, Crea R, Riggs AD, Heyneker HL, Bolivar F, Boyer HW (December 1977). "Expression in Escherichia coli of a chemically synthesized gene for the hormone somatostatin". Science. 198 (4321): 1056–63. Bibcode:1977Sci...198.1056I. doi:10.1126/science.412251. PMID412251.
^Kimoto M, Yamashige R, Matsunaga K, Yokoyama S, Hirao I (May 2013). "Generation of high-affinity DNA aptamers using an expanded genetic alphabet". Nature Biotechnology. 31 (5): 453–7. doi:10.1038/nbt.2556. PMID23563318. S2CID23329867.