Abstract
BackgroundUnprecedented progresses in high-throughput DNA sequencing and de novo gene synthesis technologies have allowed us to create living organisms in the absence of natural template.Methodology/Principal FindingsThe sequence of wild-type S13 phage genome was downloaded from GenBank. Two synonymous mutations were introduced into wt-S13 genome to generate m1-S13 genome. Another mutant, m2-S13 genome, was obtained by engineering two nonsynonymous mutations in the capsid protein coding region of wt-S13 genome. A chimeric phage genome was designed by replacing the F capsid protein open reading frame (ORF) from phage S13 with the F capsid protein ORF from phage G4. The whole genomes of all four phages were assembled from a series of chemically synthesized short overlapping oligonucleotides. The linear synthesized genomes were circularized and electroporated into E.coli C, the standard laboratory host of S13 phage. All four phages were recovered and plaques were visualized. The results of sequencing showed the accuracy of these synthetic genomes. The synthetic phages were capable of lysing their bacterial host and tolerating general environmental conditions. While no phenotypic differences among the variant strains were observed when grown in LB medium with CaCl2, the S13/G4 chimera was found to be much more sensitive to the absence of calcium and to have a lower adsorption rate under calcium free condition.Conclusions/SignificanceThe bacteriophage S13 and its variants can be chemically synthesized. The major capsid gene of phage G4 is functional in the phage S13 life cycle. These results support an evolutional hypothesis which has been proposed that a homologous recombination event involving gene F of quite divergent ancestral lineages should be included in the history of the microvirid family.
Highlights
The successful development of the high-throughput DNA sequencing technology opened the door for a quantum leap in science advancement [1]
In 2003, Smith et al [7] described a stepwise synthesis of the genome of bacteriophage WX174 just in two weeks
We describe the chemical synthesis of S13-like bacteriophages in the test tubes and characterize their biological properties
Summary
The successful development of the high-throughput DNA sequencing technology opened the door for a quantum leap in science advancement [1]. As genomes of many new species are being sequenced almost daily, genomic data are rapidly accumulating. Thanks to such sequence information, synthetic biologists are attempting to create novel living systems [2,3]. Rapid progress in DNA synthesis has been extended to the level of organism whole-genome de novo synthesis in the absence of a natural template [4,5]. In 2002, work in Wimmer’s group led to the first chemical synthesis of a DNA corresponding to the whole genome of poliovirus [6]. Unprecedented progresses in high-throughput DNA sequencing and de novo gene synthesis technologies have allowed us to create living organisms in the absence of natural template
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.