Amber codon is genetically unstable in generation of premature termination codon (PTC)-harbouring Foot-and-mouth disease virus (FMDV) via genetic code expansion

Rongzeng Hao, Kun Ma, Yi Ru, Dan Li, Gaoyuan Song, Bingzhou Lu, Huanan Liu, Yajun Li, Jiaoyan Zhang, Chunping Wu, Guicai Zhang, Haitao Hu, Jianxun Luo, Haixue Zheng

Research output: Contribution to journalArticlepeer-review


The foot-and-mouth disease virus (FMDV) is the causative agent of FMD, a highly infectious and devastating viral disease of domestic and wild cloven-hoofed animals. FMD affects livestock and animal products’ national and international trade, causing severe economic losses and social consequences. Currently, inactivated vaccines play a vital role in FMD control, but they have several limitations. The genetic code expansion technology provides powerful strategies for generating premature termination codon (PTC)-harbouring virus as a live but replication-incompetent viral vaccine. However, this technology has not been explored for the design and development of new FMD vaccines. In this study, we first expanded the genetic code of the FMDV genome via a transgenic cell line containing an orthogonal translation machinery. We demonstrated that the transgenic cells stably integrated the orthogonal pyltRNA/pylRS pair into the genome and enabled efficient, homogeneous incorporation of unnatural amino acids into target proteins in mammalian cells. Next, we constructed 129 single-PTC FMDV mutants and four dual-PTC FMDV mutants after considering the tolerance, location, and potential functions of those mutated sites. Amber stop codons individually substituted the selected amino acid codons in four viral proteins (3D, L, VP1, and VP4) of FMDV. We successfully rescued PTC-FMDV mutants, but the amber codon unexpectedly showed a highly degree of mutation rate during PTC-FMDV packaging and replication. Our findings highlight that the genetic code expansion technology for the generation of PTC-FMD vaccines needs to be further improved and that the genetic stability of amber codons during the packaging and replication of FMDV is a concern.

Original languageEnglish (US)
Pages (from-to)2330-2341
Number of pages12
JournalRNA Biology
Issue number12
StatePublished - 2021
Externally publishedYes


  • Amber codon
  • foot-and-mouth disease virus
  • genetic code expansion
  • genetic stability
  • premature termination codon (ptc)

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology


Dive into the research topics of 'Amber codon is genetically unstable in generation of premature termination codon (PTC)-harbouring Foot-and-mouth disease virus (FMDV) via genetic code expansion'. Together they form a unique fingerprint.

Cite this