MluI restriction enzyme

Protocol tips

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	Plasmid pAsi, the infectious cDNA clone of Asia1 FMDV Asia1/YS/CHA/05, was digested with MluI and EcoRV or SmaI and MluI (TaKaRa, Dalian, China). Positive plasmids bearing the desired mutation(s) in 3Dpol were digested with MluI and EcoRV or SmaI and MluI and reintroduced into pAsi that had been digested previously with the same restriction endonucleases. The recombinant plasmids were used for in vitro transcription and transfection.

Protocol tips

Plasmid pAsi, the infectious cDNA clone of Asia1 FMDV Asia1/YS/CHA/05, was digested with MluI and EcoRV or SmaI and MluI (TaKaRa, Dalian, China). Positive plasmids bearing the desired mutation(s) in 3Dpol were digested with MluI and EcoRV or SmaI and MluI and reintroduced into pAsi that had been digested previously with the same restriction endonucleases. The recombinant plasmids were used for in vitro transcription and transfection.

Publication protocol

Plasmid pAsi, the infectious cDNA clone of Asia1 FMDV Asia1/YS/CHA/05, was digested with MluI and EcoRV or SmaI and MluI (TaKaRa, Dalian, China). The resulting fragment was ligated, using T4 DNA ligase (New England BioLabs, Ipswich, MA), into a pOK12 vector that had been previously digested with the same restriction endonucleases. The resulting plasmid containing the 3Dpol gene was used as the template for site-directed mutagenesis using the series of primers listed in Table 1. Positive plasmids bearing the desired mutation(s) in 3Dpol were digested with MluI and EcoRV or SmaI and MluI and reintroduced into pAsi that had been digested previously with the same restriction endonucleases. The recombinant plasmids were used for in vitro transcription and transfection.

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