pRK1037

pRK1037

Description: This TVMV protease expression vector is intended to be used for controlled intracellular processing of fusion proteins (TVMV protease substrates) in E. coli.

Origin: A pZ derivative (Lutz and Bujard, 1997)

Replicon: p15A

Antibotic Resistance: Kanamycin

Promotor: Synthetic λP_L/tetO

Inducer: anhydrotetracycline (100 µg/liter)

Notes: Expression of the TVMV protease gene on pRK1037 will be constitutive unless the host strain produces Tet repressor. Regulated expression can be achieved in DH5alphaPro™ or BL21Pro™ cells (Clontech). The specificity of TVMV protease is distinct from that of TEV protease. The canonical recognition site for TVMV protease is ETVRFQS, with cleavage occurring between Q and S. The TVMV protease open reading frame on pRK1037 was constructed from synthetic oligonucleotides and has been codon-optimized for expression in E. coli. For best results, perform intracellular processing experiments at 30 °C.

Literature Citations: Lutz, R. and Bujard, H. (1997). Independent and tight regulation of transcriptional units in Escherichia coli via the LacR/O, the TetR/O and AraC/I1-I2 regulatory elements. Nucl. Acids Res. 25: 1203-10; Yoon, H. Y., Hwang, D. C., Choi, K. Y., and Song, B. D. (2000). Proteolytic processing of oligopeptides containing the target sequences by the recombinant tobacco vein mottling virus NIa protease. Mol. Cells 10: 213-219.

Nucleotide Sequence:

Description:	*This TVMV protease expression vector is intended to be used for controlled intracellular processing of fusion proteins (TVMV protease substrates) in E. coli.*

Origin:	A pZ derivative (Lutz and Bujard, 1997)

Replicon:	p15A

Antibotic Resistance:	Kanamycin

Promotor:	Synthetic λP_L/tetO

Inducer:	anhydrotetracycline (100 µg/liter)

Notes:	Expression of the TVMV protease gene on pRK1037 will be constitutive unless the host strain produces Tet repressor. Regulated expression can be achieved in DH5alphaPro™ or BL21Pro™ cells (Clontech). The specificity of TVMV protease is distinct from that of TEV protease. The canonical recognition site for TVMV protease is ETVRFQS, with cleavage occurring between Q and S. The TVMV protease open reading frame on pRK1037 was constructed from synthetic oligonucleotides and has been codon-optimized for expression in E. coli. For best results, perform intracellular processing experiments at 30 °C.

Literature Citations:	Lutz, R. and Bujard, H. (1997). Independent and tight regulation of transcriptional units in Escherichia coli via the LacR/O, the TetR/O and AraC/I1-I2 regulatory elements. Nucl. Acids Res. 25: 1203-10; Yoon, H. Y., Hwang, D. C., Choi, K. Y., and Song, B. D. (2000). Proteolytic processing of oligopeptides containing the target sequences by the recombinant tobacco vein mottling virus NIa protease. Mol. Cells 10: 213-219.

Nucleotide Sequence: