ArcticZymes R2D Ligaseᵀᴹ

Recombinantly expressed in E. coli.

One milliunit is defined as the amount of enzyme needed to ligate 1 pmol (of 18 pmol) of a nicked DNA substrate in 20 minutes at 25 °C in a 20 µl reaction volume in a buffer consisting of 62.5 mM Tris-HCl, pH 7.5 (25°C), 5 mM MgCl₂, 1 mM ATP, 10 mM DTT, 0.025 mg/ml BSA and 25 mM KCl.

52.9 k Da

ArcticZymes R2D Ligase can be completely inactivated by incubating at 70°C for 10 minutes.

The enzyme is stable at -20°C for 1 year in the supplied storage buffer

2.5 U R2D Ligase was incubated with a supercoiled plasmid (1 µg) for 4 hours at 37°C. Agarose gel electrophoresis did not reveal any transformation of closed circular DNA to nicked DNA.

2.5 U R2D Ligase was incubated with M13 ssDNA (0.5 µg) for 4 hours at 37°C. Agarose gel electrophoresis did not reveal any visible signs of ssDNA degradation.

2.5 U R2D Ligase was incubated with either 3H-dATP labelled ds or ssDNA (0.5 µg, 500 bp) for 4 hours at 37°C. Acid soluble radioactivity from labelled DNA was not significantly over blank test for either substrate

2.5 U R2D Ligase was incubated with a 2 kb RNA transcript (1 µg) for 4 hours at 37°C. Agarose gel electrophoresis did not reveal any visible signs of RNA degradation.

2.5 U R2D Ligase was analysed in a probe-based qPCR assay detecting the 23S ribosomal sub-unit in E. coli. No E. coli gDNA could be detected (LOD: < 3 E. coli genomic copies).