BspPI (AlwI) (2 U/µL)

Protocol tips

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	The final PCR product from amplification of PON1 was subjected to digestion by BspPI Restriction Endonuclease (RE) enzyme (Thermo Scientific). A 10 μL of PCR reaction mixture was mixed with 2 U/μL of BspPI RE enzyme, 2 μL of 10X Buffer Tango and 18 μL of NFW. The mixture was incubated at 55°C for 16 hours. The enzyme was then inactivated by incubation at 80°C for 20 minutes. The digested product was visualised after electrophoresis in Ethidium Bromide (EtBr)-stained 2.5% agarose gel on gel documentation system (UVI-TEC).

Protocol tips

The final PCR product from amplification of PON1 was subjected to digestion by BspPI Restriction Endonuclease (RE) enzyme (Thermo Scientific). A 10 μL of PCR reaction mixture was mixed with 2 U/μL of BspPI RE enzyme, 2 μL of 10X Buffer Tango and 18 μL of NFW. The mixture was incubated at 55°C for 16 hours. The enzyme was then inactivated by incubation at 80°C for 20 minutes. The digested product was visualised after electrophoresis in Ethidium Bromide (EtBr)-stained 2.5% agarose gel on gel documentation system (UVI-TEC).

Publication protocol

"PCR amplification was done using thermo cycler (Eppendorf Mastercycler Gradient-5331). Briefly, 50 ng of DNA was amplified in a 20 μL reaction mixture containing 15 pM of each of the PON1 primers, 1.25 mM MgCl2, 200 μM Deoxynucleotide Triphosphates (dNTPs), 2 μL 10X PCR buffer (10X buffer containing 500 mM KCl, 100 mM Tris-HCl, pH 9.0), and 1 U/μL Taq DNA polymerase (All reagents form MBI Fermentas). The PCR protocol included an initial incubation at temperature of 94°C (five minutes) followed by 35 cycles of amplification (denaturation for 30 seconds at 94°C, annealing for 30 seconds at 60°C and extension for 30 seconds at 72°C). A final 10 minutes extension step was done at 72°C. The primer sequences used are given in [Table/Fig-1]. The final PCR product from amplification of PON1 was subjected to digestion by BspPI Restriction Endonuclease (RE) enzyme (Thermo Scientific). A 10 μL of PCR reaction mixture was mixed with 2 U/μL of BspPI RE enzyme, 2 μL of 10X Buffer Tango and 18 μL of NFW. The mixture was incubated at 55°C for 16 hours. The enzyme was then inactivated by incubation at 80°C for 20 minutes. The digested product was visualised after electrophoresis in Ethidium Bromide (EtBr)-stained 2.5% agarose gel on gel documentation system (UVI-TEC)."

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