TEMPase Hot Start Master Mix BLUE

Protocol tips

Upstream tips	Protocol tips	Downstream tips
Amplicon size should be consistent for each target sequence and limited to approximately 65 - 100 bp.	Working on ice is not required.

Upstream tips
Amplicon size should be consistent for each target sequence and limited to approximately 65 - 100 bp.
Protocol tips
Working on ice is not required.

Publication protocol

Genomic DNA was extracted using the Dynabeads DNA DIRECT Universal kit (Invitrogen) on a 24 h culture. For each PCR reaction, 1 μL of extracted genomic DNA was used with primers listed in . PCR for the species identification and MLST were performed using the TEMPase Hot Start 2X Master Mix Blue II (Ampliqon, Denmark) and the Maxima Hot Start Taq Polymerase (Fermentas, Waltham, MA, United States) was used for toxin-encoding gene amplification, both were performed in a 25 μL reaction following the manufacturer’s instruction. Reactions were done in a Veriti Thermal Cycler (Applied Biosystems, 96 Well Model 9902). PCR products were purified using 10 U Exonuclease I (Thermo Fisher Scientific, Waltham, MA, United States) and 0.5 U Fast Alkaline Phosphatase (Thermo Fisher Scientific, Waltham, MA, United States) and incubated at 37°C for 15 min followed by an inactivation step at 85°C for 15 min (). Macrogen Europe (The Netherlands) sequenced the gene fragments using the same primer used for the PCR amplification.

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Papers

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Manufacturer protocol

Download the product protocol from Ampliqon for TEMPase Hot Start Master Mix BLUE below.

Download manufacturer protocol

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