Place the reagents in tube on ice. Place the reagents in tube on ice.

Collect all the contents of the reagents (EcoRI) tubes by Collect all the contents of the reagents tubes by centrifuging it
centrifuging it briefly. Keep it on ice again. briefly. Keep it on ice again.

In 1 clean eppendorf tube, add 1 µL EcoRI, 2 µL 10X H buffer, In 1 clean eppendorf tube, add 1 µL pGEM-T Easy Vector,
1 µg substrate DNA, and sterile nanopure water up to 20 µL. 5 µL 2X ligation buffer, 1 µL T4 ligase, and 3 µL ligation product.

Incubate the reaction for 37°C for 1 hour. Incubate the reaction at room temperature for 1 hour
or overnight at 4°C (for maximum transformants).

Run the reaction to gel electrophoresis.

Collect and analyze the data.