•1µl of plasmid DNA was added to 50µl of competent cell (Xl-1Blue/ Stbl).
•Incubated on ice for 30 min.
•Heat shock at 45ºC for exact 90 sec.
•Incubated on ice for 2 min
•500ml of LB broth was added
•Incubated at 30ºC (Stbl)/37ºC (XL-1Blue), 200rpm for 1hr.
•Spread on LB agar plate containing appropriate antibiotic and allowed to dry.
• Incubated at 30ºC (Stbl)/37ºC(Xl-1Blue/ DH5α) for 18-20 hrs