User:Eric M. Walters/Notebook/Spring 2012/2012/04/09

Cyanobacterial acrylate production

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Amplifying the three parts 100 uL reactions of each (50x2) This is because all of the old construct was transformed into strains which had lost their MCR (& Kan) acsA_up & acsA_down 1 ul forward primer (300 uM stock) 1 ul reverse primer (300 uM stock) 1 ul PCC 7002 gDNA 1 ul dNTP mix 1 ul Phusion 10 ul HF buffer 35 ul Water 98°C for 2:00 98°C for :30 60°C for :30 72°C for :15 Cycle 35x 72°C for 4:00 4°C forever accBCDA 1 ul forward primer (300 uM stock) 1 ul reverse primer (300 uM stock) 1 ul pΨ223 (100 uM stock) 1 ul dNTP mix 1 ul Phusion 10 ul HF buffer 35 ul Water 98°C for 2:00 98°C for :30 60°C for :30 72°C for 1:10 Cycle 35x 72°C for 4:00 4°C forever