User:Andrew Perry/Notebook/bbGibson/2011/08/05

bbGibson assembly

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PCR amplification of ethylene device parts - His-tag and RBS addition PCR program as per User:Andrew_Perry/Notebook/bbGibson/2011/06/08 (touchdown 55°Cx3 - 65°Cx25, 4min extension time) PCR reactions (50 μL) addition RBS and His6 to ethylene device parts. Agarose gel electrophoresis (1 %) stained with EZ-Vision. 0. Mass Ruler marker (Fermatas) 1. pSB1C3-SAMsynth (BBa_K417000) His6-SAMf/END.r, 2. pMA-ACCsynth His6-ACCsf/END.r, 3. pMA-ACCoxidase His6-ACCof/END.r, 4. (1) + BBpre-RBS-N-His6, 5. (2) + BBpre-RBS-N-His6, 6. (3) + BBpre-RBS-N-His6. ACCoxidase + His6 PCR gave correct sized product SAMsynth + His6 + RBS PCR gave correct sized product ACCoxidase + His6 + RBS PCR gave correct sized product All of the ACCsynth PCR reactions failed.