•Testing competent cells using iGem DNA transformation samples.
• Tested to see if they grew better on a freezer block versus an ice bath.
• Ran colony PCR and plated bacteria on antimicrobial plate.
• cPCR, chloramphenicol, and culturing protocols, made agarose gel
• gel electrophoresis for cPCR, nucleic acid purification plasmid
Week 3 (June 18-22)
•Made chloramphenicol LB plates
•Restriction enzyme digest of iGem plasmid PSB1C3 using EcoRI and PstI
•Made digest master mix; ran an ezyme digest of iGem plasmids PSB1A3 and PSB1K3
Performed ligation of
PSB1A3 and PSB1K3; transformed PSB1A3 in competent cells on ampicillin plate
•Created a 50mg/mL kanamycin solution and then alloquoted it to about 80 1.5mL tubes.
•Inoculated LB with ampicillin colonies (with insert).
•Streaked ampicillin (IA3) colony onto new plates.
•Conducted cPCR for IA3 colonies with insert.
Week 4 (June 25-29)
•Conducted cPCR for PSB1K3.
•Ran 1% agarose gel of PSBIA3 with the insert (colonies that were re-streaked).
•Ran 1% agarose gel of PSBK3 with insert.
•Cleaned laboratory material.
•Set up PCR using Q5 mastermix.
•Amplification of PSB1C3 plasmid backbone.
• Take some and digest with ec and PST according to the IGEM protocol, do more than one tube of it. And then take that and freeze it. The protocol calls for 37C digest for half and hour, no purification is needed. Go back to the plasmids using the PCR(PSB1C3). Take that and do a step, and do a PCR clean up. After clean up use eco and PST and digest that. Purify the PCR product first then add ECO and PST.
