== Notebook: Week 10 ==
''Friday''
- Planned/mapped out next 4 weeks
- Took dialyzed bag with purified eGFP-RiAFP and divided it into 12 eppendorfs, containing 1 mL each. Flash-froze tubes in liquid nitrogen, then quickly used a hot edge to poke holes into top of eppendorfs to allow sublimation. Tubes then placed in lyophilizer for 3 nights.
''Saturday''
- Began cloning all pSB1AK8 constructs into pSB1C3 for parts submission (G3, H1, RiA, RiG, T2, Z2?):
- Digested 6 constructs and linearized pSB1C3 with EcoRI, PstI
- Also, PCR amplified eGFP segment from original eGFP-RiAFP plasmid to use as a control for survival assays
- Digested PCR fragment with eGFP with XbaI, PstI, and digested BBa_I712074 (strong T7 promoter) with SpeI and PstI
- Gel extracted all digested pSB1AK8 constructs using kit, gel visualized with SYBRSafe (digestion seemed successful)
''Sunday''
- Ligated gel-extracted pSB1AK8 fragments with pSB1C3; ligated eGFP PCR fragment with BBa_I712074
- Transformed ligations into DH5alpha cells, plated them
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