Team:UCLA/Notebook/Protein Cages/7 August 2015

iGEM UCLA




Introduction: Today gel extraction and digestion of PCquad 3.0 will be done. If permitting, ligation will also be done.

Procedures: Gel extraction was performed as indicated by zymo procedures. The concentration was measured using the nanodrop. 78.78ng/uL. 260/280 = 3.23.

Since the yield was too low, another round of amplification will need to be done before digestion.

Conclusions: There weren’t any restriction enzymes available to use, so digestion will be done next week. PCR amplification will be done in a larger volume for PCquad 3.0 as well.


Intro: Performed a 50 uL PCR amplification of Mutant #10. PCR cleanup.

50 uL PCR:

10 uL 5x Q5 buffer

5 uL 2mM dNTPs

2.5 uL 10 uM forward primer

2.5 uL 10 uM reverse primer

0.5 uL 1 ng/uL template DNA

0.5 uL Q5 Polymerase

29 uL ddH2O (to 50 uL)

This was mixed in a master tube and added to two different PCR tubes.

98C 30s

98C 10s }

Tm 15s } 25x

72C 30s }

72C 5min


PCR Cleanup: Performed according to recommended instructions.