Registry/Measurement kit/Notebook/2007-7-31

E0240

• Noticed that during the scarring PCR, a point substitution was made.
• Decided to try a site-directed mutagenesis on the biobricked product.
• Ordered two pairs of primers:
  • First pair - mutation in the ends of the primers:
    • CGAAAGATCCCAACGAAAAGAGAGACCACATGGTCCTTCTTGAGTTTGTAACAGC, E0240_mut_E_F (78.8)
    • CTCTCTTTTCGTTGGGATCTTTCGAAAGGGCAGATTGTGTGGACAGG, E0240_mut_E_R (79.7)
  • Second pair - mutation in the middle of the primers:
    • CCTTTCGAAAGATCCCAACGAAAAGAGAGACCACATGGTCCTTCTTGAGTTTG, E0240_mut_M_F (80)
    • ACCATGTGGTCTCTCTTTTCGTTGGGATCTTTCGAAAGGGCAGATTG, E0240_mut_M_R (79.3)

I2057

• Re-transformed both versions with ccdB into DB3.1

I2055 and I2056

• Letting biobrick PCR run overnight.

T9002

• Re-tried, transformed ligation of F2620.

Misc

• Made chemically competent DB3.1 cells.