Team:UCLA/Notebook/Spider Silk Genetics/9 September 2015

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9/9/2015

Sequencing Results for M1-15(T7)

• Both sequences are incorrect.
• Both have the prefix, but the suffix only has PstI.
• Decided to BLAST the insert.
  • The results have almost exact similarity to E. coli K-12 genome.
• Due to lack of time, we will not finish cloning the M1-15 construct.

Digestion Gel Image

• Digest every unique ICA construct that we have made so far using EcoRI and PstI for a final gel image.
• Digest 1 ug of each (1C3) construct plasmid in a 50 uL reaction using 1 uL of each enzyme in NEBuffer 2.1
• Visualize on a 1% TAE gel using 2 uL of NEB 1 kb ladder.

Fig. 1 E,P digestion of MaSp constructs made using ICA this summer. The expected sizes are indicated on the right hand side. The expected size for the backbone is 2070 bp.

• The result for M1-9 is slightly anomalous,The band at ~3 kb may be due to incomplete digestion. Will repeat tomorrow.
• Also, there is a smear for M2-9 digestion. Will also repeat tomorrow.