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From 2011.igem.org

Notebook

Week 1

PCR primers designed according to Biobrick specifications and ordered for rsaA C-terminal and WT esp. Because of the discrepancy in GC content between Staphylococcus epidermidis and Caulobacter, we also optimized the esp sequence for Caulobacter for later synthesis with a higher resulting GC content.

Prepared competent cells of E. coli Top10. (Protocol)

Streaked plates of Caulobacter, Staphylococcus aureus, and S. epidermidis.

Template DNA for PCR prepared from colonies using [http://www.bioventures.com/products.php?cid=10005| GeneReleaser]. (Protocol)

Week 2

Performed PCR on esp and rsaA genes and run on gel. Results showed successful amplification.

Performed a transformation experiment to test the efficiency of our competent cells make in Week 1. Results were successful with about a 0.7 x 10^6 CFU/μg.

Cleaned up PCR product and digested esp, rsaA, and spB1C3 plasmid. Ligated esp, rsaA, and esp and rsaA to spB1C3 and transformed into E. coli Top10.

Inoculated liquid cultures with colonies from transformations.