IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/08/16

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Misc
Jason
 * 48-hr staph competent cell transformant didn't show growth
 * Redo using staph shuttle vector

Eric F.

 * Made 800mL LB Broth and autoclaved - probably still in the autoclave
 * Low on glassware - waiting on auto-autoclave
 * Low on some supplies, put on order sheet

Biofilm Growth Protocol Day 3

 * Removed from incubator at 1200 (25 hrs of incubation)
 * Control in F7 showed growth, surrounding control wells did not
 * Specks visible in D8 and D10
 * Will continue experiment ignoring results from:
 * D8
 * D10
 * E8 - adjacent to contaminated control
 * F8 - adjacent to contaminated control
 * Placed in bio safety cabinet to dry overnight @ 1440

Biofilm Growth Protocol Day 1

 * 3 - 3mL plastic test tubes
 * 1 control
 * 8325-4
 * RN4220
 * Into incubator w/o shaking @ 1630

Biofilm (100811) Growth Protocol Day 5+

 * Took OD readings of Test 1 plate (100811a - different glucose %s) and Test 2 plate (100811b - all locations)

Test 1 - 100811a - OD550 Readings

 * Where:
 * Red = Control at that row's Glucose level
 * Row C = 4% Glucose - RN4220
 * Row D = 2% Glucose - RN4220
 * Row E = 1% Glucose - RN4220
 * Row F = 0.5% Gluocse - RN4220

Test 2 - 100811b - OD550 Readings

 * Where:
 * Red = Control
 * All rows and columns are RN4220 w/ 4% glucose

Conclusions

 * 1% has the highest thickness biofilm
 * Edges show more biofilm growth
 * SUCCESS! Even though we already declared failure.

Phillip

 * Digested and ligated agr CA (natural RBS from PCR product; cut at EP)with PSB1C3 (cut at EP) following NEB biobrick protocols.
 * Digested and ligated agrCA natural RBS from PCR product; cut at ES) with B0014 (cut at XP) and PSB1Ce (cut at EP) following NEB biobrick protocls.
 * Started O/N culture of Staph aureus NCTC 8325 for AIP generation. TSB, 37 °C, 180 rpm.
 * Transformed ligation mixtures from above into E. coli competent DH5(alpha, following UBC iGEM procedures (e.g. incubate for 1 hour at 37°C, no shaking, in LB).
 * Plated onto LB chloramphenicol agar


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