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

From OpenWetWare

Jump to: navigation, search
iGEM Project name 1 Main project page
Previous entry      Next entry

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 Track

Eric F.

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
25 <> 3 4 5 6 7 8 9 10
26 C 0.1052 0.1602 0.1652 0.1402 0.1432 0.1454 0.1769 0.0945
27 D 0.1736 0.1404 0.1528 0.0964 0.1629 0.0974 0.1596 0.1941
28 E 0.2193 0.1934 0.1758 0.1013 0.1759 0.1916 0.1981 0.1949
29 F 0.0977 0.1874 0.1776 0.1997 0.1656 0.1815 0.186 0.0967
  • 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
24 <> 1 2 3 4 5 6 7 8 9 10 11 12
25 A 0.2131 0.1653 0.1571 0.1857 0.1535 0.1532 0.1423 0.1573 0.1678 0.2214 0.1711 0.152
26 B 0.201 0.2033 0.1522 0.151 0.1429 0.1834 0.152 0.1574 0.1609 0.1633 0.1213 0.1499
27 C 0.198 0.1578 0.1561 0.1541 0.0927 0.1646 0.1497 0.1536 0.1507 0.1609 0.1763 0.1882
28 D 0.1929 0.1983 0.1703 0.1542 0.1465 0.1528 0.1506 0.1492 0.1593 0.1723 0.1558 0.1703
29 E 0.1601 0.178 0.1611 0.1407 0.1673 0.1611 0.1516 0.1492 0.1658 0.168 0.1668 0.1932
30 F 0.2193 0.1551 0.1721 0.1531 0.1731 0.1686 0.1608 0.1613 0.1696 0.1444 0.1573 0.1673
31 G 0.1875 0.165 0.1592 0.159 0.1534 0.1365 0.1681 0.1556 0.182 0.1536 0.0898 0.1627
32 H 0.1667 0.1863 0.1668 0.1943 0.1776 0.1687 0.1867 0.1869 0.1487 0.1749 0.1692 0.1764
  • 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.

Quorum Sensing Track

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


Personal tools