IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/08/18

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Biofilm Track

Eric F. + Melody

Misc.

  • Streaked TSA plates of 8325-4 and RN4220, previous ones did not have enough individual colonies to be useful
  • Autoclaved pipette tips
  • Prepared 0.1% + 0.01% Crystal Violet (by accident) - was of no use

Biofilm Growth Protocol Day 5+ - 100811a + 100811b

  • Resolubilized 100811a + b with 95% ehtanol
  • Corresponding plates were labelled 100811a-b and 100811b-b
  • Placed in biosafety cabinet, waiting for plate reader to be free
  • Took readings

Biofilm Growth Protocol Day 3 - 100816M + 100816E

  • Followed day 3 protocol

Biofilm Growth Protocol Day 4 - 100814M + 100814E

  • Stained with (what we later realized to be) 0.01% Crystal Violet
  • Took readings, did NOT resolubilize today

Readings

  • Took readings on 100811a-b, 100811b-b, 100814M and 100814E
  • All readings were garbage will place below soon
  • Decided to ethanol lid and bottom of plates before re-doing measurements
100811a-b Readings
25 <> 3 4 5 6 7 8 9 10
26 C 0.0792 0.0845 0.0855 0.0807 0.0814 0.0777 0.0828 0.0891
27 D 0.0773 0.0789 0.0823 0.0829 0.0805 0.08 0.081 0.0819
28 E 0.0803 0.0812 0.0813 0.0846 0.0788 0.084 0.0827 0.085
29 F 0.0778 0.0861 0.0796 0.0849 0.0834 0.0812 0.0841 0.0796
  • Red = control
100811b-b Readings
24 <> 1 2 3 4 5 6 7 8 9 10 11 12
25 A 0.0895 0.081 0.086 0.0854 0.074 0.075 0.0775 0.0763 0.077 0.0745 0.0874 0.0932
26 B 0.081 0.0871 0.0825 0.0859 0.0785 0.081 0.0866 0.0805 0.0793 0.0854 0.1023 0.0841
27 C 0.0823 0.0786 0.0827 0.0847 0.076 0.0803 0.0824 0.0791 0.0775 0.0841 0.0857 0.095
28 D 0.0818 0.0903 0.0783 0.0816 0.0821 0.0828 0.0812 0.0766 0.0812 0.0831 0.0787 0.0833
29 E 0.0901 0.0916 0.0799 0.0806 0.0816 0.0829 0.0822 0.0884 0.0866 0.0822 0.0824 0.0911
30 F 0.0823 0.0823 0.0839 0.0811 0.0821 0.0814 0.0818 0.0821 0.0839 0.0824 0.0849 0.0812
31 G 0.0781 0.0778 0.0804 0.0833 0.088 0.0794 0.0831 0.0905 0.0825 0.0869 0.083 0.0826
32 H 0.0987 0.0907 0.0902 0.0814 0.0814 0.0825 0.0844 0.0882 0.0792 0.0843 0.0821 0.0938
  • Red = control
100814M Readings
25 <> 3 4 5 6 7 8 9 10
26 C 0.0888 0.2545 0.1799 0.2674 0.1814 0.1559 0.1363 0.1061
27 D 0.2585 0.3001 0.299 0.1342 0.0908 0.1863 0.1828 0.1336
28 E 0.225 0.2138 0.2287 0.1308 0.0944 0.1467 0.1435 0.1338
29 F 0.0957 0.1552 0.1509 0.159 0.1202 0.1103 0.1079 0.0906
  • Red = control
100814E Readings


25 <> 3 4 5 6 7 8 9 10
26 C 0.0938 0.1238 0.1038 0.0931 0.1034 0.1422 0.109 0.0994
27 D 0.1042 0.164 0.2106 0.1101 0.1065 0.101 0.1104 0.124
28 E 0.1003 0.1712 0.1296 0.1099 0.1092 0.1079 0.1577 0.1189
29 F 0.1638 0.1444 0.148 0.0999 0.1926 0.1471 0.1397 0.1453
  • Red = control

dspB

Mini-prep

  • 3T1,3T2,4T1,4T2
  • Protocol: Rafael's alkaline lysis

Nanodrop [ng/uL]:

  • 3T1: 2541.7
  • 3T2: 3053.8
  • 4T1: 3357.5
  • 4T2: 3587.8
SampleVolume of sample (uL)Volume of H2O
3T11.5748.426
3T21.3098.691
4T11.1918.809
4T21.1158.885
  • Primers: VF2 and VR
    • 5uM each; 6uL in total (require 1uL of 5uM primers)

=QS Track

  • Miniprepped agrCA (following Raf's alkaline lysis protocol) from distribution (after following cPCR program using UBC iGEM protocols)
  • No colonies for agrCA (natural form) showed up yesterday (both agrCA by itself and agrCA with terminator)
  • Therefore, PCR'd more agrCA.
  • Used same program, amount of reagent, and protocol as August 11 agr CA PCR.
  • Ran on gel with 1.25% agarose, 0.5X TBE, 40 minutes, 100 V
  • Bands showed up where expected (~2.1 kb)
  • Used Qiagen PCR purification kit to purify agr CA product.
  • Checked DNA quality with nanodrop. A260/280=1.90. A260/230=1.70. [DNA]=68.7 ng/μL.


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