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1. Plate reader results very strange and somewhat sporadic. After a discussion with Barry, we decided to try to grow up the cells for 9 hours and see if we can get a smaller range of OD600 values. There is also the possibility that the plate reader is malfunctioning.

2. None of the cultures grew on the M9 minimal media after a weekend or on the 4-amino acid-supplemented media after a night. We need to rethink this.

3. Late last night, Reshma let me know that some expired gels are available which may help me set up the protein gels earlier.

Things to Do

1. Make LCs of J45995, J45996, R0040.E0840, and B0015 adding the antibiotics to the individual cultures and using the same LB media for all of them- DONE (11:07 to be removed at 8:07)

2. Make LCs of J45120, J45181, J45200, J45250, and J45800. J45120 and J45181 will be regrown tomorrow and extracted at various time points for protein gel analysis. J45800, induced and uninduced, will be done in parallel. J45200 and J45250 could be used for a time course later on in the week- DONE

3. Contact Tom about the next step for the media question- DONE

4. Contact Drew about meeting with him about abstract/general project- DONE

5. Ask Reshma if cells can be placed on ice before running on protein gel- DONE

6. Come up with title/description of project for Amgen- DONE

7. Clean up your area- DONE

8. Made 4 25-mL cultures with the following added for both J45700 and J45800- DONE (Dr. Knight agrees with this plan I came up with):

  • A: 24 mL minimal media, .5 mL 10% casamino acids, 1 mL 20% fructose ( 1 mL instead of .5 mL at Francois' suggestion since the fructose looked really old), + antibiotics
  • B: 24 mL minimal media, .5 mL 10% casamino acids, .25 mL 40% glycerol, + antibiotics
  • C: 24 mL minimal media supplemented with proline, serine, aspartic acid, and alanine, 1 mL 20% fructose, + antibiotics
  • D: 24 mL minimal media supplemented with proline, serine, aspartic acid, and alanine, .25 mL 40% glycerol, + antibiotics
  • Note: Proline, serine, aspartic acid, and alanine were added and completely dissolved by adding 1/10 the amount added to a liter (.142 g, .0655 g, .114 g, & .0392 g, respectively) to 100 mL minimal media and filter sterilizing the mixture.

OD600s at 8:07

J45995- 1.58

J45996- 0.88

R0040.E0840- 1.70

B0015- 0.51

  • Still very different- discuss with Barry tomorrow
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