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August/7 August 2009
From 2009.igem.org
Today we carried out the following procedures (in rough chronological order):
1. Checked the cell cultures transformed and plated out yesterday (8/6) for colony formation and made an approximate count of the number of colonies.
(plate number)-(location on plate) (no. of colonies)
F16010 (2-24G) none M0100 (2-15F) none I742158 (1-19B) >50 B0034 (1-2M) >50 I0462 (1-8O) ~10 K118002 (2-16F) ~10 K118003 (2-16H) ~10 B0015 (1-23L) >50 I1466 (1-23J) ~10 C0077 (1-14F) ~10 C0076 (1-14D) ~10 K081020 (2-12H) none S03878 (2-16M) ~10 C1079 (2-8M) ~10
2. Conducted a 'miniprep' to harvest and check concentration of the EpsE (molecular clutch) plasmids from 3 test tube cultures incubated since yesterday (8/6). Result of Nanodrop concentration check:
(test tube no.) [260/280] [260/230] (concentration)
1 2.14 1.65 37.6 ng/ul
2 2.07 1.95 62.9 ng/ul
3 1.95 1.33 60.6 ng/ul
3. Prepared kanamycin antibiotic solution from kanamycin sulfate and MilliQ water. 100 mg kanamycin sulfate + 10 ml MilliQ water -> 10 ml of 10 mg/ml kanamycin solution.
4. Transformed compe[tent cells with the following parts to amplify them:
(plate number)-(location on plate) (antibiotic resistance)
F16010 (2-24G) A K081020 (2-12H) K K118013 (2-18F) A R0071 (1-12C) A I14017 (1-18L) K R0062 (1-6O) A R0079 (1-12A) A R0077 (1-10K) A
Note: 2-24G and 2-12H were transformed yesterday but produced no colonies. 2-18F was marked for transformation yesterday but 2-15F was mistakenly transformed instead.
5. Treated a sample of EpsE-containing plasmids with EcoRI and SpeI restriction endonucleases to isolate and check length of amplified EpsE part. The plasmid-buffer-endonuclease mixture was incubated and left to react overnight (Incubation at 37 degrees Celsius began at 16:00).
