Week Eight - 5 August to 11 August


Lajja and Mike performed PCR on Bacillus and Citrobactor. They then ran a gel of the samples.


Sarah, Tae and Lajja made a stock solution of Chloramphenicol since tetracycline does not seem to be working as hoped. Chloramphenicol plates were also made.

Yuan and Grant PCR purified the Bacillus and Citrobactor following the usual protocol
The lab PCR’d out the amp backbone using the linearized plasmid backbone protocol
Pbad and citrobacter were digested according to the usual protocol

6.29 uL of 79.5 ng/uL of Pbad

36.2 uL water

14.04 uL of 35.6 ng/uL of Citrobacter

28.5 uL water

Sarah, Tae and Lajja prepared and ran a gel for analysis. The gel contained the digested Pbad and citrobacter parts. After analysis, the Pbad and citrobacter parts did not appear on the gel as expected so a redo of the gel should be done tomorrow.


Brian transformed pSB1C3 RFP plate 1 well 3A, pSB1K3 RFP plate 1 well 5A

Mike and Grant performed a digestion on the pBad promoters, lysis parts, and K backbone according to the usual protocol.

Yuan digested the amp backbone with dpn1 and then PCR purified it following the normal protocol
Yuan performed a ligation of the citrobacter phytase part into the linearized amp plasmid backbone


Brian transformed the following parts:
ligated Citrobacter phytase that is in the pSB1A3 backbone
Strongest RBS (kit plate 1 well 2M)
Strong RBS: (kit plate 1 well 1H)
Strongest con. pro.: (plate 1 well 18A)
Strong con. pro. : (plate 1 well 18K)
Medium con. pro. : (plate 1 well 20C) -Not Transformed, removed from plate and frozen


Pbad WT 77.5 ng/uL, 12.9 uL DNA, 39.6 uL water

Pbad nWT 92.6 ng/uL, 10.9 uL DNA, 31.7 uL water

Cut both with E and then (E and P) for a diagnostic

Friday August 10th:

Strongest RBS 6.87 uL, 35.63ul water

Strong RBS: 4.88 uL, 37.62ul water

Strongest con. pro.: 6.24 uL, 36.66ul water

Strong con. pro. : 1.95 uL, 40.55ul water

Chloroamp bb: 2.50 uL, 40ul water

Pbad nbWT: 5.43 uL, 37.07ul water


Yuan miniprepped the promoters for Grand and Sarah
Grant and Michael PCR’d and PCR Purified the kanamycin and chloramphenicol plasmid backbones
Grant and Michael ran a gel on the PCR’d plasmid backbones

Yuann overnighted the citrobacter phytase(both a red and a white colony that was grown on amp)


Miniprepped the the citrobacter phytase(both the red and white colony)

Yuan eluted with too much solution so she redid the miniprep of the citrobacter