Notebook
Notebook - important experiments
26.07.2017
- transformation of kerUS (BBa_K1498000)
- preparation of antibiotic stock solutions: chloramphenicol (35 mg/mL) and ampicillin (100 mg/mL)
27.07.2017
- growing of a single colony (kerUS (BBa_K1498000)) in 5 mL LB media chloramphenicol (35 µg/mL] at 37 °C
- single colonies are plated on agar plates and incubated at 37 °C over night
28.07.2017
- Preparation of: miniprep (Jena Biosciences Kit)and
glycerol stock (kerUS (BBa_K1498000))
03.08.2017
- transformation of BBa_K1149002 and BBa_K1149003
04.08.17
- single colonies (BBa_K1149002 and BBa_K1149003) are plated on agar plates and incubated at 37 °C over night
09.08.17
- growing of a single colony (BBa_K1149002 and BBa_K1149003) in 5 mL LB media + chloramphenicol (35 µg/mL] at 37 °C
10.08.17
- Preparation of miniprep (Jena Biosciences Kit) and
glycerol stock (storage: -80 °C) (BBa_K1149002 and BBa_K1149003)
- SOC media preparation
- transformation pet19-LipB
21.08.2017
- preparation: preliminary test of esterase assay with EstCS2: Bba_K1149002 - preparation of o/n cultures (37°C)
22.08.2017
- preparation: preliminary test of esterase assay with EstCS2: Bba_K1149002 - glycerol stocks and induction with arabinose
23.08.2017
- preliminary test of esterase assay with EstCS2: Bba_K1149002 (link results)
28.08.2017
- preparation of electro-competent cells
31.08.2017
- transformation/electroporation with iGEM competent cells test kit DNA (o/n incubation, 37°C - transformation failed)
01.09.2017
- transformation/electroporation with pUC19 plasmid (o/n incubation, 37°C - transformation failed)
06.09.2017-08.09.17
- enzyme activity assay (cell pellet and supernatant) with BBa_K1149002 and pet19-LipB
12.09.2017
- preparation of chemo-competent cells
13.09.2017
- transformation with iGEM competent cells test kit DNA and pUC19 plasmid (o/n incubation, 37°C) - transformation successful
14.09.2017
- calculation - efficiency of the chemo-competent cells/pUC19:
efficiency E =(56cfu/0.001ng)*1000ng/(µL)=5.6*((10)^7 )cfu/µL
18.09.2017
- preparation InterLab study - transformation of 8 parts into DH5alph E. coli cells
19.09.17 – 21.09.17
- InterLab study LUDOX measurement
- enzyme activity assay with BBa_K1149002 of the supernatant - measurement in biological triplicates
- InterLab study Fluorescein measurement
22.09.2017
- InterLab study sample measurement (link results)
27.09.17 - 29.09.17
- enzyme activity assay with pet19-LipB of the supernatant - measurement in biological triplicates
06.10.2017
- Collaboration iGEM team Heidelberg: preparation mutagenesis plasmid activity assay - preparation of media, antibiotic-stocks and sugar-stocks + agar plates with different antibiotics
10.10.2017
- Collaboration iGEM team Heidelberg: preparation of different cultures + induction of the cells with arabinose (o/n incubation, 37°C)
11.10.2017
- Collaboration iGEM team Heidelberg: spread the o/n cultures on prepared agar plates (o/n incubation, 37°C)
16.10.2017
- preparation semi-quantitative keratinase-assay: spread kerUS, KerA and KerP on chloramphenicol/kanamycin agar plates (o/n incubation, 37°C)
- Preparation skim-milk-plate assay: preparation of skim-milk-agar-plates with chloramphenicol/kanamycin
18.10.2017
- M9 media preparation
- preparation semi-quantitative keratinase-assay: preparation of o/n cultures (37°C)
- Preparation skim-milk-plate assay: preparation of o/n cultures (37°C)
19.10.2017
- PCR - amplification of LipB - cloning of LipB in the iGEM standard plasmid (BBa_K1149002 is used)
- PCR - amplification of BBa_K1149002 (without EstCS2)
- semi-quantitative keratinase-assay: add 0.005 g of dried hair (65 °C, 1h) to o/n cultures (KerUS and KerA are induced with 1mM IPTG before) - 5 days incubation, 37°C (link results)
- skim-milk-plate assay kerP - spread on prepared skim-milk agar plates (supernatant and cells) - 4 days incubation, room temperature (link results)
20.10.2017
- SDS page, Gibson Assembly, Transformation (Zymos) - cloning of LipB in the iGEM standard plasmid (BBa_K1149002 is used)
- skim-milk-plate assay kerUS and KerA - spread on prepared skim-milk agar plates (supernatant and cells) - 4 days incubation, room temperature (link results)
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