Team:BIT/Interlab Study
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Summary
The objective of this year’s iGEM Interlab study is to quantify expression of five different reporter constructs. These constructs have GFP under the control of different promoter and ribosome binding sequences (RBS). Our Interlab work was carried out with the DH5-alpha as an expression host, and the Thermo Scientific Varioskan Flash spectral scanning as the equipment. We transfer the five reporter constructs into E.coli cells separately. And we measure the fluorescence data and optical density data with spectral scanning machine every 1 hour during a 6 hours cultivation.
Methods
We use endonuclease (EcoRI and PstI, XbaI and PstI) and T4 DNA ligase (NEB) to construct them. Finally, the ligation product was introduced into the pSB1C3. All constructs used were transformed into DH5α cells.
The cultivation of our bacteria was performed in tubes filled with 7 mL LB liquid medium. The cultures were kept at 37 °C and 110 rpm in shaker.
Antibiotics were added to each media (chloramphenicol 25mg/mL for Positive control、Negative control、Device 1：J23101+I13504、Device 2:J23106+I13504、Device 3:J23117+I13504).
We used100mL Luria Broth Liquid media(102μg/mL chloramphenicol) in 250 mL conical flask to grow our cells(37℃，110rmp). Every 1h, we took 100μL in 96-well plate (round wells, flat bottom) to test the OD600 and fluorescence intensity.
Measurement of fluorescence and OD were performed using the Thermo Scientific Varioskan Flash with the 'Thermo Scientific SkanIt' software and with the 96-Well Multiwell Plates (Corning).
Photometry
Contact Us
Beijing Institute of Technology,
No. 5 South Zhong Guan Cun Street,
Haidian Beijing 100081, P. R. China
Twitter : @igem_BIT
Sina Weibo : @igem_BIT
Website : http://www.bit.edu.cn
