Team:Pasteur Paris/Notebook

Labwork

July
M	T	W	T	F	S	S
					1	2
3	4	5 Operators: Deshmukh, Ersin, Gabriel, Alexis Transformation of bacteria DH5α with pet43.1a.C162, for the positive control we will use the plasmid pUC19 + a negative control according to the transformation protocol except for volumes. Observations 16h later: • The culture for the positive test revealed mean bacterial cultures (more than 50) this test confirms our DH5α cells are competent • The culture for the negative test did not reveal any bacterial culture, which proved that the cells without any plasmids couldn’t survive in an antibiotic based media • The cell cultures containing pET43.1a.C162 do not present any colonies; there must be a problem with the pET43.1a.C162 solution, because the DH5α cells are truly competent • By spectroscopy UV-visible using the equipment UV5 Nano Mettler Toledo, at 260 nm with a 1µL sample, we measured a plasmid concentration of 0, the solution used did not contain plasmids, which explains the death of our cultures For the detailed protocols followed, refer to the PDF documents below	6 Operators: Deshmukh, Ersin, Gabriel, Alexis Transformation of plasmids pET43.1a.C162 in DH5α (2nd test) according to the transformation protocol except for volumes For the detailed protocols followed, refer to the PDF documents below Operators: Deshmukh, Ersin, Gabriel, Alexis Petri Dish preparation according to the preparation protocol except for volumes. 33 Dish total Then the Petri Dish are stored at 4°C or placed at the incubator at 37°C for immediate use. For the detailed protocols followed, refer to the PDF documents below	7 Culture in liquid media of DH5α containing pET43.1a.C162 according to the protocol of liquid culture except for the volumes. The pellet is stored at -20°C named: DH5α pET43.1a.C162. For the detailed protocols followed, refer to the PDF documents below	8	9
10	11 Operators: Deshmukh, Karima, Pierre-Louis, Juliette, Paul, Nathan, Diane Culture in liquid media of DH5α containing pET43.1a.C162 according to the protocol of liquid culture except for the volumes. Note: the Erlenmeyer number 5 was given 2 doses of antibiotics so we have 50µL of antibiotics for 25mL of LB broth media. For the detailed protocols followed, refer to the PDF documents below	12 Operators: Juliette, Karima, Paul Midiprep of pET43.1a.C162 previous liquid culture with the Qiafilter Midi kit from Qiagen. Four samples with an average concentration of 60 ng/µL where store at -20°C. Operators: Ersin, Gabriel, Alexis Digestion of the plasmid pET43.1a.C162 according to the digestion protocol except for the volumes with BamHI and XbaI. Launch of an electrophoresis in order to extract the empty plasmid pET43.1a. The cutted gel were stored at 4°C For the detailed protocols followed, refer to the PDF documents below	13 Operators: Ersin, Gabriel, Alexis Gel extraction of the plasmid pET43.1a with the Qiagen gel extraction kit. The sample is stored at -20°C, with an average concentration of 7,418 ng/µL. For the detailed protocols followed, refer to the PDF documents below Operators: Gabriel, Paul Electrophoresis of pET43.1a.C162 midiprep from the 12/10/2017. Check of the plasmid purity. For the detailed protocols followed, refer to the PDF documents below	14	15	16
17 Operators: Paul, Ersin, Diane, Alexis Digestion of plasmids pET43.1a.C162 Launch of an electrophoresis in order to extract the empty plasmid pET43.1a. The cutted gel were stored at 4°C For the detailed protocols followed, refer to the PDF documents below	18 Operators: Paul, Ersin, Gabriel, Diane Gel extraction of the plasmid pET43.1a with the Qiagen gel extraction kit. Four samples with an average concentration of 61 ng/µL where store at -20°C. For the detailed protocols followed, refer to the PDF documents below	19 Operators: Ersin, Alexis Solubilisation of Eurofins Plasmid E2 gene in TE 0.1X : pEXA128-E2. Operators: Diane, Ersin, Gabriel, Alexis, Karima Protocol for Bacteria DH5α Transformation with pEXA128-E2 For the detailed protocols followed, refer to the PDF documents below Operators: Gabriel, Ersin, Alexis, Diane, Karima Dephosphorylation of plasmid or DNA: pET43.1a (X-B) For the detailed protocols followed, refer to the PDF documents below	20	21	22	23 Operator: Deshmukh Liquid Culture for miniprep on transformed Bacteria DH5α pEXA128-E2. For the detailed protocols followed, refer to the PDF documents below
24 Operators: Alexis, Juliette Petri Dish preparation according to the preparation protocol except for volumes. 20 Dish total Then the Petri Dish are stored at 4°C or placed at the incubator at 37°C for immediate use. Operator: Gabriel Solubilisation of the lyophilized Eurofins plasmid pEXA128 E1_2, for its future transformation Operators: Alexis, Juliette, Karima Transformation of DH5α with pSB1C3 Operators: Diane, Nathan Transformation of DH5α with pEXA128 E1_2. For the detailed protocols followed, refer to the PDF documents below Operators: Ersin, Gaétan Miniprep of previous liquid culture with the QIAprep Spin Miniprep kit from Qiagen. Four samples with an average concentration of 280 ng/µL. Operators: Ersin, Gabriel Digestion of the plasmid pEXA128 E2 according to the digestion protocol except for the volumes with BamHI and XbaI to retrieve our gene E2. Operator: Gabriel Electrophoresis gel preparation Electrophoresis run For the detailed protocols followed, refer to the PDF documents below	25 Operators: Juliette Alexis Liquid Culture for Miniprep on transformed Bacteria DH5α pEXA128-E1_2 Operators: Karima, Diane Gel Extraction of the gene E2 with QIAquick Gel Extraction Kit: Operators: Karima, Juliette, Alexis Transformation of DH5α by pSB1C3 Operators: Karima, Diane Ligation between dephosphorylated pET43.1a and E2 For the detailed protocols followed, refer to the PDF documents below	26 Operators: Karima, Diane Miniprep on transformed Bacteria DH5α pEXA128-E1_2 • Four samples with an average concentration of 320 ng/µL. Plasmid Digestion and Gel extraction of pEXA128-E1_2 Ligation of plasmid with DNA insert pEXA128-E1_2 For the detailed protocols followed, refer to the PDF documents below Operators: Ersin Gabriel DNA Digestion pEXA128-E2 For the detailed protocols followed, refer to the PDF documents below	27	28	29	30
31 Operators: Alexis, Ersin Solubilisation of Eurofins Plasmid E3 gene: pEXA258-E3 Solubilisation of Eurofins Plasmid E1_1 gene: pEXA258-E1_1 Operators: Karima, Diane DNA Digestion of pEX-E2 and pEX-E1_2 with XbaI and BamHI For the detailed protocols followed, refer to the PDF documents below Operators: Alexis, Ersin Transformation of DH5α with pEXA258-E1_1: Protocol for Bacteria Transformation of DH5α with pEXA258-E3 For the detailed protocols followed, refer to the PDF documents below

August
M	T	W	T	F	S	S
	1 Operators: Diane, Ersin Digestion of plasmid pET43.1a.C162 Electrophoresis Gel Extraction with QIAquick Gel Extraction Kit For the detailed protocols followed, refer to the PDF documents below Operators: Diane, Alexis Liquid Culture for Miniprep on transformed Bacteria DH5α pEXA258-E1_1 and pEXA258-E3 For the detailed protocols followed, refer to the PDF documents below	2 Operators: Ersin, Diane Miniprep on transformed Bacteria DH5α pEXA258-E1_1 and pEXA258-E3 Ligation of plasmid pET43.1a with DNA insert E2 col 2 and E1_2 col 2 Gel Extraction of pET43.1a with QIAquick Gel Extraction Kit Digestion of DNA pEXA258-E1_1 and pEXA258-E3 with XbaI and BamHI For the detailed protocols followed, refer to the PDF documents below Operators: Ersin, Diane Transformation of DH5α with pET43.1a-E1_2 Transformation of DH5α with pET43.1a-E2 or pET43.1a Dephosphorylation of plasmid pET43.1a with XbaI and BamHI For the detailed protocols followed, refer to the PDF documents below	3 Operators: Ersin, Diane Gel Extraction of E1_1 and E3 with QIAquick Gel Extraction Kit Ligation of plasmid pET43.1a with DNA inserts E3 col 1 and E1-1 col 3 For the detailed protocols followed, refer to the PDF documents below Operators: Diane, Alexis Liquid Culture for Miniprep on transformed Bacteria DH5α pET43.1a-E2 Operators: Ersin, Diane Protocol for Bacteria Transformation of DH5α with pET43.1a-E1-1 and pET43.1a-E3 For the detailed protocols followed, refer to the PDF documents below	4 Operators: Alexis, Paul Miniprep on transformed Bacteria DH5α pET43.1a-E2 Operators: Alexis, Paul Digestion of pET43.1a-E2 with XbaI and BamHI For the detailed protocols followed, refer to the PDF documents below	5	6
7 Operators: Ersin, Diane Digestion of pET43.1a-E1_1 and pET43.1a-E1_2 with XbaI and BamHI DNA pET43.1a-E3 Digestion X-B For the detailed protocols followed, refer to the PDF documents below Operators: Ersin, Diane Transformation BL21DE3 pET43.1a-E1_1 Transformation BL21DE3 pET43.1a-E1_2 Transformation BL21DE3 pET43.1a-E2 Transformation BL21DE3 pET43.1a-E3 For the detailed protocols followed, refer to the PDF documents below	8 Operators: Gabriel, Diane Preparation of solutions of plasmids pET43.1a-E1_1, pET43.1a-E1_2, pET43.1a-E2 and pET43.1a-E3, for sequencing by Eurofins Operators: Diane, Azéline Liquid Culture of transformed Bacteria BL21DE3 pET43.1a-E1-2 and pET43.1a-E2 For the detailed protocols followed, refer to the PDF documents below Operators: Ersin, Diane Transformation BL21DE3 pET43.1a-E1_1 Transformation BL21DE3 pET43.1a-E1_2 Transformation BL21DE3 pET43.1a-E2 Transformation BL21DE3 pET43.1a-E3 For the detailed protocols followed, refer to the PDF documents below	9 Operators: Gaétan, Gabriel Expression culture and kinetic BL21 pET43.1a-E1_2 and pET43.1a-E2 For the detailed protocols followed, refer to the PDF documents below Operators: Diane, Azéline Transformation BL21DE3 pET43.1a-E1_1, pET43.1a-E1_2, pET43.1a-E2, pET43.1a-E3 For the detailed protocols followed, refer to the PDF documents below	10 Operators: Gaétan, Gabriel Liquid culture of BL21DE3 pET43.1a-E1-2 and pET43.1a-E2 For the detailed protocols followed, refer to the PDF documents below Operators: Gabriel, Karima Bacteria lysis and SDS Page gel migration of Bl21DE3 pET43.1a-E2 and pET43.1a-E1_2 For the detailed protocols followed, refer to the PDF documents below	11 Operators: Gaétan, Diane Liquid culture of BL21De3 pET43.1a-E1-1 and pET43.1a-E3 For the detailed protocols followed, refer to the PDF documents below	12	13
14	15	16	17	18 Operators: Gabriel, Diane, Alexis Bacteria lysis and SDS Page gel migration For the detailed protocols followed, refer to the PDF documents below	19	20
21 Operators: Diane, Nathan InterLab Transformation For the detailed protocols followed, refer to the PDF documents below	22 Operators : Ersin, Diane Transformation BL21DE3 pET-E3 Transformation DH5α and BL21DE3 pET-E1_1 For the detailed protocols followed, refer to the PDF documents below	23 Operators: Alexis Solubilisation of Eurofins Plasmid E4 gene: pEXA258-E4 Transformation DH5α pEX-E4 For the detailed protocols followed, refer to the PDF documents below	24	25	26	27
28 Operators : Nathan, Diane Transformation of DH5α with pSB1C3 of E1_1, E1_2, E2, E3 and E4 For the detailed protocols followed, refer to the PDF documents below	29 Operators : Paul, Diane Transformation of DH5α with pSB1C3 of E1_1, E1_2, E2, E3 and E4 For the detailed protocols followed, refer to the PDF documents below	30	31

September
M	T	W	T	F	S	S
				1	2	3
4	5	6 Operators: Maud, Paul, Diane DNA Digestion and Gel extraction of pET32a For the detailed protocols followed, refer to the PDF documents below Operators: Maud, Diane Dephosphorylation of pET32a For the detailed protocols followed, refer to the PDF documents below	7 Operators : Maud, Diane Transformation of DH5α with pET32a of E1_1, E1_2, E2, E3 and E4 For the detailed protocols followed, refer to the PDF documents below	8	9	10
11	12	13	14	15	16	17
18	19	20	21	22	23	24
25	26	27	28 Operators: Ersin, Gaétan, Deshmukh Miniprep on pEX-S1 transformed Bacteria For the detailed protocols followed, refer to the PDF documents below Operators: Ersin, Gaétan, Deshmukh Transformation of DH5α with pSB1C3-E1_1; E1_2 and E2 Transformation of DH5α pET32a-E1_1; E1_2 and E2 For the detailed protocols followed, refer to the PDF documents below	29	30

October
M	T	W	T	F	S	S
						1
2	3	4	5 Operators: Juliette and Ersin Digestion of pET32a with E1_1; E1_2 and E2	6	7 Operators: Deshmukh and Juliette Liquid culture of transformed bacteria and induction of protein expression For the detailed protocols followed, refer to the PDF documents below Operators: Juliette Digestion if pSB1C3 by EcoRI and PstI DNA Electrophoresis of pSB1C3 digested by EcoRI-PsTI For the detailed protocols followed, refer to the PDF documents below	8 Operators: Paul and Azéline DNA Electrophoresis of S1 digested by EcoRI-PstI or XbaI-BamHI For the detailed protocols followed, refer to the PDF documents below
9	10 Operators: Juliette and Alexis Gel Extraction with QIAquick Gel Extraction Kit of pSB1C3, S1 E-P and S1 X-B Ligation of pSB1C3 with our parts E1_1, E1_2, E2, E3, E4, S1 DH5α Transformation with previous plasmid For the detailed protocols followed, refer to the PDF documents below	11 Operators: Juliette Ligation of pET32a with S1 Transformation of DH5α with pET32a-S1 For the detailed protocols followed, refer to the PDF documents below	12	13	14	15
16	17	18	19 Operators: Gabriel Digestion of Biobricks made the 10-10-2017 For the detailed protocols followed, refer to the PDF documents below	20	21	22
23	24 Operators: Paul Liquid culture of DH5α with all of our biobricks for -80°C storage For the detailed protocols followed, refer to the PDF documents below	25	26 Operators : Juliette, Gabriel, Ersin Reaction kinetics of the degradation of Anthracene Please check our Demonstrate page for further information For the detailed protocols followed, refer to the PDF documents below	27 Operators : Juliette, Gabriel, Ersin Reaction kinetics of the degradation of Anthracene Please check our Demonstrate page for further information For the detailed protocols followed, refer to the PDF documents below	28	29
30	31

For blank protocols, please check our protocol page here

For the link between our Biobricks and the abreviations used in the protocols, please check our parts page here

Week 1 protocols

Week 2 protocols

Week 3 protocols