E coli transformation

Addition of 10 μL H2O to appropriate well, wait for 5 min, transfer into sterile tube

Transformation of E. coli with pSEVA281 A-C2 (sfGFP), pSEVA271 C-A9 (mCherry) and piG0030:
Addition of 1 μL DNA to 75 μL competent cells (thawed on ice), put samples on ice for approximately 20 min
Heat shock: 90 s at 42 °C
Addition of SOC to samples, let cells recover for ca. 1h at 37 °C, 220 rpm
Plate 100 μL of bacteria suspension on LB-agar-plates containing the appropriate antibiotics (pSEVA281 A-C2, pSEVA271 C-A9: Kanamycin (50 μg/L), piG0030: Chloramphenicol (34 μg/L))
Let bacteria grow overnight at 37 °C

Sequencing results

Biobricks of the plasmids piG0001, piG0015 and piG002 show the correct sequences

E coli Transformation

Transformation of E. coli with pSEVA181, pSEVA271, pSEVA291, pSEVA561.
Addition of 1 μL DNA to 75 μL competent cells (thawed on ice), put samples on ice for approximately 20 min Heat shock: 90 s at 42 °C
Addition of SOC to samples, let cells recover for ca. 1h at 37 °C, 220 rpm
Plate 100 μL of bacteria suspension on LB-agar-plates containing the appropriate antibiotics (pSEVA181: Ampicillin (200 μg/L), pSEVA271 and pSEVA291: Kanamycin (50 μg/L), pSEVA561: Tetracycline (20 μg/L))
Let bacteria grow overnight at 37 °C

Antibiotic stock preparation

Preparation of antibiotics stock:
2 g Ampicillin were dissolved in 10 mL H2O and sterile filtered
0.5 g Kanamycin were dissolved in 10 mL H2O and sterile filtered
0.34 g Chloramphenicol were dissolved in 10 mL Ethanol