Part:BBa_K1468000:Experience

Applications of BBa_K1468000

For testing the behaviour of BBa_K1468000, we used six different E. coli strains: DH5α, XL1Blue, JM109, DH10B, HB101, BL21(DE3); which were grown in standard conditions: 37°C, 200 rpm (revolutions per minute) and LB with a standard composition (1% NaCl, 1% peptone and 0.5 % yeast extract).

To carry out the assay, we did as follow:

- Transformed strains -with Bba_K1468000 and with the empty plasmid, no biobrick- were plated on LB with the appropriate antibiotic (chloramphenicol).

- Overnight incubation at 37°C.

- Bacterial colonies were transfered to 1 mL of LB supplemented with chloramphenicol.

- A 20 minutes at 37°C incubation was carried out.

- These precultures (100 µL) were used to inoculate a tube containing 2 mL of LBC. Four biological replicates were perfomed.

- The cultures were incubated (standard conditions) until OD595 is between 0.2-0.4 (1 hour aprox.).

- The OD at 595 nm was measured.

- GFP fluorescence (λex= 493nm; λem=505nm) was measured.

With all this data the average, fluorescence /cell density, and standard deviation were calculated taking into account the four biological replicates.

The results obtanied were:

Figure 1. Behavior of Bba_K1468000 in six different E. coli strains. Fluorescence intensity of Bba_K1468000 was corrected by the fluorescence of cells containing an empty plasmid.

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