Characterization:

We characterized 2 parts this year: BBa_K2890001 and BBa_K1334002.

I. Data of BBa_K2890001:

(1)The toxic effect of formaldehyde on the cells growth:

We added the reporter gene GFP to the downstream of the promoter, then detected the tolerance of the cells and the sensitivity of the promoter to formaldehyde using BL21 as chassis.

Formaldehyde is toxic to cells. It can inhibit the cells growth if it is added to the culture medium too early. In order to know at which OD₆₀₀ the formaldehyde should be added into the culture medium, we used 0.5mM formaldehyde to treat cells at different original OD₆₀₀, then we collected different group bacteria at certain time to measure cells’ DO₆₀₀ absorbance (Figure 1). The result showed that the toxic effect of formaldehyde on the cells growth was decreased with the cells original OD₆₀₀ increased.

(2)The sensitivity of promoter to formaldehyde:

We cultured E.coli BL21 transformed with the plasmid (PfrmR-GFP) and added different concentration formaldehyde to different groups when the OD₆₀₀ = 1, then we collected bacteria at certain time to measure the fluorescence and the absorbance. From the Figure 2, we found that 0.8 mM formaldehyde is the best concentration for inducing GFP expression.

II. Data of BBa_K1334002:

(1)The effect of formaldehyde to the cells growth:

Formaldehyde is toxic to the cells as we know, inhibiting cells growth if it is added early to the culture medium. In order to know at which OD₆₀₀ the formaldehyde should be added into the culture medium, we added the 0.5mM formaldehyde to cells at different original OD₆₀₀, then we collected different group bacteria at certain time to measure cells’ DO₆₀₀ absorbance (Figure 4). The result showed that the toxic effect of formaldehyde on the cells growth was decreased with the cells original OD₆₀₀ increased.

(2)The sensitivity of the promoter to formaldehyde:

Different concentration of formaldehyde has different ability to induce expression. In order to find the best concentration of formaldehyde to induce this promoter, we measured the sensitivity of the promoter to formaldehyde using BL21 as chassis. We cultured E.coli BL21 transformed with the plasmid （P-formaldehyde plus GFP）and added different concentration formaldehyde to different groups at OD₆₀₀ = 1. Then we collected different group bacteria at certain time to measure the fluorescence and the absorbance (Figure 5). The result showed that 0.8 mM formaldehyde is the best concentration for inducing GFP expression.