Difference between revisions of "Part:BBa K3498029:Experience"

IGEM FCB-UANL

PvanCC promotor + Rsn-2 RBS + Rsn-2 + Terminator

This composite part is regulated by vanillic acid sensitive promoter PvanCC. This part contains an histidine tag used for purification. All assembled in pSB3K3 plasmid, a low number of copies plasmid

Introduction

Ranaspumin-2.

Ranaspumin-2, also known as Rsn-2 is a recombinantly produced protein in E. coli Top 10. Its foamability capacity is being tested for fire fighting foam production purposes

Results.

Protein expression in E. coli Top 10.

A characterization of the expression in E. coli was made. For this, different concentrations of inducer (vanillic acid) and different final growing times were tested. Concentrations were determined by imageJ pixel density analyzer program

Figure 1: Induction conditions for Rsn-2

The results indicate that the best induction condition for Rsn-2 are 800 mM and 20 hours of growing at 37°C with a production of 0.4253 mg/mL

Foamability

The foamability of the expressed Rsn-2 was determined by testing the sonicated cultures, manually agitating a 15 mL corning tube containing 5 mL of the sonicated protein and observing the volume of the foam produced Measurements were taken at 0, 1, 3, 5, 10, 15 and 30 minutes, after that, the measurements were taken every 30 minutes until 12 hours after the agitation

Figure 2: Water (negative control), Rsn-2, Rsn-3, Rsn 3-5, SDS (positive control), all agitated 30 second

The foamability is obtained by FV/SV where FV is foam volume and SV is solution volume. Ranaspumin-2 made a Foam volume of 4 mL with a 5 mL solution, hence its foamability is of 0.8 (relative number)

Figure 3: Foamability of Rsn-2

Passing the 12 hours the behaviour of the foam was analyzed in order to see how durable it is

Figure 4: Foam volume over 12 hours

It took 8.5 hours for the foam to go from 4 mL to 2 mL (Its half life), and at the final measurement it still had 1 mL of foam