Team:TzuChiU Formosa/Modeling

Gluconacetobacter hansenii which carries the acsABCD protein has the ability to produce cellulose is our main character in our project. Our project is to transform acsABCD genes into E. coli to monitor the production of cellulose production. We collect the protein produced by the transformed E.coli and Gluconacetobacter hansenii in different time point to monitor the expression of acsABCD gene products and the production of cellulose.

Method 2 : Benedict’s test

The bacterial culture which have been induced were collected and the bacteria is to remove, the end product is added with cellulase until the cellulose is fully decomposed. Benedict solution caused the formation of brick red precipitate, the concentration of cellulose is then tested with OD645.

Cellulase is used to break down the cellulose into monosaccharide. Cellulose is a glucose polymer connected through a beta (1-4) glycosidic linkages. Benedict’s test is carried out to test the presence of reducing sugar, such as. The reducing sugar reduces copper(II) ions in these test solutions to copper(I), which then forms a brick red copper(I) oxide precipitate. The color would range from green to brick red respectively depends on the amount of reducing sugar present in the solution. It can detect the concentration of the reducing sugar under the absorbance condition _OD645.

Cellulose (beta-1,4 glucan) is the most plentiful biopolymer in nature and is an crucial raw material for many industries. It is synthesized as extracellular fibrils by cellulose synthase not only in plants but also in some bacteria.

Bacteria with cellulose synthase gene use Isopropyl-β- D -1-thiogalactopyranoside (IPTG) as the inducer stimulating the production of protein. In our system, IPTG act as the inducer of the R0011 promoter, which then activate the operon. The activated acs operon encodes the cellulose synthase to synthesize cellulose. The reaction lasts two hours and every two hour we would need to collect the purified cellulose. The nett weight is recorded and the mechanism of cellulose activity on pure cellulose substrates is identified. Lastly, the Benedict’s solution is added to find out the absorbance (optical density [O.D.]) value, in order to calculate the amount of monosaccharide that cellulose can produce.

Bacterial cellulose