Implementation
We demonstrate a new Tyrian Purple-producing process using Trp 6-halogenase, tryptophanase and monooxygenase in E. coli by applying a consecutive two-cell reaction system, and increased the reaction efficiency by In-Fusion and CRISPR/Cas9 technologies. This way could be an alternative solution for overcoming the problem of environmental pollution and high costs in chemical synthesis.
We successfully produce the Tyrain Purple in laboratory
Product making and Dyeing
In order to make Tyrian purple come onto the market and popularize bacterial dyes to the public, we make a great effort on dyeing. We looked up the literature and explored in the laboratory, also we consulted experts and companies in dye field. Finally we got a good dyeing scheme which has a successful effect and environmentally friendly steps. The steps are as follows:
· After the reaction ,we collected the cells and washed with water several times and freeze-dried, added dyeing solutions.
· Then we prepared by heating with stirring on the 75–80 °C hot plate for 15 min.
· After the heating, NH4Cl 0.5 g was added to the dyeing solution and subsequent heating was repeated at 75–80 °C.
· The fabrics were soaked into the solution and dyed using an automatic dyeing machine.
· Finally, obtain a piece of beautiful purple fabric by washing with acetic and water.
Target user and Commercial availablity
We also compared cell dye with chemical dye, the result suggesting that both types have similar color features and dyeing efficiency. So, our product is commercially available and can provide materials to dye factories (more is discussed in our Human Practices section). Moreover, we expect the concept of cell dye could be promoted, more dyes could be synthetized to reduce pollution and waste caused by pure chemical synthesis.
Compared biosynthetic method with chemical synthesis:
Synthesis costs 6890$/g.
Method costs 4.124$/g.
Compared our cell dye with those dyes in the market:
✔ Friendly to the environment
✔ Safe for production workers
More products for market
To meet the different needs of the market, we have designed two product forms-- dry products and wet products.
The dry products are adapted to long-distance transportation and long-term deterioration. We produce dyes by lyophilization, which is convenient for transportation and storage without compromising the dyeing effect. In the laboratory, we were surprised to find that 50 mg of lyophilized powder can be reconstituted into over 200 ml of staining solution.
The dry products in our lab
The wet products are simpler and cheaper, it will be widely used in short-distance applications. Moreover, we will continue to explore the establishment of biological factories to realize the integration of "fabric-dye-dyeing".
Assembly line
Safety and risk assessment
To ensure the safety of dying process and prevent our bacteria from escaping to environment, we used the Light Activated Kill Switch which designed by team USTC 2019. N-terminal（http://parts.igem.org/Part:BBa_K2660006）and C-terminal（http://parts.igem.org/Part:BBa_K2660007） domain of Cas9 are separately linked to nMag - Light-activated protein(http://parts.igem.org/Part:BBa_K2660009) and pMag - Light-activated protein(http://parts.igem.org/Part:BBa_K2660008)by a 10 aa linker (http://parts.igem.org/Part:BBa_K105012). The cut process is guided by gRNA, and Cas is used as the "killer".When irradiated by a blue light， pMag and nMag will dimerize，then Cas9 is activated to cut E.coli's essential genes .
References
[1] Ramaiah, G. B. & Ari, A. P . Evaluation of color strength (K/S) values of cotton fabrics dyed with reactive dye and treated with silver nanoparticles. AIP Conf. Proc. 2162, 020111 (2019).
[2] Wardman, R. H. An Introduction to Textile Coloration: Principles and Practice (John Wiley & Sons, 2017).