Overview

     In our project, the melA gene was used to produce L-dopaquinone with the precursor Tyr. After the addition of Sec, selenomelanin is expected to be produced. We improved the biobrick BBa_K274001 from 2009 iGEM Cambridge team by adding tyrP gene, which encodes for tyrosine transporter protein. With the addition of TyrP, E. coli was allowed to have higher tyrosine uptake. Therefore, E. coli would be capable of producing a greater amount of melanin.

Results

Construction Success

    As seen in Fig. 1, tyrP was successfully constructed into the plasmid pSUI-P_trc-melA-B0015 (BBa_K4171019).

Fig. 1. Confirmation of pSUI-NPS-tyrP-P_trc-melA-B0015 by double digestion. M: Marker; Lane 1: pSUI-tyrP-P_trc-melA-B0015 (6800 bp); Lane 2: pSUI-NPS-tyrP-P_trc-melA-B0015 without digestion (negative control).

Melanin Production Comparison

    To confirm the effectiveness of TyrP, we cultured bacteria with different plasmid, pSUI-P_trc-melA-B0015 (BBa_K4171019) and pSUI-NPS-tyrP-P_trc-melA-B0015 (BBa_K4171024), respectively. The effect of TyrP was examined through the comparison of melanin production. The results of melanin production were measured by OD₄₀₀.

Fig. 2. Relative melanin production of E. coli with and without TyrP.

    As the result shows, the production of melanin increased significantly with the addition of TyrP and indicated the effectiveness of TyrP on melanin production optimization. For more information, please visit our Results page.