(Difference between revisions)

Categories :

Home Team Sponsors Parts Submitted to Registry Image Gallery Leave a Message!

Project :

Overview Sensitivity Tuner --- Characterisation --- Modelling Colour Generators --- Carotenoids (Orange/Red) --- Melanin (Brown) --- Violacein (Purple/Green) The Future Safety

Notebook :

Week 1 Week 2 Week 3 Week 4 Week 5 Week 6 Week 7 Week 8 Week 9 Week 10

Team Logistics :

Protocols Stock List Research Shared Links and Help

Melanin Pigment

Introduction

The MelA gene codes for a tyrosinase. Tyrosinases catalyze two reactions, as described in the figure below. Melanin is a macromolecular compound produced by the polymerization of the quinone product of the second reaction, and has a characteristic brown colour.

From H. Claus and H. Decker, Bacterial tyrosinases, Syst. Appl. Microbiol. 29 (2006), pp. 3–14. [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B7GVX-4H21K91-1&_user=1495569&_coverDate=01%2F24%2F2006&_fmt=full&_orig=search&_cdi=20442&view=c&_acct=C000053194&_version=1&_urlVersion=0&_userid=1495569&md5=de451c5e1d1d18ad12b7e39b10b80408&ref=full]

Our MelA gene is from Rhizobium etli. Further, it is a mutant; it has a C to T substitution at the 1,000th nucleotide, which reduces the amount of time before melanin production is visible. (Santos, C. N., and G. Stephanopoulos. 2008. Melanin-based high-throughput screen for L-tyrosine production in Escherichia coli. Appl. Environ. Microbiol. 74:1190-1197 [http://aem.asm.org/cgi/reprint/74/4/1190])

Previous Work

Currently our MelA gene is held on the following plasmid, provided by Duncan Rowe.

We isolated it using miniprep to make stocks of our own.

Action plan of our team

Our action plan is as follows:

1. Test for melanin production

2. Isolate MelA gene in biobrick form

3. Integrate Mel biobrick into system (e.g amplification of logic gate system)

1. Melanin Production

Grew the plasmid in TOP10 E. coli

Pigment was clearly produced.

2. Constructing MelA biobrick

Primer design:

Primers were designed to isolate the melanin with the prefix and suffix. We are currently looking at adding a ribosome binding site to the the primers in order to produce a complete biobrick.

PCR reactions

Practical details to follow as soon as primers arrive.

Characterisation of Biobricks and colour output

Test compatibility with other biobricks

Hopefully we will be able to integrate this into the amplification processing system.