Team:Vanderbilt/Project

From 2014.igem.org

Revision as of 02:57, 16 October 2014 by Shiltsj (Talk | contribs)




Home
Team Official Profile Project Parts Notebook
Safety Attributions

Introduction


Sage plant, <i>Salvia officinalis</i>, growing in the Vanderbilt greenhouse The production of plant essential oils and their derivatives represents an over 9 billion dollar industry when considering just their applications in the food and fragrance industries 1. A staggering 23 million kilograms of citrus oil alone are produced worldwide each year. Up until only a couple decades ago, the production of these essential oils was done exclusively by chemical extraction from plant material. However, the sudden emergence of synthetic biology a versatile and efficient tool has the potential to transform this immense industry, the products of which nearly everyone will come in contact with on a daily basis.

By harnessing existing biosynthetic pathways and introducing enzymes taken from plants into more maleable model systems, it will be possible to significantly improve on current methods of the active components of essential oils, most notably the terpenoids. While most plants express terpenes in the range of parts per million and thus require very large scale operations to be commercially viable, early forays into the biological production of terpenes have proven that it is possible to improve yields 100-fold 2. The first aspect of our project involves using the great potential of synthetic biology to design a commercially-viable strategy for the production of any class of terpenoid.

Just as important to the economic benefit of this approach is its environment benefit. With chemical terpene extraction being such a relatively inefficient process, it is necessary to process large amounts of plant material to get a substantive yield. This may not pose as significant of an issue for citrus growers, but many of the most prized compounds are taken from the rarest species of plant. The continuation of the status quo in terms of terpene extraction is not an environmentally sustainable solution. In our selection of terpenes, we placed a large emphasis on choosing compounds from some of the most rare species possible.

The best example of this idea behind our project can be seen in the gene for santalene synthase. The only species know to have genes to produce this terpene are found in remote regions of India and Australia, and one of them is listed as a vulnerable species by the IUCN. The trees can live for hundreds of years, but are the target of widespread over-exploitation, to the point that, for example, the Indian government has banned the export of sandalwood. Synthetic biology can produce the exact same active ingredients of sandalwood oil in a way that is both more economically and environmentally sound.

In order for our idea to truly be applicable to exotic and endangered species of plant, we had to take an approach that was quite different from the one most iGEM teams have historically taken. What we were looking for was a quick, inexpensive method of cloning genes that was also compatible with species that have not had their entire genomes sequenced. Often, iGEM teams resort to synthesizing their genes through third parties. However, this can be fairly costly especially given the moderately large size of many of these synthase genes, may take several weeks or even a month to finish synthesizing, and cannot be done unless the gene's sequence is known in its entirety. By going back to basics and taking raw plants as our source material, we were able to avoid these issues and demonstrated how our approach was a practically viable one.


Cadenine

Main essential oil: Cade oil
Applications: Antifungal, bactericidal, and antioxidant
Plant Species: Gossypium hirsutum (cotton) Synthase Gene: δ-cadenine synthase (E.C. 4.2.3.13)

Carene

Main essential oils: Rosemary and Cedar oil
Applications: Insecticide, anti-inflammatory, and central nervous system depressant
Plant Species: Picea abies (norway spruce) Synthase Gene: carene synthase (E.C. 4.2.3.107)

Humelene

Main essential oils: Hops oil
Applications: Culinary spice, and anti-inflamitory
Plant Species: Zingiber zerumbet (shampoo ginger) Synthase Gene: α-humulene synthase (E.C. 4.2.3.104)

Myrcene

Main essential oils: Thyme and Hops oil
Applications: Fragrance, analgesic, and anti-inflamitory
Plant Species: Perilla frutescens (Green Shiso) Synthase Gene: myrcene synthase (E.C. 4.2.3.15)

(R)-Linalool

Main essential oils: Lavender oil
Applications: Fragrance, and insecticide
Plant Species: Mentha citrata (lemon mint) Synthase Gene: (R)-linalool synthase (E.C. 4.2.3.26)

(S)-Linalool

Main essential oils: Citrus and Coriander oil
Applications: Fragrance, and insecticide
Plant Species Chosen: Arabidopsis thaliana (thale cress) Synthase Gene: (S)-linalool synthase (E.C. 4.2.3.25)

Sabinene

Main essential oils: Junier Coriander oil
Applications: Spice, and ntimicrobial
Plant Species Chosen: Salvia officinalis (sage) Synthase Gene: (+)-sabinene synthase (E.C. 4.2.3.110)

Santalene

Main essential oils: Sandalwood oil
Applications: Fragrance, antiviral, and tumor-suppressant
Plant Species Chosen: Santalum album (sandalwood tree) Synthase Gene: α-santalene synthase (E.C. 4.2.3.82)

Zingiberene

Main essential oils: Ginger oil
Applications: Flavoring and pesticide
Plant Species Chosen: Ocimum basilicum (basil) Synthase Gene: α-zingiberene synthase (E.C. 4.2.3.65)



References:
1. USDA Industrial Uses Reports. Essential Oils Widely Used in Flavors and Fragrances. September 1995.
2. Ajikumar PK, Tyo K, Carlsen S, Mucha O, Phon TH, Stephanopoulos G. Terpenoids: opportunities for biosynthesis of natural product drugs using engineered microorganisms. Mol Pharm. 2008;5(2):167-90.

Project Goals


Results and Directions

MOVE ALL RESULTS in terp table to table with column checking if plant successfully grown, genomic DNA successfully extracted, synthase gene successfully PCR isolated, plant RNA successfully extracted, synthase gene cDNA successfully reverse transcribed, gene successfully mutagenized, gene successfully expressed in E. coli or Yeast