Glofish

1. Glofish

2. GFP

3. Sources of GFP • GFP (Aequorea victoria, jellyfish), • GFP (Renilla reniformis, sea pansy), • dsRed (Discosoma, mushroom coral), • eqFP611 (Entacmaea quadricolor, sea anemone), • RTMS5 (Montipora efflorescens, stony coral), • dronpa (Pectiniidae, chalice coral), • KFP (Anemonia sulcata, Venus hair anemone), • eosFP (Lobophyllia hemprichii, open brain coral), • dendra (Dendronephthya, octocoral)

4. First Transgenic Animal in US market The New York Times Nov 22, 2003 Gene-Altering Revolution Nears the Pet Store: Glow-in-the-Dark Fish GloFish casts light on murky policing of transgenic animals Nature 27 November 2003 www.glofish.com Marketed without regulatory environmental review. FDA is leading authority.

5. GloFish • patented and trademarked brand of genetically modified (GM) fluorescent fish • variety of different GloFish are currently on the market • Zebrafish were the first GloFish available in pet stores • now sold in bright red, green, orange-yellow, blue, and purple fluorescent colors • Glofish retail for about $5 per fish. Normal zebrafish cost around one tenth of the price

6. Original Zebrafish • The original zebrafish (or zebra danio, Daniorerio) from which the GloFish was developed is a native of rivers in India and Bangladesh • Measures 3 cm long • Has gold and dark blue stripes • Over 200 million have been sold in the last 50 years in US • They have never established any wild populations because they are tropical, unable to survive in North America’s temperate climate

7. Researchers in Singapore added a fluorescence gene from a sea coral to zebra danio eggs to produce glofish • In 1999, Dr. ZhiyuanGong and his colleagues at the National University of Singapore worked with the gene encoding GFP or green fluorescent protein extracted from a jellyfish • They inserted the gene into zebrafish embryo and it integrated into the zebrafishgenome • which caused the fish to be brightly fluorescent under both natural white light and ultraviolet light • Goal: develop a fish that could detect pollution by selectively fluoresce in the presence of environmental toxins • Developing a constantly fluorescing fish was the first step • Patent filed for their process • Soon other fluorescent zebra fish were developed using genes that produced different color: red from sea coral and orange-yellow from a variant of the jellyfish gene

8. Fluorescentmedaka • Thena team of researchers at the National Taiwan University, headed by Professor Huai-Jen Tsai (蔡懷)succeeded in creating a medaka (rice fish) with a green fluorescent color as a model • A separate deal was made between Taikong, the largest aquarium produce in Taiwan and the Taiwanese researchers to market the green medaka in Taiwan under the name TK-1 • In spring 2003, Taiwan became the first to authorize sales of a genetically modified organism as a pet. 100,000 were sold in less than a month at US$18.60 each

9. Yorktown Technologies • Deal between scientists from NUS (National University of Singapore) and businessmen Alan Blake and Richard Crockett from Yorktown Technologies, L.P., a company in Austin, Texas, • Yorktown obtained worldwide rights to market the fluorescent zebrafish, which Yorktown branded as "GloFish” • GloFish have been marketed in the United States since late 2003 • there have been no reports of any ecological problems associated with their sale

10. Reproduction limited • speculation that the eggs are pressure treated to make them infertile • some GloFish are fertile and will reproduce in a captive environment • GloFishFluorescent Fish License states "Intentional breeding and/or any sale, barter, or trade, of any offspring of GloFish fluorescent ornamental fish is strictly prohibited”

11. Reduced fertility • GloFish have reduced fertility compared to the species, possibly due to higher energy costs associated with fluorescence, instead of other bodily functions

12. Sale of Glofish restricted • Canada prohibits import or sale of the fish, due to what they report is a lack of sufficient information to make a decision with regard to safety • The import, sale and possession of these fish is not permitted within the European Union • Sale or possession of Glofish remains illegal in California due to a regulation that restricts all genetically modified fish, due to concern about a fast-growing biotech salmon

13. Glofish chemical detector • GloFish also have been used for experimental research. • The alterations in the zebrafish's genes has given the organism the ability to fluoresce as a bio-indicator. • This genetic ability has been used to detect pollution and other chemicals. • Chemicals that mimic natural estrogens have well-documented effects on the reproductive systems of invertebrates, typically acting as endocrine disruptors, and glofishflourescence is used to detect levels of estrogenic chemicals • Muscles such as the heart have a larger effect of estrogen than the liver.

14. History of Glofish • First -the red fluorescent zebrafish, trademarked as "Starfire Red” • Yorktown Technologies released a green fluorescent zebrafish and an orange-yellow fluorescent zebrafish in mid-2006 • In 2011, blue and purple fluorescent zebrafish were released. These lines of fish are trademarked as "Electric Green", "Sunburst Orange", "Cosmic Blue", and "Galactic Purple", and incorporate genes from sea coral • In 2012, Yorktown Technologies introduced a new variety of "Electric Green" GloFish, derived from a different species of fish, the black tetra • Followed in 2013 by the “electric green” barb which is a variety of tiger barb and a “sunburst orange” tetra and a “moonrise pink” tetra, the first flourescent pink to be marketed (from YT)

15. Cosmic blue glofish 36085b

16. Starfire red zebra glofish 36055b red glo 36055a red

17. Galactic purple zebra? • GloFish® Galactic Purple fluorescent fish bring color and excitement to any aquarium • A traditional white aquarium light will work for daytime viewing, but won’t give the fish its best color • An actinic lightwill give the fish an exciting and uniquely fluorescent look, particularly in low lighting environments - the coloryoulook for in a glofish • Ifthe room is completely dark, a black light will create the appearance that the fish are glowing in the dark http://www.petsolutions.com/C/Live-Freshwater-Fish-Danios/I/Glofish-Cosmic-Blue-Zebra.aspx

18. Glofish: green tetra GloFish® Electric Green Tetras are not injected or dyed. They breed and reproduce naturally with this fluorescent color. Electric Green Tetras look great under either regular daylight bulbs or under an actinic bulb.

19. Green Tetra • GloFish® Electric Green Tetras are beautiful freshwater Tetra fish that glow under black lights. • Unlike other GloFish® fluorescent fish, which are Zebra Danios, the Electric Green Tetra is a variation of the White Skirt Tetra. Electric Greens are created with selective breeding of the White Skirt Tetra, using a certain gene in the fish to produce the Electric Green color • One of the hardier Tetras,itprefers to school.They do best when there are more than 3 Electric Green Tetras in the school • While requiring some swimming space, Electric Greens will also appreciate cover along the perimeter of the aquarium • Generally peaceful, they can match up with just about any other type of fish that is not large enough to eat them • While the Electric Green Tetra might prefer softer, acidic water conditions, most can adapt to a wide range of water parameters and will do fine in a well-lit aquarium

20. Purposes of Transgenics • Industrial • Medical • xenotransplantation • nutritional supplements and pharmaceuticals • Basic Research • gene function • gene expression patterns • Agriculture • size • quality • disease resistance

21. Blastocyst Injection Blastocyst ES cells http://tasq.uq.edu.au/blasto.html

22. Transgenic Fish • Tilapia • Salmon/trout • Catfish • Can grow up to 6 times faster than wildtype fish • Most have extra copies of growth hormone (GH) gene TransgenicWildtype http://www.nature.com/nbt/journal/v19/n6/images/nbt0601_500a_I1.jpg

23. Fish Transgenics • The transgene used to increase growth utilizes an antifreeze protein promoter connected to the GH cDNA • As water temperature drops the GH gene is turned on • The fish continue to grow when normally they would not • What happens if they escape?

24. Could become first transgenic animal approved for large-scale farming and human food The New York Times Age = 7.5 months Transgenic = 1.2kg Unmodified = 200 g Engineered with ocean pout antifreeze gene promoter + chinook salmon growth hormone gene Http://webhost.avin.net/afprotein/peidof.htm