Bozeman Fish Health Center

DNA ploidy analysis of hatchery-produced fish using flow cytometry technology at the Bozeman Fish Health Center

Status: Ongoing

States: The Bozeman Fish Health Center provides services to western states including Colorado, Kansas, Montana, Nebraska, Nevada, North Dakota, South Dakota, Utah, and Wyoming, plus additional states upon request. 

Contact:

USFWS Bozeman Fish Health Center, Bozeman, MT

Author: Renee Yamamoto

Triploid fish:

Triploid fish that are created at fish hatcheries have three sets of chromosomes instead of the normal two sets of chromosomes making them sterile. This condition can occur naturally in populations. Triploid fish are stocked into waters to support recreational fisheries while reducing the risk of genetic mixing between hatchery and wild fish. Triploid fish may have a smaller impact on natural systems than reproducing fish. 

Triploid induction at a fish hatchery:

The process of creating triploid fish is called triploidy induction. This involves giving a normal egg with two sets of chromosomes a third set of chromosomes. Each species of fish has a slightly different process for inducing triploidy. For rainbow trout, creating triploids at a fish hatchery adds a few more steps to the normal fish rearing process . First, the eggs are stripped from the female fish. Then the eggs are fertilized with milt from the male fish. After this, there is a set waiting period, which is determined each day based on the water temperature. At the precise time, the eggs are exposed to high amounts of pressure. This pressure causes the eggs to retain a third set of chromosomes that would normally be expelled. The triploid eggs can then be shipped to partners requesting them, and the resulting fish have extra copies of DNA in each cell, which causes the fish to be sterile. Stocking triploids can support recreational sport fisheries.

Determination of Ploidy Status:

The Bozeman Fish Health Center uses flow cytometry to determine the triploidy induction success of hatchery fish. A triploid fish has three copies of nuclear DNA, whereas a diploid, or naturally produced fish, has only two copies. Nuclei from a triploid fish contain more DNA than diploid nuclei because of the increased nuclear DNA content. A flow cytometer measures fluorescence from tagged DNA and compares samples to a known standard. Samples that can be used for ploidy testing include fresh fish blood, live whole fish, and eyeball fluid.

Principles of flow cytometry laboratory methods at the Bozeman Fish Health Center are based on standardized methods (Jenkins and Thomas 2007; Jenkins et al., 2017; Jenkins et al., 2019) and the Ploidy Predictor Calculator for use with mixed batches of cells/larvae of unknown ploidy status (https://warcapps.usgs.gov/gs-eco/warc/ploidy).

References:

Jenkins, J.A. and R.G. Thomas. 2007. Use of eyeballs for establishing ploidy of Asian carp. North American Journal of Fisheries Management 27:1195-1202.

Jenkins, J.A., Draugelis-Dale, R.O., Glennon, R.P., Kelly, A.M., Brown, B.L., and Morrison, J.R. 2017. An accurate method for measuring triploidy of larval fish spawns. North American Journal of Aquaculture 79:224-237

Jenkins J.A., Chauvin M.D., Johnson D, Brown, B.L., Bailey, J., Kelly, A.M., and Kinter, B.T. 2019. Defensible standardized ploidy assessments for Grass Carp (Ctenopharyngodon idella, Cyprinidae) intercepted from the commercial supply chain. Journal of Great Lakes Research 45:371-383