1. Scott Williams Manager of ARI wet lab research facilities Formerly Brood Operations Manager & Geneticist for Clear Springs Foods, Inc.

2. Clear Springs Foods’ Production 22,000,000 lbs year process 85,000 lbs/day grow more than 60,000 lbs/day approximately 60% of Idaho’s production

3. Vertical Integration of Trout Production Feed manufacturing Brood operations Farm production Processing –primary –value added Distribution Marketing

4. Brood Division Egg Production & Research Egg Production: Soda Springs 71.5 Million Snake River Brood 8.1 Million Research: Genetics of growth and survival traits Trout reproduction

5. The Brood Division Two facilities –Buhl Snake River canyon Research facility Selection site Brood propagation site 5% of egg production –Soda Springs Primary egg production site

6. Brood Division movement of fish and eggs Snake River Brood StationSoda Springs Brood Station CSF farms primary site of egg production neomale production brood female selection & propagation selection program egg production Snake River FarmBox Canyon FarmCrystal Springs Farm

7. Selection Program Animal Model Genetic merit based on: Individual records Family records

8. Selection Program Primarily Growth rate Indirectly for feed conversion rate Secondarily Disease resistance

9. Size Selection Family and Within Family Selection 100 to 110 full-sib families per generation 10 families per generation from randomly bred line Families selection: –Families with poor early survival were discarded –Families with high mean weights at 273 days were saved Within the selected families: –pit tag top 15 to 20% of individuals within the largest families at 273 days

10. The Steps for Size Selection 40 days post-spawn, reduce to 600 individuals 63 days, reduce to 100 individuals 76 days, pool with another family in 100 gal tank 138 days, each family is given unique clip/brand 139 days, to raceway 273 days, final measurement 274 days, PIT tag the largest 10% of fish within the families having the largest mean weights

12. Family and Within Family Selection for Size 100 to 110 full-sib families per generation 10 families per generation from a randomly bred non-selected (control) line At 273 days Families with low mean weights are discarded Top 10-15% of the individuals from the largest families are PIT tagged

13. Odd year generation

14. Even year generation

15. Transfer of Genetic Improvement to the Farms Selected females join the ranks of the brood females after first spawn Amplification through use of selected females’ daughters become brood replacements Selected males are sires of brood replacement females

16. Brood Replacement Females Progeny of 30 sires and 30 dams Undergo mass selection Produced in 15 lots per year In the months of July through January Spawn twice in natural photoperiod Spawn two more times in light-control Used for a total of four spawns

17. Rainbow Trout Egg Production Maturation in 24 months Some females will mature precociously Depending on stock, 50% of the males will mature at 12 months Photoperiod cues reproduction Naturally spring spawners Wild stocks will spawn from Feb to June Selective breeding of domesticated fish has moved spawn time to fall - winter

18. Egg Production methods to affect egg availability Photoperiod manipulation of brood dams Skeletal photoperiod 6 hours of light  18 hours of light  6 hours of light Thermal manipulation at incubation Water chilled to slow or heated to speed development Hormonal control of reproduction Exogenous hormone applications to synchronize maturation LHRHa implants

19. Current Culture Practices modifying populations for growth and carcass traits Mono-sex culture Sexual Dimorphism: –growth rates –carcass confirmation Examples –Male culture Tilapia –Female culture Trout

20. Triploid culture 3N ploidy Are sexually non-viable (99.9%) Males will exhibit secondary sexual characteristics Current Culture Practices modifying populations for growth and carcass traits

21. Production of Normal Diploids and Triploids

22. Triploid culture 3N ploidy Are sexually non-viable (99.9%) Males will exhibit secondary sexual characteristics Triploid mono-sex female populations Will not undergo sexual maturation Will be a uniform population Will have improved growth rates over diploid female population after 18 months Current Culture Practices modifying populations for growth and carcass traits

23. Sex determination in rainbow trout Females are homogametic –Have two copies of the X chromosome (XX) Males are heterogametic –Have both an X and Y chromosome (XY) Male gametes (sperm) are sex determining –They will carry either an X or Y chromosome

24. The perfect population – (from a trout culturist perspective) All female or Monosex female First Path- Direct feminization of a mixed sex population Oral doses of estradiol Population of XX and XY genotypes All female phenotypes Problem Regulatory concerns with hormone use on food fish

25. Mono-Sex Female Population Second Path through gynogenesis Gynogenesis is an induced form of parthenogenesis –e.g. “virgin birth” of Komodo dragons Diploid organism produced without the contribution of a male parent Forms a population of XX animals in trout Gynogens are highly inbred –Heterozygosity through crossing over during meiosis

26. Production of Haploids and Gynogens

27. Mono-Sex Female Population Third path - start with masculinized females Direct masculinization of a mixed sex population Oral dose of methyltestosterone Identify XX males (neomales ) Progeny testing Use morphology to separate neomales from genetic males Neomales become sires to mono-sex female population

28. Mono-Sex Female Population (commercial method) Generate female population through gynogenesis Sex-reverse gynogens Gynogens become neomales (XX males) Neomales are sires for all female offspring Perpetuate female population by sex-reversing offspring for sires of the next generation Total elimination of the Y chromosome from the population

29. Sex-Reversal and Mono-Sex Female Production