genetic manipulation in fish

july 08

Sex Ratio Manipulation
– For aquaculture or stocking, it may be desirable to use only one sex due to differences in growth rate or characteristics concerned with maturity and reproduction,
– Sex ratio can be manipulated either directly (using steroid hormones to alter the sex ratio in fry during sexual differentiation) or indirectly (manipulating the sex determination system of broodstock so that selected fish produce monosex offspring).

Chromosome Manipulation: Polyploidy
– Nearly all species of fish are diploid (2n),
– Triploidy (3n) fish exhibit good growth and condition over the spawning period comparing with 2n fish,
– Tetraploidy (4n) fish produce eggs and sperm which are 2n, thus a cross between a 4n fish and a 2n fish will produce 3n offspring without the need to use shocks to induce triploidy.

Gynogenesis and Androgenesis
– Gynogenesis is a tool for producing all-females where females are homogamety, has been used in the study of sex determination and gene recombination and as a tool for inbreeding and the production of clonal lines,
– Androgenesis is a tool for producing all-males where males are homogamety.

recapped from N. Pongthana

genetic manipulation in practice

july 08

Sperm:
– Strip milt from males,
– dilute sperm in 0.85% NaCl (1/200 dilution and then 1/4000 dilution),
– count all of the sperm in 5 blocks (5×5) of hemacytometer,
– calculate sperm concentration (10^10)=(total count/80)x4000Ax4000Bx1000C (A : inverse of dilution, B : volume of hemacytometer cell = 1/4000 uL, C : to convert from concentration per uL to concentration per ml,
– prepare sperm at a final concentration of 8×10^8 sperm/ml, volume of sperm (uL) = 5000x8x10^8/(sperm concentration)

Egg
– Strip the eggs into a glass container
– Place eggs ~10 ml

Triploid (3n)
– Fertilize eggs in one dish with 5 ml of diluted sperm,
– mix eggs and sperm gently with just enough water to cover the eggs, then slowly keep adding water,
– cold shock eggs by pouring eggs through a fine mesh sieve at about 90 seconds post-fertilization and immersed in a water bath at 2 oC, and kept there for 10 min,
– Empty the eggs into a large container of water at the same temperature as the incubation system and from there transferred to one incubator

Meiotic gynogenesis (2n)
– Prepare one petri dish containing 5 ml of diluted sperm,
– place the petri dish into the UV chamber and irradiate for 60 seconds (agitating continuously during UV-irradiation),
– fertilize eggs in one dish with 5 ml of UV-irradiated sperm, mix eggs and sperm gently with just enough water to cover the eggs, then slowly keep adding water,
– cold shock eggs by pouring eggs through a fine mesh sieve at about 90 seconds post-fertilization and immersed in a water bath at 2 oC, and kept there for 10 min.
– empty the eggs into a large container of water at the same temperature as the incubation system and from there transferred to one incubator

recapped from N. Pongthana

fish mating system

july 07

…….under corrected…….