IF BLUE CATFISH STEM CELLS COULD BE TRANSFERRED TO THE GONADS OF STERILE TRIPLOID CHANNEL CATFISH MALES, THE CHANNELCATFISH MALES COULD PRODUCE BLUE CATFISH SPERM AND NATURALLY MATE WITH CHANNEL CATFISH FEMALES. THE RESULTING PROGENY WOULD BE HYBRIDS AND A MORE NATURAL AND EASIER METHOD OF MAKING HYBRIDS (A PROCESS CALLED XENOGENESIS) WOULD THEN BE POSSIBLE, WHICH WOULD GREATLY BENEFIT THE US CATFISH INDUSTRY. OUR LONG-TERM GOAL IS TO DEVELOP XENOGENESIS (METHOD OF REPRODUCTION IN WHICH SUCCESSIVE GENERATIONS DIFFER FROM EACH OTHER) RESULTING IN XENOGENS (AN ORGANISM COMPRISED OF ELEMENTS TYPICALLY FOREIGN TO ITS SPECIES) TO INCREASE EFFICIENCY OF REPRODUCTION FOR CATFISH HYBRIDIZATION.WE HAVE MADE CHANNEL FEMALE X BLUE MALE CATFISH HYBRID USING XENOGENESIS ON A SMALL-SCALE. OUR OVERALL OBJECTIVE IS TO IMPROVE THE EFFICIENCY OF XENOGEN PRODUCTION BY IMPROVING CELL IMPLANTATION AND PROLIFERATION TO PRODUCE XENOGENIC BROOD STOCK WITH GOOD FERTILITY TO MAKE LARGE-SCALE APPLICATION FEASIBLE.TO ACCOMPLISH THE OVERALL OBJECTIVE, THE PRIMARY OBJECTIVES OF THE PROJECT ARE:1) INCREASE THE AVAILABILITY AND EFFICIENCY OBTAINING DONOR BLUE CATFISH STEM CELLS A) IDENTIFICATION OF THE LIFE STAGE YIELDING THE GREATEST PERCENTAGE AND QUANTITY OF STEM CELLS. B) DEVELOP A DEPENDABLE PROTOCOL FOR FREEZING/THAWING SPERMATOGONIA 2) INCREASE THE TRANSPLANTATION, COLONIZATION AND PROLIFERATION RATES OF DONOR BLUE CATFISH CELLS INTO HOST CHANNEL CATFISH TRIPLOIDS A) IDENTIFY LIFE STAGES OF THE RECIPIENT WITH THE GREATEST TRANSPLANTATION RATES B) EVALUATE SURVIVAL AND COLONIZATION OF SPERMATOGONIAL STEM CELLS TREATED WITH ROCK INHIBITOR Y-27632 AFTER ENZYMATIC DISSOCIATION C) COMPARE THE COLONIZATION OF FRESHLY ISOLATED AND SORTED SSCS.THE EXPECTED RESULT IS AN IMPROVEMENT IN THE XENOGENESIS METHOD FOR MAKING HYBRID CATFISH TO ALLOW THE PROCESS TO BE USED ON A COMMERCIAL SCALE. THE ANTICIPATED IMPACT IS THE INCREASE IN HYBRID CATFISH FRY PRODUCTION AND PRODUCTION EFFICIENCY, RESULTING IN GREATER PROFITABILITY AND SUSTAINABILITY FOR THE US CATFISH INDUSTRY. THIS SHOULD ENABLE THE CATFISH INDUSTRY TO BE MORE COMPETITIVE WITH FOREIGN IMPORTS OF CATFISH.