** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** CATFISH IS THE MOST IMPORTANT AQUACULTURE SPECIES IN THE US. ALTHOUGH CHANNEL CATFISH IS MORE USED IN THE US AQUACULTURE THAN BLUE CATFISH, THE INTERSPECIFIC F1 HYBRID PRODUCED BY MATING FEMALE CHANNEL CATFISH WITH MALE BLUE CATFISH IS CURRENTLY THE PREDOMINANT GENOTYPE USED IN THE US CATFISH INDUSTRY. HOWEVER, PRODUCTION OF THE INTERSPECIFIC HYBRIDS IS HINDERED BY REPRODUCTIVE ISOLATION. THE SUCCESS RATES FOR HYBRID PRODUCTION VARY GREATLY DEPENDING ON THE COMPATIBILITY OF BLUE CATFISH USED IN THE MATING. UNDERSTANDING REPRODUCTIVE ISOLATION AND MOLECULAR BASIS OF SEX DETERMINATION WILL ALLOW THE SCREENING OF CHANNEL CATFISH AND BLUE CATFISH FOR EFFICIENT AND RELIABLE PRODUCTION OF THE HYBRIDS. AT PRESENT, THE MECHANISM OF SEX DETERMINATION IN CHANNEL CATFISH IS KNOWN TO BE CONTROLLED BY DNA METHYLATION, BUT THAT IN BLUE CATFISH IS UNKNOWN. WE HYPOTHESIZE THAT WHILE THE MECHANISMS OF SEX DETERMINATION BETWEEN CHANNEL CATFISH AND BLUE CATFISH CAN BE SIMILAR, THE DETAILED PATTERNS OF METHYLATION AND DIFFERENTIALLY EXPRESSED GENES MAY BE DIFFERENT SUCH THAT THEY ARE NOT COMPATIBLE. THE SPECIFIC OBJECTIVES OF THIS PROJECT ARE TO DETERMINE THE MECHANISM OF SEX DETERMINATION IN BLUE CATFISH, TO COMPARE THAT WITH THE MECHANISM OF SEX DETERMINATION IN CHANNEL CATFISH, AND DETERMINE THE GENES INVOLVED IN REPRODUCTIVE ISOLATION AND HYBRID INCOMPATIBILITY. THESE GOALS WILL BE ACHIEVED BY ANALYSIS OF WHOLE GENOME PROFILES OF DNA METHYLATION DURING EARLY SEX DEVELOPMENT AND DIFFERENTIATION BETWEEN CHANNEL CATFISH AND BLUE CATFISH, BETWEEN COMPATIBLE AND INCOMPATIBLE BLUE CATFISH STRAINS, AND BETWEEN RECIPROCAL HYBRIDS (HYBRIDS PRODUCED BY MATING FEMALE CHANNEL CATFISH WITH MALE BLUE CATFISH, AND BY MATING FEMALE BLUE CATFISH WITH MALE CHANNEL CATFISH). ACHIEVING THESE GOALS WILL ALLOW US TO DETERMINE THE DIFFERENCES OF GENOMIC ARCHITECTURE OF THE SEX CHROMOSOMES OF CHANNEL CATFISH AND BLUE CATFISH, AND THE DIFFERENCES IN METHYLATION PROFILES AND DIFFERENTIALLY EXPRESSED GENES.UPON SUCCESSFUL COMPLETION OF THIS PROJECT, THE FUNDAMENTAL BASIS FOR REPRODUCTIVE ISOLATION BETWEEN CHANNEL CATFISH AND BLUE CATFISH WILL BE REVEALED, WHICH WILL FACILITATE THE DEVELOPMENT OF TECHNOLOGIES AND APPROACHES TO OVERCOME THE REPRODUCTIVE ISOLATION. IN THE LONG TERM, THIS PROJECT WILL LAY THE FOUNDATION FOR MORE EFFICIENT PRODUCTION OF THE INTERSPECIFIC HYBRIDS FOR THE CATFISH AQUACULTURE INDUSTRY.