DETECTION OF A SINGLE-LOCUS GENE ON CHANNEL CATFISH CHROMOSOMES BY IN-SITU POLYMERASE CHAIN-REACTION

An in-situ polymerase chain reaction (ISPCR) procedure was applied to chromosomal localization of the gene, Ig H, encoding the immunoglobuli n heavy chain of channel catfish (Ictalurus punctatus). Metaphase chro mosomes were prepared by a replication banding procedure and subjected to ISPCR using biotin-labeled primers. The hybridization signals were detected with an avidin-fluorescein isothiocyanate (FITC) based metho d, and chromosome bands revealed by simultaneous or sequential treatme nt methods. Standard fluorescent in-situ hybridization (FISH) was perf ormed on chromosome preparations to compare with the ISPCR procedure. The Ig H gene was detected at the telomeric position of a chromosome w ith a relative length of 3.2 +/- 0.2%. The Ig H-bearing chromosome det ected by the FISH method was identical to that found by ISPCR procedur e. Visibility of chromosome bands was reduced by heat and salt treatme nts and could not be analyzed after thermocycling. Therefore, specific identity of the chromosome bearing the Ig H gene remains unknown. Ban ding of fish chromosomes is difficult and poses a barrier for applying current molecular techniques to physical mapping of teleost genomes. Application of the ISPCR to chromosomal mapping is new for fish specie s and is only in initial stages of development for higher vertebrates. (C) 1997 Elsevier Science Inc.