Plant & Animal Genome VI Conference

W4

PCR TEST FOR VARIANT ALLELES OF THE LATE-FEATHERING (K) LOCUS OF THE DOMESTIC CHICKEN

Poultry Science Department, POSC O-114, University of Arkansas, Fayetteville, AR 72701

The sex-linked late-feathering allele of the K-locus of the chicken is associated with a duplication of a segment of the Z chromosome. An intact avian leukosis retrovirus (EV21) has inserted into one copy of this repeated region. Negative production characteristics have been associated with the presence of the ev21 gene. Evidence indicating the existence of ev21 minus late-feathering alleles has been published, but these alleles were never genetically verified. The most common late-feathering allele can be identified by a HaeIII RFLP differentiating the ev21 occupied repeat from the unoccupied repeat. Two PCR primer sets were designed to amplify a 327 bp ev21 junction fragment and a fragment containing the HaeIII polymorphic site. An 8 bp insertion disrupting the HaeIII site of the occupied repeat allowed the differentiation of the 234 and 242 bp HaeIII site-containing PCR fragments without resorting to digestion with HaeIII restriction enzyme. The agarose gel additive Synergel^TM (equivalent to 4% agarose) was found to be an economical replacement for MetaPhor^TM agarose in the electrophoretic separation of the two polymorphic fragments. A third polymorphic HaeIII site-containing fragment was found in broiler chicken populations which was intermediate in size (approximately 237 bp). Because of the duplication some males were found to have all three of the HaeIII site-containing polymorphic fragments. Synergel^TM allowed the detection of all three fragment sizes (234, 237, and 242). Both primer sets are co-amplified in a single PCR reaction and provide a rapid test for the different alleles of the late-feathering locus. This PCR test is currently being used to screen late-feathering lines for variant late-feathering alleles, especially those which may lack ev21.