Allelic polymorphism of Makoei sheep leptin gene identified by polymerase chain reaction and single strand conformation polymorphism
Leptin, a 16-kDa protein secreted from white adipocytes has been implicated in the regulation of food intake, energy expenditure and whole-body energy balance in animals. In the present study, the polymorphism of the leptin gene (LEP) of Makoei sheep was investigated by polymerase chain reaction and single strand conformation polymorphism technique (PCR–SSCP). Genomic DNA was extracted from whole blood samples collected from 130 sheep. A 471 bp LEP exon 3 segment was amplified by standard PCR, using the locus specific primers. PCR products were subjected to SSCP denaturation and polyacrylamide gel electrophoresis. SSCP bands were visualized with silver staining. Five SSCP patterns, representing five different genotypes, were identified. The frequencies of the observed genotypes were 0.17, 0.09, 0.14, 0.37 and 0.23 for AB, BB, AC, BC and CC, respectively. Allele frequencies were 0.15, 0.37 and 0.48 for A, B and C, respectively. The observed heterozygosity (Hobs) value for LEP gene was 0.6769. The chi-square test showed that there was no significant deviation (P>0.01) from Hardy-Weinberg equilibrium for this locus in Makoei sheep population.
Key words: Leptin gene (LEP), polymerase chain reaction (PCR), single strand conformation polymorphism (SSCP) Makoei sheep.