Simultaneous genotyping of 4 SNPs in promoter III of the ovine ACACA

• The genetic variability in the PIII promotor of the ACACA gene in different sheep breeds kept in the Czech Republic.
• By sequencing analysis has been identified 2 novel SNPs.
• Developed rapid, efficient and sensitive method for typing 4 SNPs using PEA.
• PEA will serve as a useful genotyping technique for routine simultaneous typing of 4 genetic variants in a single gene.
• Genotype AG/GT/CG/CC occurred in all 4 observed sheep breeds.

The acetyl-CoA carboxylase-α (ACACA) gene is the rate-limiting enzyme for the biosynthesis of fatty acids in animal tissues. By sequencing analysis in promoter III of the ovine ACACA we identified 2 novel SNPs (A > G at 1168 bp and a T nucleotide insertion at 1182 bp) (GenBank accession number AJ292286). The novel variant (A > G at 1168 bp) was present in Cigaja, Sumavka, and Improved Valachian breed kept in the Czech Republic and T insertion at 1182 bp was present in the East Friesian breed. A protocol for rapid efficient simultaneous genotyping of ACACA variants using the primer extension method (PEA) was described, and a total of 171 animals belonging to sheep breeds: Cigaja (9), Sumavka (87), East Friesian (59) and Improved Valachian (16) were genotyped. Variant alleles are: A/G at 1168 bp, G/T at 1330 bp, C/G at 1138 bp and C/T at 1430 bp.The genotype combination AG/GT/CG/CC occurred in all 4 breeds kept in the Czech Republic with frequency of occurrence ranging from 15.25% to 44.45%. Allele frequencies of the Cigaja breed differ significantly from the other Czech sheep breeds.