PCR-based genotyping of SNP markers in sheep

Autor(es)

Meo Niciura, Simone Cristina ^[1] ; Cruvinel, Giovanna Gabrielle ^[2] ; Moraes, Caroline Valerio ^[3] ; Bressani, Flavia Aline ^[1] ; Malago Junior, Wilson ^[1] ; Benavides, Magda Vieira ^[4] ; Souza de Chagas, Ana Carolina ^[1]

Afiliação do(s) autor(es)

^[1] Embrapa Pecuaria Sudeste, Rodovia Washington Luiz, Km 234, POB 339, BR-13560970 Sao Carlos, SP - Brazil

^[2] Ctr Univ Cent Paulista, Rua Miguel Petroni 5111, BR-13563470 Sao Carlos, SP - Brazil

^[3] Univ Fed Sao Carlos, Rodovia Washington Luiz, Km 235, BR-13566905 Sao Carlos, SP - Brazil

^[4] Embrapa Pecuaria Sul, Rodovia BR-153, Km 632, 9 Vila Ind, BR-96401970 Bage, RS - Brazil

• Tipo de documento: Artigo Científico
• Fonte: MOLECULAR BIOLOGY REPORTS; v. 45, n. 4, p. 651-656, AUG 2018.
• Citações Web of Science: 0

Resumo

Single nucleotide polymorphisms (SNPs) are the main type of variation in genome, enabling them to be associated with traits of economic importance in livestock. Genome-wide association studies (GWAS) have led to the discovery of SNPs associated with desirable traits in sheep. However, in these studies, SNPs are genotyped by high-throughput methods in genome scale, which are expensive and require sophisticated equipment and analysis methods. Therefore, the goal of this study was to develop a reliable, rapid, and inexpensive polymerase chain reaction (PCR)-based method to genotype a medium number of animals for a few candidate SNPs previously associated with desirable phenotypes in sheep by GWAS, using markers associated with gastrointestinal nematode resistance as a model. DNA extracted from white-blood cells of 150 sheep was submitted to PCR amplification followed by agarose gel electrophoresis and determination of banding pattern. Tetra-primer ARMS-PCR was successfully optimized after changes in annealing temperature; annealing and extension times; concentration of MgCl2 and DNA; ratios of inner, outer, forward and reverse primer; and addition of adjuvants, for genotyping the OAR2\_14765360, OAR6\_81718546, OAR11\_62887032, and OAR12\_69606944 SNPs in sheep. An extensive optimization of tetra-primer ARMS-PCR resulted in a suitable, simple, cost-effective PCR-based method of genotyping four SNP markers previously detected by chip arrays. (AU)

• Processo FAPESP: 17/01626-1 - Caracterização genética e de respostas imunes associadas ao fenótipo de resistência parasitária em rebanho ovino da raça Morada Nova
• Beneficiário: Ana Carolina de Souza Chagas
• Modalidade de apoio: Auxílio à Pesquisa - Regular