Padronização da técnica de extração do DNA com pelos de búfalos e análise da diversidade genética em um rebanho de búfalos utilizando marcadores microssatélite (SSRs)

Abstract

In Brazil, the herd of buffaloes is relatively low compared to other crops such as cattle herds. The largest herds of buffalo in Brazil are located in the northern region. Knowing the genetic variability contained in buffalo populations from different regions is essential for establishing a good breeding program. However, few studies using molecular markers in buffalo have been conducted. However, there are few works that make use of different tissues to obtain the DNA, fundamental material to realize this type of study. Thus, the objective of the present study was to perform the standardization of DNA extraction, to later make the analysis of genetic diversity with the microsatellite markers. The microsatellite markers that were used are multi-allelic and polymorphic, being efficient in the detection of molecular polymorphisms. Thirty animals were used to extract the total genomic DNA, and it was necessary to test three different protocols for the standardization of the extraction, being them; protocol with bovine blood, bee wing and 2 animal tissue. Afterwards PCR standardizations were performed to verify the best annealing temperature of the microsatellite primers. For the standardization of extraction the best protocol was the 2 animal tissue. The PCR reactions were performed by touchdown of 55 to 60 ° C, amplified in metaphor agarose gel. The data obtained through the genotyping of three molecular markers of the microsatellite type were analyzed by the Darwin program, separating into two different groups, showing that the animals used are genetically different. The PIC values were 0.5732 for the BL41 locus, 0.4786 for IDVGA 53 and 0.5149 for CSSM054, being shown to be an efficient marker for detecting polymorphism. However, a larger number of microsatellite markers should be used to verify the genetic diversity among the buffaloes evaluated, since the number of markers is still considered low to estimate the genetic variability that may actually exist between the genotypes.