Tatiana Puga Torres, Xavier Blum Rojas, Medardo Blum Narváez, Edith López Montanero, Alexandra Narváez Sarasti
JBRA Assist. Reprod. 2015; 19 (2):37-43
Received January 25, 2015
Accepted March 30, 2015
Abstract
Objetive: Y chromosome microdeletions at the "Azoospermia Factor" regions (AZFa, AZFb, AZFc) are the second genetic cause of spermatogenic failure in infertile men. Despite its importance for the infertile patient treatment, any investigation about it has been published previously in Ecuador. The purpose of this study is to optimize a molecular technique that allows detection of microdeletions in the AZF region.
Methods: Using a genomic DNA of healthy male with natural conceived offspring’s, a multiplex real time polymerase chain reaction (qPCR) was standarized with eigth sequence-tagged site (STS) sY85, G34990, sY133, sY127, sY254, sY255, and using as internal control sex-determine region Y (SRY) and Ameologenin Y (AMELY). With this technique, 35 DNA samples taken from peripheral blood of patients with severe oligozoospermia were analyzed.
Results: A triplex qPCR was standardized using EvaGreen DNA-binding dye to obtain melting temperature (Tm) of the STS previously mentioned. Three of the patients evaluated were detected to have partial microdeletion in the AZFa region, with a frequency of 8.8%; being losses in the G34990 section (one patient) and sY85 section (two patients). No cases of microdeletions in other AZF regions were found.
Conclusions: The triplex qPCR optimizated allows the identification of microdeletions in AZFa, AZFb and AZFc region in infertile men and a better clinical management of the patient's treatment decision. This first report for Ecuador reveled a higuer prevalence of microdeletion in the AZFa region in comparison with those previously described in other populations.