High-throughput screening for spermatogenesis candidate genes in the AZFc region of the Y chromosome by multiplex real time PCR followed by high resolution melting analysis.
Microdeletions in the AZF region of the Y chromosome are among the most frequent genetic causes of male infertility, although the specific role of the genes located in this region is not fully understood. AZFa and AZFb deletions impair spermatogenesis since no spermatozoa are found in the testis. Deletions of the AZFc region, despite being the most frequent in azoospermic patients, do not correlate with spermatogenic failure. Therefore, the aim of this work was to develop a screening method to ascertain the presence of the main spermatogenesis candidate genes located in the AZFc region in the light of the identification of those responsible for spermatogenic failure. DAZ, CDY, BPY2, PRY, GOLGA2LY and CSGP4LY genes were selected on the basis of their location in the AZFc region, testis-only expression, and confirmed or predicted protein codification. AMEL and SRY were used as amplification controls. The identification of Real Time PCR products was performed by High Resolution Melting analysis with SYTO 9 as intercalating dye. The herein described method allows a rapid, simple, low-cost, high-throughput screening for deletions of the main AZFc genes in patients with spermatogenic failure. This provides a strategy that would accelerate the identification of spermatogenesis candidate genes in larger populations of patients with non-obstructive idiopathic azoospermia.
Servicio de Huellas Digitales Genéticas, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
Evguenia
E
Alechine
Servicio de Huellas Digitales Genéticas, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
Daniel
D
Corach
D053713:Azoospermia / Q000235:genetics; D002460:Cell Line; D002872:Chromosome Deletion; D041322:Chromosomes, Human, Y / Q000235:genetics*; D006801:Humans; D007248:Infertility, Male / Q000235:genetics; D008297:Male; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D029589:Seminal Plasma Proteins / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics; D016324:Sequence Tagged Sites; D013091:Spermatogenesis / Q000235:genetics*; D013737:Testis / Q000378:metabolism
D029589:Seminal Plasma Proteins