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All patients had previously been tested for the presence of Y-chromosome microdeletions using the multiplex PCR method of Simoni et al.
Defining regions of the Y-chromosome responsible for male infertility and identification of a fourth AZF region (AZFd) by Y-chromosome microdeletion detection.
Analysis of Y-chromosome microdeletions by polymerase chain reaction (PCR): The genomic DNA from the infertile as well as normal subjects, were isolated by Genomic DNA Isolation Kit (Promega, USA) using the manufacturer's protocol.