Y CHROMOSOMAL STR LOCUS DYS385 IN AZOOSPERMIC AND FERTILE MEN FROM THE AEGEAN REGION OF TURKEY: IS THERE ANY FORENSIC RELEVANCE?
Unuvar D1, Baransel Isir A2, Cankus G3, Pehlivan S3,*
*Corresponding Author: Associate Professor Sacide Pehlivan, Department of Medical Biology and Genetics, Faculty of Medicine, Gaziantep University, Gaziantep, Turkey; Tel./Fax: +90-34- 2360-6028; e-mail: spehlivan@ gantep.edu.tr; aybaransel@yahoo.com
page: 39

MATERIALS AND METHODS

Forty-nine azoospermic males referred to the Family Planning and Infertility Research Treatment Center in the Aegean Region of Turkey and 51 males with proven fertility, were analyzed. Forty-nine azoospermic males, who had non obstructive azoospermia and normal endocrine or hereditary findings, were obtained for the present study [13], together with 51 males with proven fertility, who had had a child in the last 5 years. The study was designed in accordance with the Helsinki Declaration of 1975 on human experimentation and signed informed consent was obtained from all enrollees.

      Blood samples were taken from all cases. Genomic DNA was extracted from 2 mL of peripheral blood samples which were collected in tubes containing EDTA as anticoagulant by using a NucleoSpin (Macherey-Nagel GmbH & Co. KG, Düren, Germany) isolation kit. The primer sequences, as originally described by Gerken et al. (GDB accession nos. L36701 and L36702, unpublished), were used in the PCR. DYS385-1 and DYS385-2B primers were used. These primer sequences are as follows: (DYS385-1) 5'-AGC ATG GGT GAC AGA GCT A-3'; (DYS385-2B) 5'- CCA ATT ACA TAG TCC TCC TTT C-3'.

      The PCR was performed according to the method described by Schneider et al. [9]. The PCR amplification protocol was carried out as follows (using a TC-1 thermo­cycler; Perkin Elmer, Langen, Germany): initial denaturation for 2 minutes at 94C then a touchdown PCR [14] with denaturation for 30 seconds at 94C, annealing for 30 seconds at decreasing temperatures of 59-57C (3X two cycles at each temperature), extension for 1 minute at 72C, followed by 29 cycles as above with at a 56C annealing temperature, and a final extension for 7 minutes at 72C. Electrophoretic separation of amplified PCR products was performed using a non denaturing 7% polyacryl­amide gel followed by silver staining. Genotype frequencies and gene diversity value were calculated according to Nei [15].

 




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