Effect of Freeze-Drying Extenders on Ram Sperm DNA Integrity

Abstract

DNA hasarının belirlenebilmesi, sperma saklama yöntemlerinin geliştirilmesi açısından önemlidir. Biz çalışmamızda farklı liyofilizasyon medyumları kullanılarak saklanan sperm hücrelerinin DNA bütünlüğünün değerlendirilmesinde akridin oranj (AO) ve therminal deoxynucleotidyl transferase-medited dUDP nick and labelling (TUNEL) florasan boyama yöntemlerini kullandık. Koçlardan alınan sperma dört gruba bölündü ve her bir grup: I) %10 fötal buzağı serumu (FCS) içeren TCM 199 solüsyonu II) %10 FCS ve 0.2 mol/L trehaloz içeren TCM 199 solüsyonu III) 50 mmol/L NaCl ve EGTA [Ethylen glycol-bis (β-aminoethyl ether)-N, N, N, N,-tetraacetic acid] 10 mmol/L Tris solüsyonu ve IV) %20 yumurta sarısı ve %7 gliserol içeren Tris bazlı solüsyonlardan biri ile konsantrasyonu 10 x 106 spermatozoa/100 μL olacak şekilde sulandırıldı. Taze alınan spermada DNA fragmentasyon oranı AO ve TUNEL yöntemlerinde sırasıyla %2 ve %4 olarak tespit edildi (P>0.05). AO ve TUNEL yöntemiyle değerlendirilen liyofilizasyon sonrası DNA bütünlüğüne sahip spermatozoa oranlarının, sulandırıcı farklılığına göre etkilenmediği tespit edildi (P>0.05). Aynı şekilde her iki boyama yöntemi karşılaştırıldığında, liyofilizasyon sonrası elde edilen DNA bütünlüğünün değerlendirilmesinde AO ve TUNEL sonuçlarının istatistiksel olarak farklı olmadığı belirlendi (P>0.05).

Keywords

• koç sperması
• liyofilizasyon
• DNA bütünlüğü
• TUNEL
• AO

References

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