CANLILARDA “TEK HÜCRE JEL ELEKTROFOREZ” YÖNTEMİ İLE DNA HASAR ANALİZİ (TEKNİK NOT): COMET ANALİZ YÖNTEMİ

Year 2010, Volume: 14 Issue: 2, 77 - 89, 21.02.2014

Murat Dikilitaş , Abdurrahim Koçyiğit

Abstract

Comet analiz yöntemi olarak da bilinen tek hücre jel elektroforez yöntemi son yıllarda genişleyen uygulama alanı, güvenilirliği ve uygulaması kolay olması bakımından kimyasal ve
fiziksel etmenlerin canlılar üzerinde yol açtığı genotoksik ve sitotoksik etkilerin bir göstergesi olan DNA hasar seviyelerinin ölçülmesini sağlayan önemli bir metotdur. Bu metot, izole edilen DNA içeren tek hücrelerin DNA’larının lam üzerinde hazırlanan agar jel içerisinde elektroforetik ortamda yürütülmesi ve hasar seviyesine göre göç eden farklı yük ve molekül ağırlıklarına sahip DNA parçalarının DNA spesifik floresan boya ile boyandıktan sonra, floresan mikroskop altında değerlendirilmesi esasına dayanmaktadır. Metot, oksidatif stres, toksik ağır metaller, kimyasal ajanlar, ilaçlar ve ultraviole gibi çeşitli genotoksik ajanların DNA sarmalları üzerinde oluşturduğu tek veya çift zincir kırıklarını doğru, hassas, hızlı, ucuz ve az bir örnek hacmi kullanarak ölçen bir yöntem olup, tüm canlı hücreleri üzerinde yapılan çalışmalarda yaygın olarak kullanılmaktadır.

Keywords

Alkali Tek Hücre Jel Elektroforezi, Comet Analizi, SCGE, DNA Hasarı, DNA Tamiri.

ANALYSIS OF DNA DAMAGE IN ORGANIZMS VIA “SINGLE CELL GEL ELECTROPHORESIS” (TECHNICAL NOTE): COMET ASSAY METHOD

Abstract

Single cell gel electrophoresis known also as the comet assay is an important method which enables to measure DNA damage and the evaluation of mechanisms of genotoxic and cytotoxic
effects of physical and chemical agents on organisms. The method is reliable, easily applicable and has been widely used in recent years. This method depends on the isolated DNA of single cells running in agarose gel on a microscopy slide in an electrophoretic condition and the migration of damaged DNA fragments with different polarity and molecular weights and the evaluation of DNA fragments following the use of DNA specific fluorescence dyes under a microscopy. The assay method determines the single or double-strand breaks caused by the
agents such as oxidative stress, chemical agents, drugs, heavy metal toxicity and ultraviolet lights with fast, sensitive, reliable and cheap way as well as requiring the little amount of cell samples. The method is commonly applied in all living cells. 

Keywords

The Alkaline Single Cell Gel Electrophoresis, Comet Assay, SCGE, DNA Damage, DNA Repair

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