Sezen ARAT, S.Jacek RZUCIDLO, Steven L. STİCE

Transgenic bovine nuclear transfer embryos from adult somatic cell lines

Bu çalışma olgun hayvandan elde edilen fibroblast hücrelerinin genetik değişikliğe uğratılıp uğratılamıyacağını ve bu hücrelerden geliştirilen hücre hatlarının embriyonik gelişimi destekleyip desteklemiyeceğini araştırmak için yapıldı. Hücre hattı yaşlı bir sığırın kulak dokusundan geliştirildi. Yeşil floresan protein genini (EGFP) taşıyan hücreler ve transfekte edilmemiş olgun hücreler klonlama için kullanıldı. Yeşil floresanın expresyonu olgun hücre klonlannın 35/49 (% 71,4)'ünde tespit edildi. Yeşil floresan proteini exprese eden hücrelerden oluşan embriyoların gelişme oranı (% 11,4) transfekte edilmemiş hücrelerden oluşanlardan (% 20,1, P < 0,05) daha düşüktü. Bununla beraber EGFP ile transfekte edilen ve EGFP'yi exprese etmeyen ancak neomisin dirençlilik geninin taşıyan hücrelerin nükleusları kullanıldığında, nükleer transfer embriyolarının gelişme oranında azalma olmadı (% 21,5). Transfekte edilmiş ve transfekte edilmemiş hücrelerden oluşan embriyolar benzer morfoloji ve hücre sayısına sahipti. Bu sonuçlar olgun hücrelerin transfeksiyon ve seleksiyonu içeren klonal çoğalmayı tamamlayabileceğini ve bu hücrelerin transgenik nükleer transfer embriyolarının üretiminde kullanılabileceğini gösterdi.

Erişkin cücut hücresi hatlarından elde edilen transgenik sığır nükleer transfer embriyoları

This study was performed to examine whether adult fibroblast cells can be manipulated genetically and whether clonal lines derived from those cells can support embryonic development. A primary adult cell line was established from the ear of an aged cow. Adult cells with a plasmid containing the enhancer green fluorescence protein (EGFP) gene, and non-transfected cells were used for cloning. Green fluorescence expression was observed in 35/49 (71.4%) adult clones. The developmental rates of embryos were significantly lower for cell lines expressing EGFP (11.4%) than for non-transfected cells (20.1%, P < 0.05). However, there was no decrease in nuclear transfer (NT) developmental rates (21.5%) when donor nuclei from EGFP transfected cell lines not expressing EGFP but retaining neomycin-resistance gene expression were used as donor nuclei. The NT embryos from transfected and non-transfected cell lines had similar morphology and cell numbers. The results indicated that adult cells can complete clonal propagation including transfection and selection and can be used to produce transgenic NT embryos.

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