Kết hợp Omics và Công nghệ CRISPR để Xác định và Xác minh Các Vùng An Toàn Gen Trong Bộ Gen Gà

Springer Science and Business Media LLC - Tập 25 - Trang 1-20 - 2023
Nima Dehdilani1, Lena Goshayeshi1,2, Sara Yousefi Taemeh1,2, Ahmad Reza Bahrami3,4, Sylvie Rival Gervier5, Bertrand Pain5, Hesam Dehghani1,2,6
1Stem Cell Biology and Regenerative Medicine Research Group, Research Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran
2Division of Biotechnology, Faculty of Veterinary Medicine, Ferdowsi University of Mashhad, Mashhad, Iran
3Department of Biology, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran
4Industrial Biotechnology Research Group, Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran
5Stem Cell and Brain Research Institute, University of Lyon, Université Lyon 1, INSERM, INRAE, U1208, USC1361, Bron, France
6Department of Basic Sciences, Faculty of Veterinary Medicine, Ferdowsi University of Mashhad, Mashhad, Iran

Tóm tắt

Một trong những câu hỏi nổi bật nhất trong lĩnh vực chuyển gen là ‘Nơi nào trong bộ gen để tích hợp một gen chuyển giao?’. Việc thoát khỏi sự im lặng epigenetic và sự đóng cửa của trình điều khiển gen chuyển giao cần những vị trí an toàn trong bộ gen (GSH) đáng tin cậy. Những tiến bộ trong công nghệ kỹ thuật gen kết hợp với dữ liệu sinh học đa omics đã cho phép đánh giá hợp lý các vị trí GSH trong bộ gen của vật chủ. Hiện tại, chưa có vị trí GSH nào được xác thực đã được đánh giá trong bộ gen gà. Tại đây, chúng tôi đã phân tích và thực nghiệm hai vị trí GSH trong bộ gen của tế bào gà. Để thực hiện điều này, các vị trí GSH khả nghi bao gồm giống HIPP của gà (cHIPP; giữa các gen DRG1 và EIF4ENIF1) và giống ROSA của gà (cROSA; trên các gen THUMPD3) đã được dự đoán bằng cách sử dụng dữ liệu sinh học đa omics. Sau đó, sự biểu hiện bền vững của gen chuyển giao đã được xác nhận thông qua việc đặc trưng thực nghiệm các dòng tế bào đồng sinh liên tục mang cassette DsRed2-ΔCMV-EGFP tại các vị trí dự đoán. Hình thức suy yếu của trình điều khiển CMV (ΔCMV) cho phép đánh giá chính xác các vị trí GSH theo cách phụ thuộc vị trí so với trình điều khiển CMV đầy đủ. Các vị trí cHIPP và cROSA được giới thiệu trong nghiên cứu này có thể được khai thác một cách đáng tin cậy cho việc sản xuất sinh học nhất quán các protein tái tổ hợp trong những con gà được kỹ thuật gen. Ngoài ra, kết quả còn cho thấy rằng bối cảnh gen quyết định sự biểu hiện của gen chuyển giao được kiểm soát bởi ΔCMV tại các vị trí GSH.

Từ khóa

#Vị trí an toàn gen #chuyển gen #công nghệ CRISPR #kỹ thuật gen #sinh học đa omics #gà

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