A rapid and simple method for the isolation of mutant variants regulating tissue-specific expression of the tni gene through drug selection |
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Authors: | Youngwon Lee Charles P Emerson Myoung Hee Kim |
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Institution: | 1. Genetic Resources Center, Korea Research Institute of Bioscience and Biotechnology, KIST, Taedok Science Town, 305-600, Taejon, Korea 2. Fox Chase Cancer Center, 7701 Burholme Avenue, 19111, Philadelphia, PA
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Abstract: | TnINEO fusion gene was constructed by fusing 3.4-kbp of quailTnI genomic DNA sequences spanning the promoter to exon 5 and aneo gene in frame. A myoblast cell line was established after transfection of pTnINEO. Since this cell line was passaged several
times, a high frequency of neomycin (G418) sensitivity conversion was detected. Two drug-resistant variants were analyzed
through genomic Southern blot and S1 nuclease protection assay. One variant has a mutation(s) in the regulatory element that
activated the dormantTnI promoter-enhancer in myoblast, and the other has shown the genomic rearrangement. This result presented the possibility of
isolating factor(s) that activate the muscle-specificTnI promoter simply by screening drug-resistant cells having appropriate mutations. |
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Keywords: | Index Entries" target="_blank">Index Entries Fast troponin I gene stage-specific expression myoblast drug selection mutant genomic rearrangement |
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