Objective: The aim of this study was to evaluate and compare alterations in gene expression using two distinct immortalization methods (hTERT and HPV16-E6/E7) in ameloblastoma cell lines.
Materials and methods: A primary cell culture derived from human ameloblastoma (AME-1) was established and immortalized by two different methods using a transfection processes to hTERT and HPV-E6/E7. The RNA-seq was used to verify which immortalization method had less influence on gene expression. It was performed in four steps: extraction and collection of mRNA, PCR amplification, comparison with the human reference genome, and analysis of differential expression. The genes with differentiated expression were identified and mapped.
Results: RNA-seq revealed genetic alterations in ameloblastoma cell lines after the immortalization process, including increased expression of tumor genes like MYC, E2F1, BRAF, HRAS, and HTERT, and a decrease in tumor suppressor genes like P53, P21, and Rb.
Conclusions: It is possible to affirm that cell immortalization is not an inert method regarding gene regulation mechanisms and the hTERT method (AME-TERT) presented fewer changes in gene expression levels.
Keywords: RNA-Seq; ameloblastoma; cell immortalization; transcriptome.
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