BACKGROUND: Our understanding of fragile X syndrome can be improved by reversing the expression of the silenced fragile X mental retardation 1 (FMR1) gene in immortalized cells from these patients. Epstein-Barr virus (EBV) infection has been extensively used to transform B cells into a permanent lymphoblastoid cell line. METHODS: We immortalized B lymphocytes from three different fragile X patients and one normal male. We analyzed the CGG triplet repeats and methylation status of the FMR1 and interferon (IFN)-gamma promoter. We also assayed FMR1 mRNA levels by real-time PCR and FMR1 protein (FMRP) by Western blot. RESULTS: We observed that EBV transformation may induce the instability of CGG repeats and DNA demethylation that can lead to the modification of mRNA expression. CONCLUSIONS: EBV transformation may induce variable changes in the genome that can lead to the misinterpretations of experimental data obtained from these cells. Thus, periodic testing of DNA from immortalized cells should be routinely undertaken to detect undesired effects.
BACKGROUND: Our understanding of fragile X syndrome can be improved by reversing the expression of the silenced fragile X mental retardation 1 (FMR1) gene in immortalized cells from these patients. Epstein-Barr virus (EBV) infection has been extensively used to transform B cells into a permanent lymphoblastoid cell line. METHODS: We immortalized B lymphocytes from three different fragile Xpatients and one normal male. We analyzed the CGG triplet repeats and methylation status of the FMR1 and interferon (IFN)-gamma promoter. We also assayed FMR1 mRNA levels by real-time PCR and FMR1 protein (FMRP) by Western blot. RESULTS: We observed that EBV transformation may induce the instability of CGG repeats and DNA demethylation that can lead to the modification of mRNA expression. CONCLUSIONS:EBV transformation may induce variable changes in the genome that can lead to the misinterpretations of experimental data obtained from these cells. Thus, periodic testing of DNA from immortalized cells should be routinely undertaken to detect undesired effects.
Authors: Roberta Pietrobono; Maria Grazia Pomponi; Elisabetta Tabolacci; Ben Oostra; Pietro Chiurazzi; Giovanni Neri Journal: Nucleic Acids Res Date: 2002-07-15 Impact factor: 16.971
Authors: Raquel Martínez; Victoria Bonilla-Henao; Antonio Jiménez; Miguel Lucas; Carmen Vega; Inmaculada Ramos; Francisco Sobrino; Elizabeth Pintado Journal: Mol Diagn Date: 2005
Authors: Malcolm Hawkins; Jennifer Boyle; Kathleen E Wright; Rob Elles; Simon C Ramsden; Anna O'Grady; Michael Sweeney; David E Barton; Trent Burgess; Melanie Moore; Chris Burns; Glyn Stacey; Elaine Gray; Paul Metcalfe; J Ross Hawkins Journal: Eur J Hum Genet Date: 2010-08-25 Impact factor: 4.246
Authors: James F Meschia; Brett M Kissela; Thomas G Brott; Robert D Brown; Bradford B Worrall; Jeanne Beck; Alexa N Skarp Journal: Clin Med Res Date: 2006-03