کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1929504 | 1050460 | 2012 | 4 صفحه PDF | دانلود رایگان |
Integration of exogenous sequence into an intact genome may cause some artificial phenotype or unspecific observations. We noticed that there is unspecific vascular expression when using fli1a:EGFP transgenic embryos for whole-mount in situ hybridization (WISH) experiments. We therefore tested whether the residual vector sequence contained in the fli1a:EGFP transgene or the integration of transgene into the genome may cause this expression ‘noise’ and/or deregulation of gene expression at a genome-wide level. RNA probes were synthesized using two different methods, i.e. vector-based and PCR-based. The vector-based dnmt3 probe showed unspecific vascular expression in fli1a:EGFP embryos, but not in wildtype embryos, by WISH. Moreover, we also found that compared to that in wildtype, there were alterations in gene expression at whole-genome level in the fli1a:EGFP embryos. Our finding that the vector sequence contained in the fli1a:EGFP genome causes unspecific vascular expression by WISH and the genome-wide expression profiling is altered in fli1a:EGFP embryos strongly argue that extra caution should be taken for data interpretation when using transgenics, such as fli1a:EGFP, in developmental biology studies.
► RNA probe synthesized using regular plasmid method gave a vascular expression in fli1a:EGFP embryos.
► The 240-bp sequence within the fli1a:EGFP transgene is the cause of the unspecific vessel expression.
► Expression profiling was disrupted in fli1a:EGFP embryos by microarray analysis.
Journal: Biochemical and Biophysical Research Communications - Volume 427, Issue 1, 12 October 2012, Pages 223–226