A FRT-flanked PGK-Neo-SV40pA selection cassette inserted a strong transcriptional stop signal (3xpA) downstream of a phylogenetically conserved region of the GATA4 bound first heart field specific enhancer. The selection cassette was removed via flp-mediated recombination. This insertion causes a premature termination of transcription and a severely shortened transcript. RNA-seq and qPCR confirmed the absence of full-length RNA in E9.5 hearts. The expression level of its cis located gene Nkx2-5 is unchanged in E9.5 hearts. And the lncRNAs IRENE-SS and IRENE-div expression levels are not affected.