Nucleic Acids Research, Vol 24, Issue 20 4023-4028, Copyright © 1996 by Oxford University Press
RL Davisson, N Nuutinen, ST Coleman and CD Sigmund
We generated transgenic mice containing a chimeric construct consisting of
the alpha-cardiac myosin heavy chain (alpha cMHC) promoter and the human
renin (hRen) gene in order to target hRen synthesis specifically to the
heart. The construct consisted of three segments: (i) an alpha cMHC DNA
segment including 4.5 kb of 5' flanking DNA and an additional 1.1 kb of
genomic DNA encompassing exons I-III (non-coding) and the first two
introns; (ii) a partial hRen cDNA consisting of exons I-VI; and (iii) a
hRen genomic segment containing exons VII through IX, their intervening
introns, and 400 bp of 3' flanking DNA. This results in the formation of a
909 bp internal fusion exon consisting of alpha cMHC, polylinker, and hRen
sequences. Despite the presence of splice acceptor and donor sites
bracketing this exon, transcription of this transgene resulted in a major
alternatively spliced mRNA lacking the exon and therefore a majority of the
hRen coding sequence. Cloning and sequencing of RT-PCR products from
several heart samples from two independent transgenic lines confirmed
accurate and faithful splicing of alpha cMHC exon II to hRen exon VII thus
bypassing the internal fusion exon. All other exons (alpha cMHC exons I and
II and hRen exons VII, VIII and IX) were appropriately spliced. These
results are consistent with the hypothesis on exon definition which states
that internal exons have a size limitation. Moreover, the results
demonstrate that transgenes present in the genome at independent insertion
sites and in either a single copy or multiple copies can be subject to exon
skipping. The implications for transgene design will be discussed.
ARTICLES
Inappropriate splicing of a chimeric gene containing a large internal exon results in exon skipping in transgenic mice
Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242, USA.
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