| Summary: | An analysis of LDL-receptor gene was performed on an Italian patient with heterozygous familial hypercholesterolemia. Restriction enzyme analysis showed that the proband was heterozygous for a deletion of 4.5 kb spanning the 5' end of exon 13 (45 nucleotide residues) to intron 15. Amplification of genomic DNA, using polymerase chain reaction (PCR), followed by direct sequencing, showed that this deletion was identical to the one reported by Lehrman et al. (1986. Proc. Natl. Acad. Sci. 83: 3679-3683). As only the normal LDL-receptor mRNA was detectable in proband fibroblasts by Northern blot, we used reverse transcription-PCR to amplify the mutant mRNA using primers complementary to exon 6 (sense) and exon 18 (antisense). The amplification of control cDNA resulted in a single fragment of 1725 nucleotides containing the normal sequence. The amplification of cDNA from the proband produced the 1725-nucleotide fragment (as in the control) and three additional fragments (F1, F2, and F3) of smaller size. The direct sequence showed that in the F1 fragment exon 12 was joined to exon 16; in the F2 fragment exon 12 was joined to exon 17; and in the F3 fragment exon 11 was joined to exon 16. Thus, the deletion-bearing allele generated three mRNAs, two of which resulted from alternative splicings leading to the skipping of exons 16 and 12, respectively. It is expected that the translation of these mutant mRNAs will generate three aberrant proteins, the synthesis of which should be negligible in view of the very low content of the corresponding mRNAs.
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