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O K Rødningen

Publications and source records attributed to O K Rødningen.

At least 19 recordsLinked to original sources
PubMed2000

The genetic algorithm applied to haplotype data at the LDL receptor locus.

Conventional statistical methods based upon single restriction fragment length polymorphisms often prove inadequate in studies of genetic variation. Cladistic analysis has been suggested as an alternative, but requires basic assumptions that usually cannot be met. We wanted to test whether it could be a workable approach to apply the genetic algorithm, an artificial intelligence method, to haplotype data. The genetic algorithm creates in-computer artificial 'individuals', all having 'genes' coding for solutions to a problem. The individuals are allowed to compete and 'mate', individuals with genes coding for better solutions mating more often. Genes coding for good solutions survive through generations of the genetic algorithm. At the end of the run, the best solutions can be extracted. We applied the genetic algorithm to data consisting of cholesterol values and haplotypes made up of seven restriction sites at the LDL receptor locus. The persons included were 114 FH (familial hypercholesterolemia) patients and 61 normals. The genetic algorithm found the restriction sites 1 (Sph1 in intron 6), 2 (StuI in exon 8), and 7 (ApaLI site in the 3' flanking region) were associated with high cholesterol levels. As a validity check we used runs of the genetic algorithm applied to 'artificial patients', i.e. artificially generated haplotypes linked to artificially generated cholesterol values. This demonstrated the genetic algorithm consistently found the appropriate haplotype. We conclude that the genetic algorithm may be a useful tool for studying genetic variation.

Algorithms↗
PubMed1999

Mutant transcripts of the LDL receptor gene: mRNA structure and quantity.

mRNA of the low-density lipoprotein receptor (LDLR) gene from 22 heterozygous familial hypercholesterolemic subjects possessing different mutations in this gene was analyzed by Northern blot analysis and reverse transcription-polymerase chain reaction (RT-PCR) in order to detect abnormally spliced transcripts. These analyses revealed abnormally spliced transcripts for the two splice-site mutations 1359-1G-->A and 1705 + 1G-->T. The abnormally spliced transcript for mutation 1359-1G-->A was caused by activation of a cryptic acceptor splice site in exon 10. As a result, seven nucleotides of exon 10 were deleted. For mutation 1705 + 1G-->T, two mutant transcripts were observed. In the first transcript, exon 10 was spliced to exon 13, and in the second transcript intron 11 was retained. The relative amount of mutant transcripts from 14 of the 22 subjects was determined by use of an RT-PCR-based method. Quantitation of the relative amounts of mutant transcripts for five missense mutations resulted in a mean value (+/-SD) of 52.8% (+/-4.55). In comparison, quantitation of the relative amounts of mutant transcripts for five nonsense mutations resulted in a mean value of 31.8% (+/-6.91). This value was significantly lower than the value of 54.2% (+/-2.38) obtained for nine healthy subjects (P < 0.0001). The relative amount of mutant transcripts for the 1705 + 1G-->T mutation was 36%. Thus, transcripts from alleles containing premature stop codons are present in reduced amounts, whereas transcripts from alleles containing missense mutations are present in normal amounts. These findings underscore the importance of determining how mutations affect mRNA structure and quantity in order to understand how mutations cause disease.

Alleles↗
PubMed1999

Phenotypic consequences of a deletion of exons 2 and 3 of the LDL receptor gene.

Screening for structural alterations of the low density lipoprotein (LDL) receptor gene by Southern blot analysis revealed an abnormal band pattern in one subject with a clinical diagnosis of homozygous familial hypercholesterolemia (FH). The molecular defect was further characterized by polymerase chain reaction and cDNA sequencing. These analyses identified a 4.8 kb in-frame deletion of exons 2 and 3, where exon 1 was spliced to exon 4. This deletion is expected to produce a receptor that has lost the two first cysteine-rich repeats of the ligand-binding domain. Previously published data of in vitro site-directed mutagenesis has shown that binding of LDL to such a receptor is reduced to 70% of normal. A mild phenotype in our FH homozygote is consistent with that observation. In contrast, heterozygotes carrying this deletion have a relatively more severe phenotype that is comparable to that of heterozygotes carrying a null-allele. A severe phenotype was also found in a compound heterozygote carrying this deletion. Possible mechanisms for this phenotypic variability are discussed.-Rødningen, O. K., S. Tonstad, J. D. Medh, D. A. Chappell, L. Ose, and T. P. Leren. Phenotypic consequences of a deletion of exons 2 and 3 of the LDL receptor gene.

Base Sequence↗
PubMed1998

Effects of a 9.6-kb deletion of the LDL receptor gene (FH Helsinki) on structure and levels of mRNA.

FH Helsinki is a deletion of the low-density lipoprotein receptor (LDLR) gene that deletes 9.6 kb from intron 15 to exon 18. Screening for mutant transcripts by Northern blot analysis from a patient heterozygous for FH Helsinki revealed two mutant transcripts. One was a transcript where the proximal part of intron 15 was retained in mRNA. The second was a transcript where exon 15 was spliced to nucleotide 4186 of exon 18. Thus, this transcript was generated using the normal donor splice site in intron 15, and a cryptic AG acceptor splice site in exon 18. Translation of the two mutant transcripts is predicted to give nonfunctional proteins, as both the membrane-spanning domain and the cytoplasmic domain of the receptor are deleted. Scanning of the autoradiograms showed that the amounts of each of the two mutant transcripts were approximately 10 times higher than that of the normal transcript in our heterozygous patient. The finding of higher levels of mutant transcripts was confirmed by an allele-specific transcript quantitation method, in which the amount of the two mutant transcripts together was approximately 5 times higher than the amount of the normal transcript. Deletion of destabilizing elements (AU-rich elements) by FH Helsinki are proposed to cause the increased levels of mutant transcripts.

Base Sequence↗
PubMed1997

[Application of gene technology in the diagnosis of familial hypercholesterolemia].

Familial hypercholesterolaemia is an autosomal dominant disorder characterized by hypercholesterolaemia, xanthomas and premature coronary heart disease. Treatment of hypercholesterolemia is effective and consists of dietary changes and lipid lowering drugs. Only a minor proportion of familial hypercholesterolaemia patients are adequately treated, however. One explanation for this is assumed to be the relatively vague clinical diagnostic criteria applied. Because familial hypercholesterolaemia is caused by a mutation in the gene encoding the low density lipoprotein (LDL) receptor, mutation analysis of this gene could form the basis for specific diagnosis. 29 different mutations in the LDL receptor gene have been found to cause familial hypercholesterolaemia among Norwegian patients, and a total of 681 patients from 322 unrelated families have been provided with a molecular genetic diagnosis. We conclude that the use of molecular genetic analysis is feasible, and should be used clinically.

DNA Mutational Analysis↗
PubMed1997

Molecular genetics of familial hypercholesterolaemia in Norway.

OBJECTIVES: To characterize mutations in the low density lipoprotein (LDL) receptor gene causing familial hypercholesterolaemia (FH) amongst Norwegian patients. DESIGN: Molecular genetic analyses of the LDL receptor gene have been performed in patients with a clinical diagnosis of FH. SUBJECTS: A total of 742 probands have been studied. Of these, 476 had a diagnosis of definite FH. The rest had a diagnosis of possible FH. RESULTS: Twenty-three different mutations in the LDL receptor gene as well as the apolipoprotein B-3500 mutation have been found. Six of the mutations in the LDL receptor gene are novel mutations. A molecular genetic diagnosis was achieved in 295 of the probands with definite FH (62%) and in 317 probands total. Of the 317 probands, 3% carried the apolipoprotein B-3500 mutation. When family members were included, a total of 624 persons carried a mutation in the LDL receptor gene and 20 carried the apolipoprotein B-3500 mutation. CONCLUSIONS: Approximately 5% of Norwegian FH patients have been provided with a molecular genetic diagnosis. Our data suggest that molecular diagnosis of FH in Norway is feasible and should be implemented in clinical medicine.

Adult↗
PubMed1996

Application of long polymerase chain reaction in the study of the LDL receptor gene.

In this report we have applied an improved method of the polymerase chain reaction (PCR) in order to detect structural aberrations and point mutations in the low density lipoprotein receptor (LDLR) gene. Except from intron 1, we were able to amplify the entire gene in two fragments of 16.1 and 20.0 kb, respectively. We were also able to detect a 9.6-kb deletion, known as FH Helsinki, as well as a restriction fragment length polymorphism in the 2.7-kb intron 6. We conclude that PCR can almost completely replace Southern blot analysis in molecular genetic studies of the LDLR gene.

Base Sequence↗
PubMed1996

Two novel missense mutations in the LDL receptor gene causing familial hypercholesterolemia.

We have employed analysis of single-strand conformation polymorphisms to identify mutations in the low density lipoprotein receptor gene causing familial hypercholesterolemia. Two familial hypercholesterolemia heterozygotes had abnormal single-strand conformation polymorphism patterns of exons 4 and 8. DNA sequencing revealed that the abnormal pattern of exon 4 was due to heterozygosity (G/T) at nucleotide 502. Nucleotide 502 is the first base of codon 147, and the G->T mutation (D147Y) changes this codon from AspGAC to TyrUAC. The abnormal pattern of exon 8 was due to heterozygosity (A/G) at nucleotide 1097. Nucleotide 1097 is the second base of codon 345, and the A->G mutation (Q345R) changes this codon from GlnCAG to ArgCGG. Based upon screening of 437 unrelated familial hypercholesterolemia heterozygotes, both D147Y and Q345R account for about 0.5% of the mutations causing familial hypercholesterolemia in Norway.

Base Sequence↗
PubMed1995

Two novel point mutations in the EGF precursor homology domain of the LDL receptor gene causing familial hypercholesterolemia.

Familial hypercholesterolemia is caused by mutations in the low density lipoprotein (LDL) receptor gene. Analysis of single-strand conformation polymorphisms of exons 10 and 11 of the LDL receptor gene from familial hypercholesterolemia heterozygotes indicated the presence of two mutations, which were characterized by DNA sequencing. One mutation (delta N466) was a 3-bp deletion in exon 10 that deletes Asn in codon 466. The other (intron 11 +1, G-->T) was a splice donor mutation at position +1 of intron 11.

Amino Acid Sequence↗
PubMed1995

Screening for known mutations in the LDL receptor gene causing familial hypercholesterolemia.

Familial hypercholesterolemia (FH) is caused by defective low density lipoprotein (LDL) receptors and is characterized by hypercholesterolemia and premature coronary heart disease. Two strategies can be used to identify the mutation in the LDL receptor gene underlying FH. One strategy is to search for novel mutations by DNA sequencing with or without prior mutation screening. The other strategy is to screen for known mutations. In this study we employed the latter strategy to screen 75 unrelated, Norwegian FH subjects for 38 known mutations. Three of the 38 mutations were detected in our group of FH subjects. Two subjects had FH-Padova, one had FH-Cincinnati-2 and one had FH-Gujerat. When additional unrelated FH heterozygotes were screened for the three mutations, the gene frequencies were 1.3%, 1.0% and 3.0%, respectively. In addition to identifying known mutations we also detected a novel stop codon in codon 541 (S541X). We conclude that screening for known mutations in the LDL receptor gene should be used as a complementary strategy to screening for novel mutations in order to understand the molecular genetics of FH.

Base Sequence↗
PubMed1995

Identification of the apo B-3500 mutation in the Norwegian population.

Familial defective apolipoprotein B-100 (FDB) is caused by a mutation in codon 3500 of the apo B gene. It is inherited in a co-dominant fashion and is characterized by hypercholesterolaemia. Thus, FDB has similar features to familial hypercholesterolaemia (FH). In order to investigate whether some of the Norwegian subjects diagnosed as having FH actually have FDB, we have screened 208 Norwegian FH heterozygotes for the apo B-3500 mutation. One of the subjects possessed the mutation which was on a haplotype compatible with the mutation-bearing haplotype found in other populations. Although, hypercholesterolaemia segregated with haplotypes both at the apolipoprotein B and low density lipoprotein (LDL) receptor loci in the proband's family, LDL receptor analysis revealed that the proband was not doubly heterozygous for FDB and FH.

Adult↗
PubMed1994

Two founder mutations in the LDL receptor gene in Norwegian familial hypercholesterolemia subjects.

DNA from 20 unrelated familial hypercholesterolemia (FH) subjects were studied by analysis of single-strand conformation polymorphisms (SSCP) for mutations in exon 3 of the low density lipoprotein (LDL) receptor gene. Four different SSCP patterns were observed. The underlying mutations were characterized by DNA sequencing. One pattern represented the wild-type sequence. Another pattern represented a C-->G mutation (FH-Svartor) that changes codon 78 into the amber stop codon. The two other patterns represented heterozygosity and homozygosity, respectively, for a G-->A splice donor mutation (FH-Elverum) in intron 3. Based upon two PCR-based assays, the frequencies of FH-Svartor and FH-Elverum among 267 unrelated FH subjects, were 8% and 25%, respectively. FH Svartor was located on a chromosome with haplotype 3 in all five families where haplotype analysis were performed. FH Elverum was located on haplotype 2 in 16 out of 20 families. The two mutations must be considered founder mutations in the Norwegian population, and their existence will be clinically useful in diagnosing FH. The presence of two founder mutations together with previously published data on the prevalence of FH in Norway, indicate that FH may be a more common disease in Norway than previously thought.

Base Sequence↗
PubMed1994

Familial hypercholesterolaemia caused by a non-sense mutation in codon 329 of the LDL receptor gene.

Analysis of single-strand conformation polymorphisms (SSCP) was employed to screen familial hypercholesterolaemia (FH) subjects for point mutations in exon 7 of the low density lipoprotein receptor (LDLR) gene. An abnormal band pattern was found in one out of 100 unrelated FH subjects. The underlying mutation was found by DNA sequencing to be due to heterozygosity (C/T) at nucleotide 1048. Nucleotide 1048 is the first nucleotide of codon 329, and is located within the domain that has a high degree of homology with the precursor for epidermal growth factor. The C-->T transition, referred to as FH-Fossum, changes codon 329 from CGAArg to TGAStop. The mutation is expected to cause a class 1 receptor defect.

Codon↗
PubMed1993

Screening for point mutations in exon 10 of the low density lipoprotein receptor gene by analysis of single-strand conformation polymorphisms: detection of a nonsense mutation-FH469-->Stop.

DNA from 40 unrelated familial hypercholesterolemia (FH) heterozygotes were subjected to analyses of single-strand conformation polymorphisms (SSCPs) of exon 10 of the low density lipoprotein receptor (LDLR) gene. Four different SSCP patterns were observed. The underlying mutations were characterized by DNA sequencing. Three of the patterns represented the three genotypes of a recently described sense mutation in codon 450. A method based upon the polymerase chain reaction (PCR) was developed to analyze this mutation. The frequencies of the wild-type (G at nucleotide 1413) and mutant (A at nucleotide 1413) alleles were 0.56 and 0.44, respectively. The fourth pattern was found in only one FH heterozygote and was caused by heterozygosity at nucleotide 1469 (G/A). Nucleotide 1469 is the second base of codon 469Trp(TGG). The G-->A mutation changes this codon into the amber stop codon, and is referred to as FH469-->Stop. The mutant receptor consists of the amino terminal 468 amino acids. Because the truncated receptor has lost the membrane-spanning domain, it will not be anchored in the cell membrane. FH469-->Stop destroys an AvaII restriction site, and this characteristic was used to develop a PCR method to establish its frequency in Norwegian FH subjects. Two out of 204 (1%) unrelated FH heterozygotes possessed the mutation.

Adult↗
PubMed1993

Evaluation of running conditions for SSCP analysis: application of SSCP for detection of point mutations in the LDL receptor gene.

We have performed analyses of single-strand conformation polymorphisms (SSCP) of the promoter region and the translated parts of the 18 exons of the low-density lipoprotein receptor (LDLR) gene. DNA from 20 unrelated familial hypercholesterolemia (FH) patients was studied. Four different running conditions were used for the nondenaturing gel electrophoresis to systematically evaluate how differences in the running conditions affect the sensitivity of the assay. These conditions were 15 W, 40 W, and 50 W in the absence of glycerol, and 50 W in the presence of 10% glycerol. SSCP analyses of the 18 PCR fragments for the 20 subjects revealed a total of 46 genotypes at 15 W, 45 at 50 W, 42 at 40 W, and 41 at 50 W with 10% glycerol. A total of 53 different genotypes were observed when the results of the four conditions were considered together. Assuming that the four conditions together detected 100% of the different genotypes, the sensitivity of the four individual conditions ranged between 87% (15 W) and 77% (50 W with 10% glycerol). There were marked differences among the different running conditions to detect abnormal SSCP patterns of individual exons. Therefore, different conditions should be used for the different exons of the LDLR gene.

Base Sequence↗
PubMed1993

Haplotype analysis at the low density lipoprotein receptor locus in normal and familial hypercholesterolemia Norwegian subjects.

We have performed haplotype analysis at the low density lipoprotein receptor (LDLR) locus in order to investigate the molecular genetics of familial hypercholesterolemia (FH) in Norway. Haplotypes were constructed using 7 restriction fragment length polymorphisms (RFLPs) in 194 subjects from 48 unrelated Norwegian FH families. Hypercholesterolemia co-segregated with haplotypes at the LDLR locus in all 48 families. Unambiguous haplotypes could be established for 190 independent chromosomes from 51 FH heterozygotes and 44 healthy normal subjects. A total of 20 different haplotypes was found. The most frequent haplotype was haplotype 3, which accounted for 32.4% or 43.1% of the normal and defective haplotypes, respectively. Haplotype 2 was significantly more frequent among the defective alleles than among the normal alleles (33.3% and 5.8%, respectively, p < 0.0001). Thus, haplotypes 2 and 3 accounted for 76.4% of the defective haplotypes. More data are needed to determine the possible existence of founder genes in the Norwegian population. Haplotypes 1, 2, 3, 5 and 8 accounted for 88.2% of the normal haplotypes. Based upon the cumulative heterozygosity index, the SphI, NcoI and 3' ApaLI RFLPs are the most informative markers in the Norwegian population.

Base Sequence↗