[Another hot point mutation of Wilson disease gene in Chinese: exon 12].
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To study the feature of disease-causing mutation of exon 12 of Wilson disease (WD) gene in Chinese and evaluate its value in direct gene diagnosis.Genomic DNA of 44 unrelated WD patients and 60 normal controls was prepared from peripheral blood leukocytes by a salt-out method. The mutation of exon 12 in these subjects was identified by PCR-single strand conformation polymorphism (SSCP) and further confirmed by direct sequencing.Two missense mutations were identified in 8 cases (18%), including 6 cases of heterozygous for Thr935Met mutation and 2 of heterozygous for Gly943Asp mutation. Different features of mutation of exon 12 were noted between Chinese and Caucasian. Both Thr935Met and Gly943Asp were novel missense mutations and exon 12 was another hot point mutation of WD in Chinese besides exon 8.The finding helps establish a fast and effective direct gene diagnosis.Keywords:
Single-strand conformation polymorphism
genomic DNA
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Carbohydrate-deficient glycoprotein syndrome type IA (CDG IA) is an autosomal recessive disease characterized clinically by severe involvement of the central and peripheral nervous system, and biochemically by complex defects in carbohydrate residues in a number of serum glycoproteins. CDG IA is caused by mutations in the PMM2 gene located in chromosome region 16p13. In this study, 61 CDG type IA patients (122 chromosomes) were screened for mutations in the PMM2 gene using a combination of SSCP and sequence analysis. More than 95% of the mutations could be detected. All of them were missense mutations. Mutations 422G>A and 357C>A were strikingly more common in the material and comprised 58% of mutations detected. Of the 20 mutations found, 10 were not reported previously. Seven mutations, e.g. 26G>A (five alleles) and 548T>C (seven alleles), were found only in Scandinavian families. The most common genotype was 357C>A/422G>A (36%). Three patients were homozygous, 357C>A/357C>A (two cases), and 548T>C/548T>C (one case). No patients homozygous for the most common mutation 422G>A were detected. The different mutations were clustered e.g., in that most were located in exon 5 (five) and exon 8 (six), while no mutation was detected in exon 2. When the frequencies of each mutation were included, exon 5 comprised 61% (65 chromosomes) of the mutations; in Scandinavian patients the frequency of these mutations was 72%. Thus, analysis of exon five in these patients enables both reliable and time-saving first screening in prenatal diagnostic cases. This could be followed by a second step of additional strategies for the detection of other mutations. Hum Mutat 16:395–400, 2000. © 2000 Wiley-Liss, Inc.
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To investigate the distribution of possible novel mutation of parkin gene in variant subset patients with Parkinson's disease (PD) in China and to explore the role of parkin gene in the pathogenesis of PD.Seventy patients were divided into early onset and late-onset groups, and 70 healthy subjects were included as controls. Genomic DNA from 70 normal controls and from those of PD patients were extracted from peripheral blood leukocytes with standard procedures. Mutations of parkin gene (exons 1-12) in all subjects mentioned above were screened by PCR-SSCP. And in the samples with abnormal SSCP result, further sequencing was performed to confirm the mutation and its location.A new missense mutation (Gly284Arg) on exon 7 was found in a sample, while 4 samples from all subjects exhibited abnormal mobility shift on SSCP electrophoresis. All mentioned DNA variants were sourced from the samples of the patients with early-onset PD.Point mutation in parkin gene also contributes partly to the development of early-onset Parkinson's disease in Chinese population.
Single-strand conformation polymorphism
Pathogenesis
genomic DNA
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Coding region
Single-strand conformation polymorphism
Stop codon
genomic DNA
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To investigate the mutation characteristics of spastin gene in Chinese patients with hereditary spastic paraplegia (HSP) and thus provide a basis for the gene diagnosis of HSP.Mutation of spastin gene was screened by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) combined with DNA direct sequencing in 31 unrelated affected HSP individuals in China, of whom 22 were from autosomal dominant families and 9 were sporadic HSP patients. Co-segregation analysis was carried out after the finding of abnormal SSCP bands.Six cases were found to have abnormal SCP bands, and among them, two missense mutations (T1258A, A1293G in exon 8) and one deletion mutation (1667delACT or 1668delCTA or 1669delTAC in exon 14) were found and all of them were not reported previously. They were all co-segregated with the disease and were localized within the functional domain of spastin gene. Besides, T1258A was seen in two unrelated families.The mutation rate (18.2%) in autosomal dominant HSP in Chinese patients is comparatively low. Point mutation is the major mutation type and exon 8 may be the mutation hot spot.
Hereditary Spastic Paraplegia
Single-strand conformation polymorphism
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Objective To investigate the characteristics of mutations in exon 3-20 of Wilson disease (WD) gene and their consequences in Chinese population Methods Sixty unrelated normal Chinese and forty four unrelated WD patients were studied Genomic DNA was prepared from peripheral blood leukocytes by a salt out method Polymerase chain reaction single strand conformation polymorphism (PCR SSCP) and subsequently direct sequencing were used to identify the mutations and polymorphisms of WD gene Results Ten different mutations have been found, accounting for 52% of the mutant genes Five of them are identified as novel missense mutations Mutations Arg778Leu, Thr935Met and Ala874Val were represented respectively in 28 4%, 6 8% and 3 4% of WD chromosomes The remaining mutations were found rare and limited to one or two patients A total of 11 patients were homozygous for a single mutation, and 17 patients were in a compound heterozygous state with or without a known mutation Conclusion In Chinese, WD seems to result from two or three relatively common mutations and a large number of rare mutations Arg778Leu and Thr935Met might be hotspots of mutation in Chinese population The results indicated that the feature of mutations of WD gene is different between Chinese and the Western Instead of exon 14 and exon 18, we had to select exon 8 and exon 12 first to detect mutations of WD gene in Chinese It is of great importance to establish a direct diagnostic method for WD This study improves our knowledge on functional domains of the WD gene, and helps elucidate the wide spectrum of manifestations of the disease as well
Single-strand conformation polymorphism
genomic DNA
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To investigate the characteristics of mutations in exon 3-20 of Wilson disease (WD) gene and their consequences in Chinese population.Sixty unrelated normal Chinese and forty-four unrelated WD patients were studied. Genomic DNA was prepared from peripheral blood leukocytes by a salt-out method. Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and subsequently direct sequencing were used to identify the mutations and polymorphisms of WD gene.Ten different mutations have been found, accounting for 52% of the mutant genes. Five of them are identified as novel missense mutations. Mutations Arg778Leu, Thr935Met and Ala874Val were represented respectively in 28.4%, 6.8% and 3.4% of WD chromosomes. The remaining mutations were found rare and limited to one or two patients. A total of 11 patients were homozygous for a single mutation, and 17 patients were in a compound heterozygous state with or without a known mutation.In Chinese, WD seems to result from two or three relatively common mutations and a large number of rare mutations. Arg778Leu and Thr935Met might be hotspots of mutation in Chinese population. The results indicated that the feature of mutations of WD gene is different between Chinese and the Western. Instead of exon 14 and exon 18, we had to select exon 8 and exon 12 first to detect mutations of WD gene in Chinese. It is of great importance to establish a direct diagnostic method for WD. This study improves our knowledge on functional domains of the WD gene, and helps elucidate the wide spectrum of manifestations of the disease as well.
Single-strand conformation polymorphism
genomic DNA
Compound heterozygosity
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This study is to explore whether there is presenilin 1 (PS1) gene mutation in Chinese familial Alzheimer's disease (FAD). There has been no such systemic research before in China. Using polymerase chain reaction, single strand conformation polymorphism (PCR-SSCP), followed by denaturing high performance liquid chromatograph (DHPLC) and DNA sequencing, we analyzed a Chinese family with early onset AD. The patients in this family showed a novel missense mutation in exon 4 of the PS1 gene (G to T change in codon 97), altering valine to leucine acid substitution. Because the change occurred in conserved domains of this gene, and is not present in normal controls, this novel mutation is likely to be causative of Chinese FAD.
Single-strand conformation polymorphism
genomic DNA
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To detect genetic mutations associated with autosomal dominant congenital stationary night blindness (ADCSNB) in a family from Henan province.Genomic DNA was extracted from peripheral blood samples of 14 family members. Based on 3 genes reported previously, PCR primers were designed and corresponding exons containing the mutation sites were amplified with PCR. PCR products were purified and directly sequenced.A c.281C>T heterozygous missense mutation was detected in RHO gene in all of the patients. This mutation can cause a change of the protein structure (p.Thr94Ile). The same mutation was not detected in normal individuals from the family and 50 normal controls.A c.281C>T mutation in RHO gene is responsible for the onset of ADCSNB in this Chinese family and results in symptoms of night blindness.
genomic DNA
Mutation Testing
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To study the feature of disease-causing mutation of exon 12 of Wilson disease (WD) gene in Chinese and evaluate its value in direct gene diagnosis.Genomic DNA of 44 unrelated WD patients and 60 normal controls was prepared from peripheral blood leukocytes by a salt-out method. The mutation of exon 12 in these subjects was identified by PCR-single strand conformation polymorphism (SSCP) and further confirmed by direct sequencing.Two missense mutations were identified in 8 cases (18%), including 6 cases of heterozygous for Thr935Met mutation and 2 of heterozygous for Gly943Asp mutation. Different features of mutation of exon 12 were noted between Chinese and Caucasian. Both Thr935Met and Gly943Asp were novel missense mutations and exon 12 was another hot point mutation of WD in Chinese besides exon 8.The finding helps establish a fast and effective direct gene diagnosis.
Single-strand conformation polymorphism
genomic DNA
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OBJECTIVE: To investigate the characteristics of mutations of exons 14 and 18 of Wilson's disease (WD) gene in Chinese patients. METHODS: The subjects of study included 60 unrelated normal controls and 44 unrelated WD patients. Genomic DNA was prepared from peripheral blood leukocytes by a salt-out method. Mutations of exons 14 and 18 in these subjects were screened by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and further confirmed by sequencing. RESULTS: One patient was homozygous for Arg1041Pro mutation of exon 14 and another patient was heterozygous for Asn1270Ser mutation of exon 18. CONCLUSION: The mutations of exons 14 and 18 of WD gene in Chinese patients were confirmed by sequencing for the first time in China. Arg1041Pro was identified as a novel missense mutation. In addition, an Asn1270Ser, previously described mutation, was detected in this study. But exons 14 and 18 are not the hot point mutations of WD gene in Chinese patients.
Single-strand conformation polymorphism
genomic DNA
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