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At least 991 records · Page 55Linked to original sources
PubMed2005

Mutational analysis of the BRCA1 gene in 30 Czech ovarian cancer patients.

Ovarian cancer is one of the most severe of oncological diseases. Inherited mutations in cancer susceptibility genes play a causal role in 5-10% of newly diagnosed tumours. BRCA1 and BRCA2 gene alterations are found in the majority of these cases. The aim of this study was to analyse the BRCA1 gene in the ovarian cancer risk group to characterize the spectrum of its mutations in the Czech Republic. Five overlapping fragments amplified on both genomic DNA and cDNA were used to screen for the whole protein-coding sequence of the BRCA1 gene. These fragments were analysed by the protein truncation test (PTT) and direct sequencing. Three inactivating mutations were identified in the group of 30 Czech ovarian cancer patients: the 5382insC mutation in two unrelated patients and a deletion of exons 21 and 22 in another patient. In addition, we have found an alternatively spliced product lacking exon 5 in two other unrelated patients. The 5382insC is the most frequent alteration of the BRCA1 gene in Central and Eastern Europe. The deletion of exons 21 and 22 affects the BRCT functional domain of the BRCA1 protein. Although large genomic rearragements are known to be relatively frequent in Western European populations, no analyses have been performed in our region yet.

Adult↗
PubMed1998

Mutational analysis of the N-ras gene in acute lymphoblastic leukemia: a study of 125 Japanese pediatric cases.

A point mutation of the N-ras gene is one of the known genetic alterations identified in patients with acute lymphoblastic leukemia (ALL), but its clinical importance is still controversial. Using polymerase chain reactions, we examined codons 12, 13 and 61 of this gene in 125 Japanese childhood ALL patients (64 common-ALL, 22 pre-B-ALL, 33 T-ALL, 2 B-ALL, 3 undifferentiated ALL, and 1 unclassified ALL) including 9 relapsed patients. An N-ras point mutation was observed in 14 (11%) patients (9 common-ALL, 3 T-ALL, and 2 undifferentiated ALL; 13 patients at diagnosis and 1 at relapse). The patients with undifferentiated ALL harbored an N-ras mutation at a significantly higher rate. However, no correlation was found between the presence of an N-ras mutation and sex, age, or white blood count. There was no significant difference in the event-free survival rate between 13 fresh patients with an N-ras mutation and 103 patients with a wild-type configuration. The N-ras mutation was present in about 10% of childhood ALL cases but it did not have a prognostic impact. The sequence analyses revealed that the majority of the patients (13/14) had an N-ras mutation of a G to A transition. This finding was consistent with previous reports on N-ras mutations in acute leukemias in which the incidence of a G to A mutation was significantly higher in ALL than in myeloid malignancies.

Child↗
PubMed2005

[FANCA gene mutation analysis in Fanconi anemia patients].

OBJECTIVE: To screen the FANCA gene mutation and explore the FANCA protein function in Fanconi anemia (FA) patients. METHODS: FANCA protein expression and its interaction with FANCF were analyzed using Western blot and immunoprecipitation in 3 cases of FA-A. Genomic DNA was used for MLPA analysis followed by sequencing. RESULTS: FANCA protein was undetectable and FANCA and FANCF protein interaction was impaired in these 3 cases of FA-A. Each case of FA-A contained biallelic pathogenic mutations in FANCA gene. CONCLUSIONS: No functional FANCA protein was found in these 3 cases of FA-A, and intragenic deletion, frame shift and splice site mutation were the major pathogenic mutations found in FANCA gene.

Cell Line↗
PubMed2000

Genotypic mutation analysis in the p53 gene of benzo[a]pyrene-treated European flounder (Platichthys flesus).

We have applied a genotypic mutation detection system (the Restriction Site Mutation (RSM) assay) to detect mutations in the marine teleost flounder (Platichthys flesus). The aim of this study was to evaluate this species as an environmental indicator of genotoxic exposure. We have used the model genotoxin benzo[a]pyrene (B[a]P) to determine the limits of mutation detection in the p53 gene of flounder liver DNA. This study has revealed two important findings. Firstly, we were able to demonstrate that a polymorphism exists in the TaqI restriction site of exon 8 of the flounder p53 gene at codon 243. This polymorphic allele was present as a heterozygote at a mean frequency of 15%, whereas 85% carried the homozygous wild type sequence. Secondly, we established that B[a]P treatment resulted in specific mutational events at the adenine base of the same TaqI site, contrasting previous reports stating that there was a guanine preference for this chemical in mammalian DNA. This difference in mutation specificity may possibly be accounted for by sequence specific factors or by species differences in metabolic activation and/or DNA repair and are worthy of further study.

Adenine↗
PubMed2003

Mutation analysis in 16 patients with mtDNA depletion.

Sixteen unrelated Southern European patients with the mitochondrial depletion syndrome (MDS) were analyzed for mutations in the TK2 and DGUOK genes. Three novel mutations were identified in TK2 (R183G, R254X, and 142insG). When we analyzed additional genes involved in the dNTPs pool, such as SLC25A19 (DNC) and NT5M (d-NT2), we did not detect mutations. The current study suggest that scanning the TK2, DGUOK, SLC25A19, and NT5M genes is likely to help about 10% of MDS families in terms of genetic counseling. Also, our findings indicate that genotype-phenotype correlations are not straightforward in MDS.

Age of Onset↗
PubMedDate not supplied

Mutational analysis and genotype-phenotype correlation in patients with classic 21-hydroxylase deficiency from transylvania (north-west Romania).

The regional incidence of 21-hydroxylase deficiency, its mutational spectrum and the correlation of genotype and phenotype has been studied by European, American and Latin-American groups. However, little information is known about the molecular background of the disease in patients from Central-Eastern Europe. The present study aimed to genotype a group of patients from Transylvania, the north-western part of Romania, in order to gain some insight into the molecular pattern and the genotype-phenotype correlation of congenital adrenal hyperplasia (CAH) in this region. We genotyped 17 patients with classic 21-hydroxylase deficiency and, whenever available, their parents in order to verify mutational segregation. The patients came from 13 unrelated families. DNA was prepared from peripheral blood leucocytes and four gene fragments were amplified by PCR. The 21-hydroxylase gene (CYP21B) was completely sequenced and analyzed for point mutations or deletions. Percentage distribution of mutations was as follows: 12G--34.6%, deletions and large conversions (del)--19.2%, P30L--15.4%, 1172N--15.4%, P30L+I2G+del8bp (triple mutation)--11.5%, and R356W--3.8%. Mutational percentage distribution compared to other Latin populations is higher for I2G and I172N and lower for deletions, while the P30L mutation was found at a higher rate than in any other analyzed population. Some differences may arise from the low patient number and from ethnic particularities. The incidence of compound heterozygotes in our group was 76.5%. The genotype seemed to correlate fairly well to phenotype, with a general concordance rate of 82.35%. Clear divergence was found in two patients with the simple virilizing form, exhibiting a homozygous status for I2G and del, respectively. This study offers the first information about the molecular pathology of CAH in Romania and should help to improve management and clinical outcome for patients with CAH in Transylvania. Hopefully, it might also be the first step towards exact and accurate prenatal diagnosis in our country.

Adolescent↗
PubMed1999

Mutational analysis of the cardiac actin gene in familial and sporadic dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) results in part from genetic disorders. Recently, missense mutations of the cardiac actin gene have been reported to cause DCM. We studied 136 Japanese DCM cases to elucidate how frequently the gene mutations are involved in its pathogenesis. Genomic DNA samples were obtained from 136 DCM cases (107 males, 29 females), containing 30 familial DCM (5 confirmed and 25 suspected). All six exons of the cardiac actin gene were analyzed by polymerase chain reaction, single-strand conformation polymorphism, and sequencing. We detected no mutations of the disease causation previously reported (G867A or A1014G) but two silent mutations (G979C and C1018T) in exon 6 and one point mutation (T1080A) in the 3'-untranslated region. As a result of screening 128 healthy subjects, these novel silent mutations were found to be mere genetic polymorphisms, not responsible for the disease. Although some genetic polymorphisms exist in the cardiac actin gene, mutations of the gene are rarely responsible for DCM, at least in the Japanese patients.

3' Untranslated Regions↗
PubMed2004

Mutation analysis of hMSH2 and hMLH1 in colorectal cancer patients in India.

The aim of this work was to study the mutation profile in hMSH2 and hMLH1 genes in hereditary nonpolyposis colorectal cancer (HNPCC) patients in India. On the basis of the Bethesda criteria, 31 colorectal cancer patients were studied first for microsatellite instability, using the five markers recommended by the Bethesda guidelines. Twelve of 31 tumor samples were found to be MSI-H, 9 of 31 were MSI-L, and the rest were MSS. The 12 patients with MSI-H were analyzed for mutations in hMSH2 and hMLH1 genes using PCR-denaturing high-performance liquid chromatography (dHPLC), followed by sequencing of samples showing abnormal peaks. Of the five mutations detected, three were found to be deleterious mutations (hMSH2-R680X, hMLH1-E671X, and a splice junction mutation IVS16-2A --> G); one had a mutation of probable significance (hMLH1-C680G) and one was of unknown significance (hMSH2-R171K). This study has also shown that most of the early-onset colon (4/7) and early-onset rectal (15/21) cancers are MSS or MSI-L. This is the first study to describe the mutation in hMSH2 and hMLH1 in Indian patients, a low incidence region for colorectal cancer. A two-stage procedure using MSI testing followed by PCR-dHPLC was found to be an efficient method in studying the mutation profile in high-risk patients.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Mutation analysis of MECP2 and clinical characterization in Korean patients with Rett syndrome.

Rett syndrome is a progressive neurodevelopmental disorder occurring predominantly in females. Recently, mutations in the MECP2 gene on Xq28, which encodes methyl-CpG binding protein 2, were identified as responsible for some cases of Rett syndrome. In the present study, we analyzed the entire coding sequence of the MECP2 gene in 20 sporadic cases of Rett syndrome in Korea. Of the 20 patients, 14 (70%) had pathogenic mutations, which included 10 different mutations. Altogether, there were five missense mutations (D97Y, L100V, R133C, T158M, R306C), four nonsense mutations (R168X, R255X, R270X, R294X), and one frameshift mutation (a 41-bp deletion at 1157-1197). Two of these were novel mutations (D97Y, L100V). Most of the nucleotide substitutions involved C to T transitions at CpG hotspots. We could find no clear phenotype-genotype correlation according to the type of mutation. However, there was a tendency for patients with no MECP2 mutation (30%) to show more severe symptoms and more rapid clinical progression than patients with mutations. Further studies are necessary to identify the other possible genetic causes of Rett syndrome.

Adolescent↗
PubMed1993

The use of selection in recovery of transgenic targets for mutation analysis.

Transgenic animal mutagenesis assays using lambda shuttle vectors have recently been described for isolation and characterization of spontaneous and chemical induced DNA mutations. Extensive information on lambda and E. coli genetics provides a wealth of techniques to allow selection of mutant target genes. Here we describe the modification of an E. coli host which permits two methods for the direct selection of mutant genes. These methods reduce the number of plates needed to be screened for a comparable amount of frequency data by 20-100-fold and thus provide a significant savings of the materials and time required for the screening of mutations. In addition, mutants selected by these approaches described here may alter or broaden the spectrum of mutations that are recoverable. Ultimately, a combination of selective and nonselective techniques may prove valuable for the analysis of mutations produced in vivo in transgenic animals.

Bacteriological Techniques↗
PubMed2002

[Mutation analysis of retinitis pigmentosa 1 gene in Chinese with retinitis pigmentosa].

OBJECTIVE: To investigate the frequency and pattern of RP1 point mutations in Chinese retinitis pigmentosa (RP) patients and to examine their effects on the development of RP. METHODS: Conformation sensitive gel electrophoresis (CSGE) and direct DNA sequencing were used to determine sequence alterations occurring in the entire coding region of the RP1 gene in 101 Chinese RP patients in Hong Kong. RESULTS: R677X was detected in one RP patient. A nonpathogenic nonsense mutation, R1933X, was identified in three normal individuals and one patient with Stargardt disease. The frequency of RP1 mutations among all RP patients in this study is 1/101. R677X is expected to lead to large disruptions of the encoded protein. Additionally, 10 more missense alterations in the RP1 gene were identified in the subjects of this study. Apart from M479I whose pathogenicity can not be determined currently, other sequence changes are just polymorphisms of the RP1 gene. CONCLUSION: The nonpathogenicity of R1933X indicates that the C-terminal 224 residues of RP1 protein may be not critical for RP1. Recently, a C-termnal truncating mutation, Y1053(1 bp del), was reported to occur in an RP patient. Thus RP can be caused by lack of the region of RP1 protein after codon 1052 but before 1933. To confirm such a proposition, a large genotyping study is necessary and is likely to reveal more RP causative mutations and uncover more sequence alterations different from those of other ethnic groups.

Adolescent↗
PubMed2001

Evaluation of denaturing high performance liquid chromatography (DHPLC) for the mutational analysis of the neurofibromatosis type 1 ( NF1) gene.

The identification of mutations in the NF1 gene causing type 1 neurofibromatosis (NF1) has presented a considerable challenge because of the large size of the gene, the lack of significant mutational clustering, the diversity of the underlying pathological lesions and the presence of NF1 pseudogenes. Denaturing high performance liquid chromatography (DHPLC), a high throughput, non-hazardous and largely automated heteroduplex-based technique, is in many ways ideally suited to mutation detection in this condition. DHPLC was therefore optimised for the rapid screening of the 60 exons and splice junctions of the NF1 gene in patients with NF1. The sensitivity of DHPLC was evaluated in a retrospective study of a cohort of 111 unrelated NF1 patients with known germline mutations; 97% of mutations were detected. In a subsequent prospective analysis of 50 unrelated NF1 patients, germline mutations were identified in 34 individuals (68%), 22 of these alterations being novel. This represents the highest rate of mutation detection so far reported for the NF1 gene with a single screening technique and genomic DNA as a target.

Base Sequence↗
PubMed2004

TCIRG1-dependent recessive osteopetrosis: mutation analysis, functional identification of the splicing defects, and in vitro rescue by U1 snRNA.

Human malignant infantile osteopetrosis (arOP) is a genetically heterogeneous autosomal recessive disorder of bone metabolism. The TCIRG1 gene, encoding the a3 subunit of the vacuolar proton pump, which mediates the acidification of the bone/osteoclast interface, is responsible for more than one-half of the arOP patients. We performed genetic analysis of TCIRG1 in 55 arOP patients including 25 new cases and identified nine novel mutations. The two most frequent mutations, c.1674-1G>A (aberrant splicing: r.1674_1884del) and c.2005C>T (protein variation: p.Arg669X), found in 17 and 16 alleles, respectively, constituted 30% of all TCIRG1 abnormalities. They both originated in Northern Europe, p.Arg669X quite recently from West Flanders, Belgium. As substitutions in splicing regulatory sequences represented a large portion (40%; 44 alleles) of the TCIRG1 variations, we developed a functional splicing assay to distinguish between polymorphic variants and disease-causing mutations. Three intronic nucleotide substitutions flanking the splice sites (c.117+4A>T; c.1673+5G>A; and c.504-8G>A) were studied using hybrid minigenes and an abnormal processing of the transcripts was demonstrated in all cases. Cotransfection experiments with complementary U1 snRNAs performed in c.117+4A>T and c.1673+5G>A mutations showed that only in the first case was the defect at the 5' splice site corrected, indicating that mutations near the invariant GT donor sites are mechanistically different. These findings indicate the feasibility of the hybrid minigene approach to detect splicing defects, particularly in patients in whom the RNA is not available. In addition, the present results suggest that modified U1 snRNAs may represent a new therapeutic strategy for arOP patients with a U1 snRNP-dependent splicing defect.

Alternative Splicing↗
PubMed2003

Mutational analysis of TGF-beta type II receptor, Smad2, Smad3, Smad4, Smad6 and Smad7 genes in colorectal cancer.

Transforming growth factor-beta(TGF-beta) is known to play an important role in controlling embryonal development, cell proliferation and homeostasis. The purpose of this study is to elucidate the involvement of the TGF-beta pathway in colorectal carcinogenesis. DNA was extracted from 100 patients with colorectal cancer. Then, all coding regions of the TGF-beta type II receptor (TRII) and the genes for Smad2, Smad3, Smad4, Smad6, and Smad7 were analyzed by PCR-SSCP and direct sequencing. Also, a LOH analysis of 18q21, where the Smad2 and Smad4 genes are located, was performed. We detected 11 cases of frameshift mutation in the TRII gene (11%) and 5 cases of point mutations in the Smad4 gene (5.0%); LOH at 18q21 was detected with 33% frequency. No abnormalities were found in the genes for Smad2, Smad3, Smad6, and Smad7. These results suggest that the abnormalities of TRII and Smad4 play an important role inhibiting TGF-beta signaling in colorectal carcinogenesis.

Base Sequence↗