Identification of 12 novel mutations in the CFTR gene

Over 200 mutations, besides the deletion     

Spectrum of CFTR mutations in the middle north of Spain and identification of a novel mutation (1341G→A)

We have analyzed 39 unrelated cystic fibrosis (CF) families by denaturing gradient gel electrophoresis (DGGE) and direct sequencing in order to determine the spectrum of CF mutations in our population. This approach has allowed us to detect 72 out of the 78 CF chromosomes (92.3%). The DF508 mutation was found to be present in 51/78 (65.4%) CF chromosomes, in accordance with the predicted Northwest‐Southeast gradient within the European population. Another 14 known mutations, and the novel 1341G→A mutation were identified. Nine out of fifteen non DF508 mutations were present in a single chromosome. The 1341G→A mutation, found in 2 unrelated patients, is a new mutation associated to severe phenotype, causing pancreatic insufficiency and chronic lung infections. Our data suggest a different distribution of non‐DF508 mutations in our population when compared with previous studies carried out in Spanish CF families. Six out of the 14 non‐F508 in our study were not present in a recent study carried out in 640 Spanish families with CF. These six mutations account for 29.6% non DF508 chromosomes in our sample. Hum Mutat 14:89, 1999. © 1999 Wiley‐Liss, Inc.     

Analysis of the CFTR gene in the Spanish population: SSCP-screening for 60 known mutations and identification of four new mutations (Q30X,A120T, 1812-1 G→A,and 3667de14)

In order to determine the spectrum of CF mutations in the Spanish population, we have analysed 40 unrelated Spanish CF patients, with at least one chromosome negative for mutations ΔF508, G542X, and N1303K. Exons l–7,10–14a,15,16,17b,18–21 of the CFTR gene were studied by Single Strand Conformation Polymorphism (SSCP) analysis, using 60 known CF mutations as controls. SSCP screening allowed us to detect 28 different mutations in 52 CF chromosomes, and to identify four new mutations (Q30X in exon 2, A120T in exon 4, 1812-lG→A in intron 11 and 3667del4 in exon 19). Further analysis of the four new mutations in a total of 950 Spanish CF chromosomes showed a final frequency of 0.4%, 0.1%, 0.1%, and 0.1% for 1812-1G→A, Q30X, A120T, and 3667del4, respectively. No mutations were detected in exons 1, 3, 14a, 16, and 18. We have also detected 10 intragenic polymorphisms and DNA sequence variants and have analysed their frequencies in our population. The total of 28 mutations identified in the 80 CF chromosomes highlight the molecular heterogeneity of CF in the Spanish population. © 1994 Wiley-Liss, Inc.     

Ataxia-telangiectasia in the Japanese population: Identification of R1917X,W2491R,R2909G,IVS33+2T→A,and 7883del5, the latter two being relatively common mutations

We analyzed the data regarding six Japanese ataxia-telangiectasia (A-T) patients from four unrelated families, at the DNA level, to search for possible common mutations in the Japanese population. Among eight mutant alleles in the four families, c. 4612del165 (exon 33 skipping) was identified in two alleles, and c. 5749A to T (R1917X), c. 7471T to C (W2491R), c.7883del5, and c. 8725A to G (R2909G) were identified in one allele each. We found no mutations in the other two alleles. The IVS33+2T→A mutation was identified at the genomic level as the cause of exon 33 skipping. We also identified the IVS33+2T→A mutation in a Japanese patient ATL105 who was previously found to be a homozygote of c. 4612del165. W2491R and R2909G mutations were not detected in more than 100 control Japanese alleles. The latter is located in a highly conserved PI-3 kinase domain and is a completely conserved residue among ATM-related proteins. Taken together with previously documented mutations in five other Japanese A-T patients, IVS33+2T→A and 7883del5 were identified in four and five alleles, respectively, in a total of 18 mutant alleles of Japanese A-T patients. These results suggest that these two mutations are relatively common mutations in the Japanese population. Hum Mutat 12:338–343, 1998.© 1998 Wiley-Liss, Inc.     

Identification and characterization of CFTR gene mutations in Indian CF patients

Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. This study was performed on Indian CF patients (n = 50) to investigate the spectrum of mutations in the CFTR gene and their association with intragenic and extragenic marker haplotypes. We report identification of 14 previously known and eight novel mutations, namely 3986-3987delC, 876-6del4, 1792InsA, L69H, S158N, Q493L, I530L and E1329Q. The frequency of delta F508 was found to be 27%. Absolute linkage between delta F508 and the KM.19-GATT-TUB9-M470V-T854T haplotype (2-2-1-1-1) predicts a relatively recent appearance of delta F508 in Indian CF patients. Low frequency of delta F508 mutation and detection of eight novel and thirteen rare mutations reflect a heterogeneous spectrum of mutations in Indian CF patients. Failure to detect mutations in 34% of alleles indicates the possible presence of gross deletions involving one or more exons or may indicate the location of the molecular defects in either the noncoding parts of the gene or in the promoter region, which warrants analysis of those regions.     

Development of RNA-SSCP protocols for the identification and screening of CFTR mutations: Identification of two new mutations

A strategy is described that allows a rapid and accurate identification and screening of cystic fibrosis gene mutations. It consists of setting up and developing RNA single strand conformation polymorphism (rSSCP) protocols, a technique based on the large repertoire of secondary structure of single-stranded RNA. By incorporating the T7 phage promoter sequence into PCR primers, it is possible to carry out rSSCP and compare it to standard single-strand conformation polymorphism (SSCP). Several parallel tests indicate that rSSCP detects a higher fraction of single base changes, and is less time consuming than SSCP since it requires only one fairly short electrophoretic run. Using this technique we were able to identify two new splicing mutations in introns 5 (711+5G→A) and 10 (1717–8G→A) of the CFTR gene. © 1994 Wiley-Liss, Inc.     

两个斑驳病家系KIT基因突变研究

目的 对2个斑驳病家系进行致病基因突变分析,为患者及其家系成员提供遗传咨询和生育指导.方法 分别采集家系1两例患者(先证者及其父亲)、家系2先证者及3名表型正常家系成员的外周血,提取外周血DNA和RNA.应用PCR、逆转录PCR及测序等技术,从基因组水平和表达水平对此两家系先证者和患者进行KIT基因诊断,并初步探讨检测到的突变对KIT基因功能的影响.结果 家系l中两例患者KIT基因均存在IVS12+ 2_+7delinsACATCTTTA的杂合突变,该突变在cDNA水平导致KIT基因c.1765-1779del突变,在氨基酸水平导致p.Gly592Ala/del:E12突变,使得KIT基因剪切位点发生改变,即其中一条cDNA第12外显子被跨越、未转录.家系2中先证者KIT基因存在c.2401A＞C突变,3位表型正常的家系成员未见该突变.结论 确诊了两个斑驳病家系的致病原因.家系1患者KIT基因均存在IVS12+ 2_+ 7delinsACATCTTTA的杂合突变,该突变为人类基因突变数据库未记载的、新的剪切突变;家系2先证者KIT基因存在c.2401A＞C突变,结合3位表型正常的家系成员KIT基因未见c.2401A＞C突变,推测该突变为先证者患斑驳病的致病突变可能性大.为此两家系进行遗传咨询和产前诊断提供了理论依据.     

Identification of two novel frame shift mutations of the NF1 gene in Korean patients with neurofibromatosis type 1

Neurofibromatos is type 1 (NF1) is one of the most common inherited disorders and is characterized by abnormalities in multiple tissues derived from the neural crest. The NF-1 gene has been cloned and mapped to human chromosome 17q11.2. The NF-1 gene has an open reading frame that predicts a protein consisting of 2,818 amino acids, known as neurofibromin. Here, we report two kinds of novel frame shift mutations of the NF1 gene from 2 out of 56 unrelated Korean NF1 patients. These mutations were detected using polymerase chain reaction and single strand conformational polymorphism analysis. Sequencing analysis revealed four base pair insertion at codon 1270 of exon 22, and a base pair deletion at codon 1398 of exon 24. These mutations resulted in premature termination of the mutant alleles and may encode truncated forms of neurofibromin.     

Identification of 12 novel mutations in the SLC3A1 gene in Swedish cystinuria patients

Cystinuria is an autosomal recessive disorder that affects luminal transport of cystine and dibasic amino acids in the kidneys and the small intestine. Three subtypes of cystinuria can be defined biochemically, and the classical form (type I) has been associated with mutations in the amino acid transporter gene SLC3A1. The mutations detected in SLC3A1 tend to be population specific and have not been previously investigated in Sweden. We have screened the entire coding sequence and the intron/exon boundaries of the SLC3A1 gene in 53 cystinuria patients by means of single strand conformation polymorphism (SSCP) and DNA sequencing. We identified 12 novel mutations (a 2 bp deletion, one splice site mutation, and 10 missense mutations) and detected another three mutations that were previously reported. Five polymorphisms were also identified, four of which were formerly described. The most frequent mutation in this study was the previously reported M467T and it was also detected in the normal population with an allelic frequency of 0.5%. Thirty‐seven patients were homozygous for mutations in the SLC3A1 gene and another seven were heterozygous which implies that other genes may be involved in cystinuria. Future investigation of the non‐type I cystinuria gene SLC7A9 may complement our results but recent studies also suggest the presence of other potential disease genes. Hum Mutat 18:516–525, 2001. © 2001 Wiley‐Liss, Inc.     

Identification of two novel mutations in the OCRLI gene in Japanese families with Lowe syndrome

Kubota T, Sakurai A, Arakawa K, Shimazu M, Wakui K, Furihata K, Fukushima Y. Identification of two novel mutations in the OCRL1 gene in Japanese families with Lowe syndrome. Clin Genet 1998: 54: 199–202. 0 Munksgaard, 1998
The oculocerebrorenal syndrome of Lowe (OCRL) is a rare X-linked disorder with features of congenital cataracts. Fanconi syndrome of the renal tubule, and mental retardation. The OCRLI gene has been positionally cloned and shown to encode a phosphatidylinositol 4.5–biphos-phate-5–phosphatase. OCRL is thus thought to be an inborn error of inositol polyphosphate metabolism. We analyzed the gene in two Japanese OCRL patients and their families by DNA sequencing and mismatch polymerase chain reaction (PCR) followed by restriction digestion. A novel nonsense mutation (C1399T) replacing the glutamine of codon 391 (Gln 391 Stop) was identified in exon 12 in 1 patient and also in his mother. A novel missense mutation (C1743G) was identified in exon 15 in the second patient, his mother and maternal grandmother. The missense mutation predicts a substitution of serine for arginine (Ser 505 Arg) in a domain highly conserved among the inosi-tol-5–phosphatase family. Our observations expand the range of OCRLI mutations that cause Lowe syndrome. and will be useful for genetic counseling in these two Fdmilies.