两个新RUNX2基因突变引起家族性锁骨颅骨发育不全

Objective To identify the RUNX2 gene mutation in two unrelated Chinese families with cleidocranial dysplasia (CCD), and to assess the feasibility of gene diagnosis for patients with CCD. Methods Genomic DNA was isolated from peripheral blood samples of 4 patients and 4 healthy members in the two pedigrees as well as 102 unrelated healthy controls. All 7 coding exons and their flanking intronic sequences of the RUNX2 gene were amplified by PCR, then the PCR products were sequenced bi-directionally. The sequencing results were compared with normal sequences in GenBank to identify the mutation. The mutation was confirmed by RFLP with restriction endonuclease. Results In one family, a novel heterozygous missense mutation c. 346T＞A (W116R) in exon 1 of the RUNX2 gene was detected in the two affected individuals, and the mutation was further confirmed with Bsr Ⅰ restriction endonuclease digestion. In the other family, a novel nonsense mutation c. 610A＞T (K204X) was identified in the two patients. No above sequence change was found in the 102 healthy controls. Conclusion Two novel RUNX2 mutations were found in two unrelated Chinese families with cleidoeranial dysplasia. The identification of these mutations further extended the mutation spectrum of RUNX2 gene and will facilitate prenatal diagnosis and gene diagnosis of CCD.     

两个新RUNX2基因突变引起家族性锁骨颅骨发育不全

Objective To identify the RUNX2 gene mutation in two unrelated Chinese families with cleidocranial dysplasia (CCD), and to assess the feasibility of gene diagnosis for patients with CCD. Methods Genomic DNA was isolated from peripheral blood samples of 4 patients and 4 healthy members in the two pedigrees as well as 102 unrelated healthy controls. All 7 coding exons and their flanking intronic sequences of the RUNX2 gene were amplified by PCR, then the PCR products were sequenced bi-directionally. The sequencing results were compared with normal sequences in GenBank to identify the mutation. The mutation was confirmed by RFLP with restriction endonuclease. Results In one family, a novel heterozygous missense mutation c. 346T＞A (W116R) in exon 1 of the RUNX2 gene was detected in the two affected individuals, and the mutation was further confirmed with Bsr Ⅰ restriction endonuclease digestion. In the other family, a novel nonsense mutation c. 610A＞T (K204X) was identified in the two patients. No above sequence change was found in the 102 healthy controls. Conclusion Two novel RUNX2 mutations were found in two unrelated Chinese families with cleidoeranial dysplasia. The identification of these mutations further extended the mutation spectrum of RUNX2 gene and will facilitate prenatal diagnosis and gene diagnosis of CCD.     

Genetic analysis of two children patients affected with CHARGE syndromeCSCD

Objective To analyze two Chinese pediatric patients with multiple malformations and growth and development delay. Methods Both patients were subjected to targeted gene sequencing, and the results were analyzed with Ingenuity® Variant Analysis™ software. Suspected pathogenic variations were verified by Sanger sequencing. Results High-throughput sequencing showed that both patients have carried heterozygous variants of theCHD7 gene. Patient 1 carried a nonsense mutation in exon 36 (c. 79570T, p. Arg2653 ∗), while patient 2 carried a nonsense mutation of exon 2 (c. 7180T, p. Gln240 ∗ ). Sanger sequencing confirmed the above mutations in both patients, while their parents were of wild-type for the corresponding sites, indicating that the two mutations have happened de novo. Conclusion Two patients were diagnosed with CHARGE syndrome by high-throughput sequencing. © 2018 West China University of Medical Sciences. All rights reserved.     

中国人圆锥角膜患者的TGFBI基因编码区点突变及多态性的检测和分析

Objective To investigate the point mutations and polymorphisms of transforming growth factor β-induced gene (TGFBI) in Chinese patients with keratoconus and discuss the relationship between the feature of gene mutations and single nucleotide polymorphisms of TGFBI gene and keratoconus. Methods Polymerase chain reaction-single strand conformation polymorphism and DNA direct sequencing were performed in 30 keratoconus cases and 30 healthy controls. All 17 exons of the TGFBI gene were analyzed for point mutations and single nucleotide polymorphisms. Results Totally two heterozygous nucleotide changes were identified in exon 12 of the TGFBI gene. The codon 535 is changed from GGA to TGA in 1 patient, leading to a substitution of glycine to a stop codon at the protein level (G535X). The codon 540 is changed from TTT to TTC in 2 patients and 1 control individual, resulting in a nonsense mutation (F54F),and is a single nucleotide polymorphism of the gene. Conclusion Mutation and polymorphisms of the TGFBI gene were detected in Chinese patients with keratoconus in this study. The results suggest that TGFBI gene might play an important role in the pathogenesis of keratoconus.     

中国人圆锥角膜患者的TGFBI基因编码区点突变及多态性的检测和分析

Objective To investigate the point mutations and polymorphisms of transforming growth factor β-induced gene (TGFBI) in Chinese patients with keratoconus and discuss the relationship between the feature of gene mutations and single nucleotide polymorphisms of TGFBI gene and keratoconus. Methods Polymerase chain reaction-single strand conformation polymorphism and DNA direct sequencing were performed in 30 keratoconus cases and 30 healthy controls. All 17 exons of the TGFBI gene were analyzed for point mutations and single nucleotide polymorphisms. Results Totally two heterozygous nucleotide changes were identified in exon 12 of the TGFBI gene. The codon 535 is changed from GGA to TGA in 1 patient, leading to a substitution of glycine to a stop codon at the protein level (G535X). The codon 540 is changed from TTT to TTC in 2 patients and 1 control individual, resulting in a nonsense mutation (F54F),and is a single nucleotide polymorphism of the gene. Conclusion Mutation and polymorphisms of the TGFBI gene were detected in Chinese patients with keratoconus in this study. The results suggest that TGFBI gene might play an important role in the pathogenesis of keratoconus.     

Liddle's syndrome caused by a novel mutation of the γ-subunit of epithelial sodium channel gene SCNN1G in Chinese

Objective To screen the mutation of the β and γ subunits of epithelial sodium channel gene SCNN1 in two families with Liddle's syndrome. Methods Two patients clinically diagnosed as Liddle's syndrome and their family members were enrolled. Peripheral blood samples were collected and total genomic DNA was prepared. Polymerase chain reaction (PCR) was used to amplify the exon 13 of the SCNN1B and SCNN1G gene. PCR products were purified and subjected to direct DNA sequencing. Results A heterozygous nonsense mutation at codon 564 of the SCNN1B gene from CGA(Arg) to stop codon(TGA)was detector in the proband of family 1. More importantly, a novel heterozygous nonsense mutation of CAG (Gln) to stop codon TAG at codon 567 of the SCNN1G gene was detected in the proband and another two members of family 2. Conclusion Screening for specific mutations of the SCNN1 gene in relatives of patients with Liddle's syndrome can be used to identify the previously unrecognized cases within the family.A new nonsense mutation(Q567X) of the SCNN1G gene is likely the cause of Liddle's syndrome in family 2.     

Liddle's syndrome caused by a novel mutation of the γ-subunit of epithelial sodium channel gene SCNN1G in Chinese

Objective To screen the mutation of the β and γ subunits of epithelial sodium channel gene SCNN1 in two families with Liddle's syndrome. Methods Two patients clinically diagnosed as Liddle's syndrome and their family members were enrolled. Peripheral blood samples were collected and total genomic DNA was prepared. Polymerase chain reaction (PCR) was used to amplify the exon 13 of the SCNN1B and SCNN1G gene. PCR products were purified and subjected to direct DNA sequencing. Results A heterozygous nonsense mutation at codon 564 of the SCNN1B gene from CGA(Arg) to stop codon(TGA)was detector in the proband of family 1. More importantly, a novel heterozygous nonsense mutation of CAG (Gln) to stop codon TAG at codon 567 of the SCNN1G gene was detected in the proband and another two members of family 2. Conclusion Screening for specific mutations of the SCNN1 gene in relatives of patients with Liddle's syndrome can be used to identify the previously unrecognized cases within the family.A new nonsense mutation(Q567X) of the SCNN1G gene is likely the cause of Liddle's syndrome in family 2.