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Chen X.,Shenzhen Institute of Pediatrics | Lv R.,Shenzhen Childrens Hospital | Wen F.,Shenzhen Childrens Hospital | Chen Y.,Clinical Laboratory | Liu F.,Shenzhen Childrens Hospital
Indian Journal of Hematology and Blood Transfusion | Year: 2016

Human glucose-6-phosphate dehydrogenase deficiency (G6PD) is mostly caused by single nucleotide change in the G6PD gene which leads to single amino acid substitution. Previous trials suggested a few samples had decreased ratio of G6PD/6PGD(<1.00) but no mutation detected by multiple methods. In 138 cases of Chinese children with G6PD deficiency, RT-PCR combined with DNA Sequencing was performed to screen the mutations in the coding region and promoter region of G6PD gene. The mutation detection frequency by this method was 100 %, including a novel missense mutation (1088 A>T) and 13 mutations reported before. The novel mutation predicted an Asn-to-Ile substitution at codon 363, which was identified in a male infant patient. The variant caused by this mutation had reduced enzymatic activity, belonging to WHO Class I. Synonymous or missense mutation was not found in the proximal promoter region of the G6PD gene, which was consistent with earlier findings that G6PD deficiency was not associated with promoter mutations in the G6PD gene. RT-PCR combined with DNA Sequencing could be another alternative for clinically molecular diagnosis of G6PD deficiency. © 2016, Indian Society of Haematology & Transfusion Medicine.

Wen P.,Shenzhen Institute of Pediatrics | Chen Z.,Dongguan Peoples Hospital | Wang G.,Shenzhen Institute of Pediatrics | Su Z.,Shenzhen Institute of Pediatrics | And 5 more authors.
Chinese Journal of Medical Genetics | Year: 2016

Objective: To investigate the clinical phenotype and ACATI gene mutation in a family affected with β-ketothiolase deficiency (BKTD). Methods: Clinical features and laboratory test data were collected. The probands were monozygotic twin brothers. Genomic DNA was isolated from peripheral blood leukocytes obtained from the probands and their family members. Molecular genetic testing of the ACAT1 gene was carried out. Results: The probands have presented with fever, vomiting and severe ketoacidosis. By arterial blood gas testing, pH was determined to be 7. 164, bicarbonate was 4. 0 mmol/L, and urine ketone was + + + +. Urinary organic acid gas chromatography-mass spectrometry analysis showed excessive excretion of 3-hydroxybutyric acid, 2-methyl-3-hydroxybutyric acid and tiglylglycine. Increased 3-hydroxybutyrylcarnitine (C4-OH), tiglylcarnitine (C5:1) and 3-hydroxyisovalerylcarnitine (C5-0H) levels. The clinical phenotype of proband's parents were both normal, but an elder sister turned out to be an affected patient. Genetic analysis has identified two heterozygous mutations [c. 622C>T(p. R208X) and c. 653C>T(p. S218F)] in the proband, which were respectively detected in the mother and father. The c. 653C>T(p. S218F) mutation was not found among the 100 healthy controls and has not been included in the Human Gene Mutation Database (HGMD). Conclusion: The primary clinical manifestations of BKTD is ketoacidosis. Urine organic acid and blood acylcarnitine analyses play an important role in the diagnosis of the disease. The compound heterozygous of ACAT1 gene mutations probably underlie the BKTD in our patient. © 2016, West China University of Medical Sciences. All rights reserved.

Zhang H.,Shenzhen Institute of Pediatrics | Yue L.,Shenzhen Institute of Pediatrics | Yuan X.,Shenzhen Childrens Hospital | Zheng M.,Shenzhen Institute of Pediatrics | And 2 more authors.
Chinese Journal of Clinical Oncology | Year: 2012

Objective: This study aims to investigate the distribution of γ-glutamyl hydrolase gene (GGH) T16C genotype and allele frequency in children with acute leukemia (AL) and healthy children and to identify the novel single nucleotide polymorphism (SNP). Methods: Bone marrow samples from 90 children with AL and peripheral blood samples from 120 healthy children (controls) were obtained to prepare complementary DNA (cDNA). The cDNA was analyzed for the polymorphism in GGH T16C by direct sequencing. Results: A novel missense mutation was a T→G transition in the second nucleotide of codon 64 in exon 2 of the GGH gene, which was predicted to result in the substitution of leucine with tryptophan (Leu 64 Trp). The T16C polymorphism in the coding region of the GGH gene was first identified in Chinese children. The T16C allelic frequency in the AL children was 20%, whereas that in the control group was 17.1%. The allelic frequency of SNP was not associated with susceptibility to AL. Conclusion: This work is the first report concerning the examination of T16C polymorphism of GGH in Chinese Han children. A novel GGH gene missense mutation T191G was identified in healthy Chinese children.

Wen P.-Q.,Shenzhen Institute of Pediatrics | Wang G.-B.,Shenzhen Institute of Pediatrics | Chen Z.-L.,Dongguan Peoples Hospital | Cui D.,Shenzhen Institute of Pediatrics | And 6 more authors.
Chinese Journal of Medical Genetics | Year: 2012

Objective: To assess the feasibility of high-resolution melting (HRM) analysis for screening patients with neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD). Method: Based on previous studies on SLC25A13 gene in Chinese patients with NICCD, four hotspot mutations (851del4, 1638ins23, IVS6+5G>A and IVS16ins3kb) were selected. Results of the HRM analysis was validated using 50 negative controls and 20 patients with NICCD whose genotypes were confirmed previously by direct sequencing. With the established protocol, 171 suspected patients were enrolled. Samples with abnormal melting curves were further validated by DNA sequencing. Results: HRM analysis can accurately determine the genotypes of all negative controls and patients. The sensitivity and specificity of the technique reached 100% (70/70). The melting curves of samples with the same genotype were highly reproducible. In 171 suspected patients, seven NICCD patients were detected by HRM. Identified mutations included one case of 851del4 homozygote, one case of IVS6+5G>A heterozygote, 3 cases of 851del4 heterozygotes, one case of [IVS6+5G>A]+[851del4] and one case of [1638ins23+IVS16ins3kb]+[1638ins23]. All mutations were subsequently confirmed by DNA sequencing. Conclusion: HRM analysis is a convenient, high-throughput and rapid technique for the screening of NICCD patients.

Zhao W.,Guangdong Medical College | Yue L.-J.,Shenzhen Institute of Pediatrics | Chen X.-W.,Shenzhen Institute of Pediatrics
Chinese Journal of Medical Genetics | Year: 2011

Objective: To investigate the allele and genotype frequencies of reduced folate carrier gene (RFC) 80G/A polymorphism in Chinese patients with acute leukemia (AL) and healthy control children, and to provide clue for association between the single nucleotide polymorphism (SNP) of RFC and the occurrence of AL. Methods: Bone marrow samples from 98 childhood patients with AL and peripheral blood samples from 135 healthy children were obtained to prepare complementary DNAs (cDNAs). The cDNAs were analyzed for the polymorphisms in RFC 80G/A by reverse transcriptase-polymerase chain reaction-denaturing gradient gel electrophoresis (RT-PCR-DGGE) and direct sequencing. Results: The A allele frequencies of the AL patients and control children were 0.515 and 0.415, respectively (P<0.05). Chi-square test confirmed a statistical significance of the association between RFC80 G/A and AL. Conclusion: RFC 80AA or GA genotype may contribute to increasing the susceptibility to AL.

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