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Objective: To explore the genotype mutation characteristics of patients with glucose-6-phosphate dehydrogenase(G6PD) deficiency in Wuhan., Methods: A total of 1 321 neonates with positive screening and outpatients were received G6PD mutation detection, 12 kinds of common G6PD mutation in Chinese people was detected by using multicolor melting curve analysis (MMCA) method, for those with negative results, the enzyme activity and clinical information were analyzed, sequencing was recommended after informed consent when it is necessary., Results: Among 1321 patients, a total of 768 mutations were detected out, with a detection rate of 58.1%. A total of 18 types of G6PD genotypes were identified, including c.1388G>A, c.1376G>T, c.95G>A, c.1024C>T, c.871G>A, c.392G>T, c.487G>A, c.1360C>T, c.1004C>A, c.517T>C, c.592C>T, c.94C>G, c.152C>T, c.320A>G, c.1028A>G, c.1316G>A, c.1327G>C and c.1376G>C, including 683 male hemizygotes, 3 female homozygotes, 80 female heterozygotes and 2 female compound heterozygous., Conclusion: A total of 18 types of G6PD mutations are identified in the reaserch, and c.94C>G, c.1028A>G and c.1327G>C are first reported in Chinese population. The most common G6PD mutation types in Wuhan are c.1388G>A, c.1376G>T, c.95G>A.

Objective: To analyze the screening results of glucose-6-phosphate dehydrogenase (G6PD) deficiency and gene mutation distribution of G6PD deficiency in preterm infants in Chengdu, China, in order to provide a basis for the improvement of G6PD screening process in preterm infants., Methods: Fluorescent spot test for G6PD deficiency using dried blood spots was used for G6PD screening of 54 025 preterm infants born from January 1, 2015 to December 31, 2019 in Chengdu, and G6PD enzymology and gene detection were used for the diagnosis of 213 infants with positive screening results., Results: Among the 54 025 preterm infants, 192 were diagnosed with G6PD deficiency, with an incidence rate of 3.55‰. The incidence rate of G6PD deficiency in preterm infants was higher than that in full-term infants in the same period of time and tended to increase year by year. Birth in summer, gestational age <32 weeks, and birth weight <2 500 g were influencing factors for the increase in false positive rate of screening ( P < 0.05). The diagnostic accordance rate of genetic tests was significantly higher than that of enzyme activity assay in female infants ( P < 0.05). Nine gene mutations were detected in Chengdu, without compound heterozygous mutation. Homozygous mutation was not detected in female infants. In the 80 infants with gene mutations, the top three gene mutations were c.1388G>A in 26 infants (32%), c.1376G>T in 21 infants (26%), and c.1024C>T in 13 infants (16%), accounting for 75%. There was a significant difference in pathogenicity grading among the three gene mutations ( P < 0.001). The pairwise comparison showed that c.1024C>T had a significantly lower pathogenicity grade than c.1376G>T and c.1388G>A ( P < 0.0167), suggesting that c.1376G>T and c.1388G>A had greater influence on enzyme activity than c.1024C>T., Conclusions: Screening for G6PD deficiency in preterm infants should be taken seriously. It is recommended to apply cold-chain transportation of samples in summer to reduce the false positive rate of primary screening for G6PD deficiency. Genetic tests should be promoted in girls with positive screening results to improve the detection rate of G6PD deficiency in preterm female infants. There are various types of gene mutations in preterm infants with G6PD deficiency in Chengdu, and infants with c.1024C>T mutation tend to have mild conditions.

Objective: To reveal the relationship between G6PD genotypes and the G6PD enzyme activities in dried blood spots of newborn screening. Methods: Simple random sampling procedure was used in this study. The fluorescence PCR melting curve analysis was performed to classify G6PD gene variants in 635 neonates coming from Guangzhou Newborn Screening Center during October 1 to 20, 2016, including 15 reported variants. Those samples consisted of 377 cases with screening positive results (261 from males and 116 from females) and 258 cases with screening negative results (32 from males and 226 from females). The cut-off value of G6PD was less than 2.6 U/g Hb in dry blood spots. Sanger sequencing for G6PD gene was used in 7 cases with screening negative results under simple random sampling. One-way ANOVA and least significant difference method (LSD) test were performed to compare the difference of G6PD activity among genotypes. Results: The top 6 frequency of G6PD gene variants were c.1388G>A(35.07%), c.1376G>T(32.13%), c.95A>G(12.72%), c.871G>A(8.32%), c.1024C>T(4.08%) and c.392G>T(2.28%), accounting for 94.62% of all variant alleles (580/613). A total of 253 males positive for enzyme activity were detected to have gene mutations. The positive rate of G6PD enzyme activity was 98.06%(253/258). The mean values of G6PD activities for c.1376G>T,c.95A>G and c.1388G>A were 0.85, 1.10 and 1.28 U/g Hb, respectively. There were significant differences among the three groups ( F =28.7, P <0.01). A total of 105 females positive for enzyme activity were detected to have gene mutations. The positive rate of G6PD enzyme activity was 90.52%(105/116). The positive rate of G6PD enzyme activity was 26.95% among 256 females with one point mutation while it was 83.72% in females with multi-allele variants. The G6PD activity of heterozygous females was (2.9±0.8) U/g Hb, which was significant higher than that of females with multi-allele variants (1.5±1.0) U/g Hb ( t =8.6, P <0.01). Conclusions: G6PD activities in dried blood spots were related to G6PD genotypes in males. They were also associated with the numbers of allele variants in females. Newborn screening for G6PD deficiency can be used to detect most of G6PD-deficient hemizygotes and female patients with multi-allele variants, which is helpful for preventing neonatal jaundice and medicine application.

Objective: To explore the clinical significance of G6PD gene mutation detection in female heterozygote with G6PD deficiency., Methods: G6PD activity and fourteen common G6PD gene mutations in female blood samples were detected by biochemical phenotype detection and PCR-reverse dot blotting, respectively. Unidentified genotype of G6PD positive samples was further ascertained by direct DNA sequencing. The results from two methods were compared and analyzed., Results: A total of 493 unrelated females were enrolled, and the G6PD activity and G6PD mutations was detected. Among them, 473 females were found to be normal in G6PD activity and 20 females with G6PD deficiency, and the detection rate by G6PD activity method was 4.06%. In all enrolled females, G6PD gene mutations, including the mutation of c.1311 C＞T, were identified in 130 females, and the detection rate was 26.3%. Detection rate of the mutations that can lead to G6PD deficiency was 8.11%. The detection rates between the two methods were significantly different (P＜0.01). The misdiagnosis rate of the G6PD activity detection reached 49. 94% for the female heterozygotes. Eight G6PD mutations and 13 mutation patterns were identified in the research, and most of mutation patterns were single nucleotide missense mutation. In addition to c.1311C＞T mutation, the most common mutations were c.1376G＞T, c.1388G＞A and c.95 A＞G. G6PD mutations were identified in 19 of 20 females with G6PD deficiency, and were also detected in 21 of 473 females with normal G6PD activity, of which the rate of heterozygous mutation was 90.88%., Conclusion: The phenotype detection based on G6PD enzyme activity alone is not sufficient for the diagnosis of female heterozygotes. The detection of G6PD mutations that covers the common mutations in specified region can effectively identify the female heterozygotes with normal G6PD activity.

Objective: To analyze the hotspots of known pathogenic disease-causing variants of glucose-6-phosphate dehydrogenase (G6PD) and the phenotype spectrum of neonatal patients with known pathogenic disease-causing variants of G6PD. Methods: The known pathogenic disease-causing variants of G6PD were collected from Human Gene Mutation Database. Screening was performed for these variants among the 7 966 cases (2 357 neonatal, 5 609 non-neonatal) in the database of sequencing at Molecular Diagnosis Center, Children's Hospital of Fudan University. All these samples were from patients suspected with genetic disorder. The database contained Whole Exon Sequencing data and Clinical Exon Sequencing data. We screened out the patients with known pathogenic disease-causing variants of G6PD, analyzed the hotspot of G6PD and the phenotype spectrum of neonatal patients with known pathogenic disease-causing variants of G6PD. Results: (1) Among the next generation sequencing data of the 7 966 samples, 86 samples (1.1%) were detected as positive for the known pathogenic disease-causing variants of G6PD (positive samples set). In the positive sample set, 51 patients (33 males, 18 females) were newborn babies. Forty-three patients (26 males, 17 females) had the enzyme activity data of G6PD. (2) Among the 86 samples, Arg463His, Arg459Leu, Leu342Phe, Val291Met were the leading 4 disease-causing variants found in 72 samples (84%). (3) Male neonatal patients with the same variants had the statistically significant differences in enzyme activity: among 13 patients with Arg463His, enzyme activity of 9 patients was ranked as grade Ⅲ, 1 case ranked as Ⅳ, 3 cases had no activity data;among 10 patients with Arg459Leu, enzyme activity of 4 patients was ranked as Ⅱ, 4 cases ranked as Ⅲ, 2 cases had no activity data;among 2 patients with His32Arg, enzyme activity of one patient was ranked as Ⅱ, another was Ⅲ. Male neonatal patients with the same mutation and enzyme activity also had the statistically significant differences in phenotype spectrum: among 9 patients with Arg463His and level Ⅲ enzyme activity, 6 presented hyperbilirubinemia, 2 met the criteria for exchange transfusion therapy, 2 showed hemolysis;among 4 patients with Arg459Leu and level Ⅱ enzyme activity, 3 presented hyperbilirubinemia;among 4 patients with Arg459Leu and level Ⅲ enzyme activity, 2 presented hyperbilirubinemia, 1 met the standard of exchange transfusion therapy;among 3 patients with Val291Met and level Ⅲ enzyme activity, 1 presented hyperbilirubinemia. Conclusions: Arg463His, Arg459Leu, Leu342Phe, Val291Met were the hotspots variants for the G6PD. Patients with the same G6PD variants and sex present different phenotype, patients with the same G6PD variants, sex and enzyme activity also present different phenotype .

Objective: Since glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common hereditary hemolytic erythrocyte enzyme deficiency, most cases have single nucleotide mutations in the coding region, and current test methods for gene mutation have some missed detections, this study aimed to investigate the feasibility of RT-PCR sequencing in the detection of gene mutation in G6PD deficiency., Methods: According to the G6PD/6GPD ratio, 195 children with anemia of unknown cause or who underwent physical examination between August 2013 and July 2014 were classified into G6PD-deficiency group with 130 children (G6PD/6GPD ratio <1.00) and control group with 65 children (G6PD/6GPD ratio≥1.00). The primer design and PCR amplification conditions were optimized, and RT-PCR sequencing was used to analyze the complete coding sequence and verify the genomic DNA sequence in the two groups., Results: In the G6PD-deficiency group, the detection rate of gene mutation was 100% and 13 missense mutations were detected, including one new mutation. In the control group, no missense mutation was detected in 28 boys; 13 heterozygous missense mutations, 1 homozygous same-sense mutation (C1191T) which had not been reported in China and abroad, and 14 single nucleotide polymorphisms of C1311T were detected in 37 girls. The control group showed a high rate of missed detection of G6PD deficiency (carriers) in the specimens from girls (35%, 13/37)., Conclusions: RT-PCR sequencing has a high detection rate of G6PD gene mutation and a certain value in clinical diagnosis of G6PD deficiency.

Objective: To investigate the features of methylation in the promoter region of glucose-6-phosphate dehydrogenase (G6PD) gene and the association between gene promoter methylation and G6PD deficiency., Methods: Fluorescent quantitative PCR was used to measure the mRNA expression of G6PD in 130 children with G6PD deficiency. Sixty-five children without G6PD deficiency served as the control group. The methylation-sensitive high-resolution melting curve analysis and bisulfite PCR sequencing were used to analyze gene promoter methylation in 22 children with G6PD deficiency and low G6PD mRNA expression. The G6PD gene promoter methylation was analyzed in 44 girls with normal G6PD mRNA expression (7 from G6PD deficiency group and 37 from control group)., Results: Twenty-two (16.9%) children with G6PD deficiency had relatively low mRNA expression of G6PD; among whom, 16 boys showed no methylation, and 6 girls showed partial methylation. Among the 44 girls with normal G6PD mRNA expression, 40 showed partial methylation, and 4 showed no methylation (1 case in the G6PD group and 3 cases in the control group)., Conclusions: Gene promoter methylation is not associated with G6PD deficiency in boys. Girls have partial methylation or no methylation in the G6PD gene, suggesting that the methylation may be related to G6PD deficiency in girls.

Objective: To determine the incidence and molecular characteristics of G6PD deficiency in Chaozhou region of eastern Guangdong Province., Methods: G6PD enzyme activity was assayed with an auto-bioanalyzer. Reverse dot blotting (RDB) was used for detecting 6 common G6PD mutations. Samples with no mutation detected by RDB were further sequenced for unknown mutations., Results: The rate of G6PD deficiency was 3.36% (142/4224). 2.33% (47/2013) of males and 4.3% (95/2208) of females were affected. 12 mutations were detected among the 142 patients, which included c.1376G>T, c.1388G>A, c.1024C>T, c.392G>T, c.871G>A, c.95A>G, c.517T>C, c.131C>G, c.1376G>T/c.517T>C, c.871G>A/IVS-1193T>C/c.1311C>T, c.1376G>T/IVS-11, 93T>C/c.1311C>T and c.1376G>T/c.486&#95;34delT (rs3216174)., Conclusion: The incidence of G6PD deficiency in Chaozhou region was lower than that of the Hakka population of Guangdong Province, and the mutation types were diversely distributed in this region. c.1376G>T, c.1388G>A and c.1024C>T were the most common mutations, which was followed by c.517T>C. In addition, c.131C>G has been first discovered in the Chinese population. c.1376G>T/c.517T>C and c.1376G>T/c.486&#95;34delT(rs3216174) were new types of compound heterozygous mutations in females.

Organic anion transporter 2 (OATP2) is an uptake transporter located on the basolateral membrane of human hepatocytes. It mediates the transportation of various organic solutes including bilirubin and impacts bilirubin metabolism. It is encoded by the gene of solute carrier organic anion transporter family member 1B1 and the gene variants that inhibit hepatic bilirubin uptake function may reduce the normal functional level of bilirubin elimination and result in neonatal hyperbilirubinemia. In recent years, some studies have indicated that variants of SLCO1B1 are associated with neonatal jaundice. This article reviews the research advance in SLCO1B1 with respect to the structure and function and the relationship between SLCO1B1 mutations and neonatal jaundice.

Objective: To investigate the epidemiological status of glucose-6-phosphate dehydrogenase (G6PD) deficiency among children from Yunnan with unique ethnic origins., Methods: DNA samples from 11759 children were tested with fluorescent spot test, G6PD/6PGD quantitative ratio assay and hemoglobin electrophoresis., Results: The detection rate of G6PD deficiency was 2.5%, for which boys were significantly greater than girls (3.5% vs. 1.4%, P<0.05). Significant differences were also detected among children from different ethnic groups and different regions. For ethnic Han Chinese, the detection rate was 0.7%, which was lower than the majority of ethnic minorities. By regression analysis, altitude of residence and family history both have significant influence on the calculated rate., Conclusion: Occurrence of G6PD deficiency seems to be influenced by gender. It also varies substantially between different ethnic groups as well as regions, e.g., more common in south. It also showed a declining trend after years of diagnosis and intervention. This survey may provide a valuable basis for counseling of G6PD deficiency in Yunnan.

Objective: To study the correlation between glucose-6-phosphate dehydrogenase (G-6-PD) activities and three common mutations of G-6-PD gene G1388A, G1376T and A95G and investigate the effects of G-6-PD gene mutations on neonatal jaundice in Nanning, Guangxi., Methods: One hundred and twenty-four neonates from Nanning, Guangxi, with hyperbilirubinemia were enrolled. The ARMS-PCR and PCR/REA methods were used to determine G-6-PD gene mutations. G-6-PD activities were measured using the NBT method. The incidence of acute bilirubin encephalopathy and the peak bilirubin concentration 72 hrs after birth were compared between the neonates with different genotypes and between the G-6-PD mutation and normal groups. The risk of blood serum bilirubin >340 mumol/L was evaluated by logistic regression analysis., Results: Of the 124 cases, gene mutations were found in 37 cases, including G1388A (n=20), G1376T (n=14), A95G (n=4) and G1388A+A95G (n=1). Five cases (25%) showed normal G-6-PD activities in the G1388A gene mutation group and 4 (29%) had normal G-6-PD activities in the G1376T G1388A gene mutation group. All of 4 cases of A95G G1388A gene mutation showed a deficiency of G-6-PD activities. There were no significant differences in the incidence of acute bilirubin encephalopathy and the peak bilirubin concentration 72 hrs after birth between the G1388A and G1376T G1388A gene mutation groups. The incidence of acute bilirubin encephalopathy, the peak bilirubin concentration 72 hrs after birth and the risk of serum bilirubin >340 micromol/L in the G-6-PD mutation group were not different from the normal group., Conclusions: G1388A, G1376T and A95G are common G-6-PD gene mutations in Nanning, Guangxi. The false negative results may be received when the NBT method is used for diagnosis of G-6-PD deficiency. There are similar effects on the incidence of acute bilirubin encephalopathy and the peak bilirubin concentration 72 hrs after birth between different gene mutation groups. G-6-PD gene mutations alone may not contribute to the development of acute bilirubin encephalopathy and the changes of peak bilirubin concentration 72 hrs after birth and the risk of serum bilirubin >340 micromol/L.

Objective: To study the feasibility of genetic diagnosis for female carriers of human glucose-6-phosphate dehydrogenase (G6PD) deficiency by reverse transcriptase-PCR-denaturing gradient gel electrophoresis (RT-PCR-DGGE)., Methods: Blood samples were collected from suspected 54 female carriers of G6PD deficiency. Total RNAs of peripheral blood were prepared and reverse-transcripted into cDNA. Design of 6 primer pairs for DGGE was based on 17 mutation sites of G6PD cDNA described in the Chinese population. Mutations in the coding region of G6PD gene were screened and genotyped by combination of PCR-DGGE and DNA sequencing., Results: One case of 1024C/T, 20 cases of 1376G/T and 12 cases of 1388G/A were detected in the 54 samples. The total detection rate was 66.1% (33/54)., Conclusions: Heterozygous mutation rate in female carriers of G6PD deficiency detected by RT-PCR-DGGE is high. RT-PCR-DGGE is value of clinical diagnosis for G6PD-deficiency female carriers.

Objective: To detect gene mutations of children with glucose-6-phosphorate dehydrogenase (G-6-PD) deficiency and of carriers of G-6-PD deficiency gene with the technique of polymerase chain reaction and denatured gradient gel electrophoresis (PCR-DGGE), and to explore the value of the technique in the diagnosis of G-6-PD deficiency and G-6-PD deficiency gene carrying., Methods: cDNAs were harvested by reverse transcription method after RNAs had been extracted from peripheral blood of 43 children with G-6-PD deficiency and of their family members (36 lineages). Electrophoresis behaviors of the fragment from exons 11-12 of G-6-PD cDNA were detected with the technique of PCR-DGGE. Gene sequencing was then performed for the abnormal electrophoresis bands., Results: Abnormal electrophoresis bands were found in the 1304-1520 fragment of G-6-PD cDNA in 33 out of 36 family lineages. The G-6-PD/6-PGD ratio was below 1.00 in 9 mothers of patients. Three of them had the G-6-PD/6-PGD ratio lower than 0.50. The PCR-DGGE bands were the same in the 3 mothers. Gene sequencing showed double heterozygote in the 3 mothers, but the maternal carriers of G-6-PD deficiency gene who had normal G-6-PD/6-PGD ratio showed mono-heterozygote in gene sequencing. Three mutational sites were found in the 1304-1520 fragment, i.e., C1311TG1376T and G1388A. The electrophoresis behaviors were different among the 3 gene mutational sites., Conclusions: PCR-DGGE is a sensitive and reliable technique in the screening of gene mutations. It is useful in the diagnosis of G-6-PD deficiency, especially in the diagnosis of female G-6-PD deficiency gene carrying.

Objective: To molecularly analyze in Han and Li individuals of glucose-6-phosphate dehydrogenase deficiency in Hainan, China., Methods: The amplification refractory mutation system (ARMS) was employed to detect G1376T, G1388A and A95G mutations. The coding regions and flanking intronic regions from the second to the thirteenth exons of G6PD gene was analyzed by DNA sequencing to characterize the gene mutations in samples without G1376T, G1388A and A95G mutations., Results: Among 29 Han cases of G6PD deficiency, 11 had G1376T (37.9%), 2 G1388A (6.9%), 1 G1376T and G1388A (3.4%) and 1 G1376T and A95G (3.4%) were identified. Mutations of G1376T, G1388A, A95G and their complex accounted for 51.7% of G6PD deficiency in the Han individuals. Among 42 Li cases of G6PD deficiency, 25 had G1376T (59.5%), 6 G1388A (14.3%), 2 A95G (4.8%), 4 G1376T and G1388A (9.5%), 1 G1376T and A95G (2.4% )were identified. These mutations accounted for 90.5% of the Li individuals. Gene mutation of 18 cases (14 Han and 4 Li individuals) remained unknown. Sequencing results of the 18 samples indicated that one case had a single base of T deletion at nucleotide 636 or 637 in the 5th intron (IVS-5 636 or 637 T del) and two cases had C1311T with IVS-11 T93C mutation., Conclusion: G6PD G1376T and G1388A are the most common mutations in the populations of the Han and Li nationalities in Hainan. The IVS-5 636 or 637 T del mutation is first reported in Chinese, and the complex mutation of G1376T/A95G is first found in the Li nationality.

Objective: To detect the level of glucose-6-phosphate dehydrogenase (G-6-PD) mRNA expression in children with G-6-PD deficiency and their lineal family members in order to explore possible mechanisms of the disease at the transcriptional level., Methods: RNA was extracted from peripheral blood of 41 children with G-6-PD deficiency and of their lineal family members (29 father lineages and 40 mother lineages). cDNA was then harvested using the reverse transcription method. G-6-PD mRNA expression was detected by quantitative real-time PCR (QRT-PCR). The detection results of G-6-PD mRNA expression in three groups (Patient, Father lineage and Mother lineage) were compared using Statistical Software SPSS 10.0 system., Results: The mean G-6-PD mRNA value in the Patient, Father lineage and Mother lineage groups were 0.57 +/- 0.19, 0.74 +/- 0.21 and 0.67 +/- 0.21 respectively. The G-6-PD mRNA value in the Patient group was significantly lower than both Father lineage (t = -3.18, P < 0.01) and Mother lineage groups (t = -2.54, P < 0.05)., Conclusions: The expression of G-6-PD mRNA decreased in children with G-6-PD deficiency, suggesting that the pathogenesis of this disorder relates to the varying levels of gene transcription.

Objective: To explore new genotypes of glucose-6-phosphate dehydrogenase (G6PD) deficiency in Guangxi., Methods: G6PD mutations were identified by combination of denaturing high performance liquid chromatography (DHPLC), DNA sequencing and restriction endonuclease assay., Results: Three cases (10.0%) of G6PD Viangchan (871G-->A, 1311C-->T) and one case (3.3%) of G6PD Union (1360C -->T) were first identified in China mainland. G6PD Ganton (1376G-->T, 30.0%) was the commonest mutation, followed by Kaiping (1388G-->A, 26.7%), and Gaohe (95A-->G, 23.3%)., Conclusion: Besides G6PD Ganton, Kaiping and Gaohe, there are G6PD Viangchan and Union in Guangxi. G6PD Viangchan and Union is reported for the first time in China mainland.

Objective: To analyze the genotypes of glucose-6-phosphate dehydrogenase (G6PD) gene mutations, and investigate the incidence of G6PD deficiency and gene frequency in She Nationality (SN) in Fujian province., Methods: G6PD deficiency was screened by NBT method, G6PD mutations identified by amplification refractory mutation system (ARMS), mismatch-PCR with a restriction endonuclease, PCR-SSCP and DNA sequencing., Results: Among 1820 Qingshui SN and 764 Muyun SN, 101 and 104 samples were found to be abnormal, and the G6PD gene frequency accounted for 0.0607 and 0.1706 respectively. Among 54 pure ethnic descent SN samples in Muyun SN, 38 were identified as 1376G-->T, 10 as 1388G --> A and 6 as 95A-->G. The frequencies were 70.3%, 18.5% and 11.1% respectively. One case with both 1388G-->A and 95A-->G and another case with both 1376G-->T and 1388G-->A. Six 1024C-->T and one 392G -->T in Qingshui SN were identified., Conclusions: (1)G6PD nt1376G-->T and G6PD nt1388G-->A are two main variants in Muyun SN. Chinese ethnic groups share the similar G6PD gene variants, suggesting originated from the same ancestor. (2) 5 variants are found in SN: G6PD nt1376G-->T, nt1388G-->A, nt95A-->G, nt1024C-->T and nt392G-->T. (3) One with both nt1376G-->T and nt95A-->G mutations, another with both nt1376G-->T and nt1388G-->A mutations were found in SN. (4) nt95A-->G is another common G6PD gene variant in Muyun SN; nt1024C-->T is a common G6PD gene variant in Qingshui SN. (5)The gene frequency of G6PD in Qingshui SN is 0.0607, and is 0. 1706 in Muyun SN, these would facilitate the policy decision of C6PD deficiency prevention and treatment in these areas.

Unlabelled: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzymopathy. To date, about 126 mutations in the G6PD gene have been detected, among which 17 mutations were found in Chinese. The most common mutations are: 1376 G-->T and 1388 G-->A, both in exon 12; 95 A-->G in exon 2, which amounted to more than 50% of mutations representing various regions and ethnic groups in China. A large-scale screening and genotypic analysis was held in Shui people in Sandu of Guizhou. To investigate the incidence and the molecular basis of G6PD deficiency of Guizhou Shui people, NBT qualitative and G6PD/6PGD quantitative methods were used to detect G6PD deficiency in 1,090 Shui people from the general people belonging to Sandu of Guizhou. By means of mis-matched primers amplified the G6PD gene, the products were 234 bp, 280 bp and 345 bp in length, then restriction enzyme analysis was used to detect the most common Chinese G6PD mutations, 1376 G-->T, 1388 G-->A and 95 A-->G. The results showed that out of the 1,090 samples, 98 G6PD deficiency samples were found. The incidence of G6PD deficiency was 8.99%. 24 cases of 1376 G-->T, 12 cases of 1388 G-->A, 9 cases of 95 A-->G were detected. A sample with 1376 G-->T and 95 A-->G mutation was found in a girl. It was reported for the first time., In Conclusion: 1376 G-->T, 1388 G-->A, 95 A-->G mutations are the common G6PD mutations in Shui people in Sandu of Guizhou. The results indicates that different national minorities of Chinese may originated from a common ancestor.

Objective: Studying on G6PD polymorphism from Hakka population in Guangdong province., Methods: Identifying the variants of G6PD gene and determining the frequencies respectively with the use of amplified refractory mutation system(ARMS), polymerase chain reaction-single strand conformation polymorphism(PCR-SSCP) and ABI 3100 DNA sequencing technologies., Results: Mutations of G6PD gene in cDNA 1388 (G-->A), 1376 (G-->T), 95 (A-->G), 392 (G-->T), 1024 (C-->T), 1311 (C-->T) have been found., Conclusion: G6PD cDNA 1388 (G-->A), 1376 (G-->T), 95(A--> G), 392 (G-->T), 1024 (C-->T) and 1311 (C-->T) accompanied with intron 11 (93 T-->C) are the common mutations in Chinese population. cDNA 1388 (G-->A), cDNA 1376 (G-->T) are the most popular G6PD gene variants in Hakka population. In this study, no new type of G6PD gene mutation was found in the Hakkas of Guangdong.

Objective: To investigate the relationship between complex 1311 mutation of C-->T in exon 11 and 93 T-->C in intron 11 of G6PD gene and the G6PD deficiency., Methods: Using NBT paper strip method to screen and quantitative NBT method to confirm G6PD deficiency. PCR-SSCP technique was used to find the abnormal exon 11 and the amplification refractory mutation system (ARMS) to identify 1311 mutation, and DNA sequencing to identify the complex mutation at 1311 in exon 11 and 93 in intron 11., Results: Abnormal band in exon 11 was found in 12 cases. DNA sequencing showed that they were 1311 mutation together with 93 mutation., Conclusion: This complex mutation may be the cause of reduced activity of G6PD enzyme.

Objective: To identify glucose-6-phosphate dehydrogenase (G6PD) gene mutations in 23 patients with G6PD deficiency and to gain further understanding of the molecular and genetic background of G6PD gene in Yunnan province, China., Methods: The mutations located in exons 2-12 and in parts of introns of G6PD gene were analyzed by amplification refractory mutation system(ARMS), natural and mis-match primer PCR/restrict enzyme, polymerase chain reaction-single strand conformation polymorphism(PCR-SSCP ) analysis and automatic DNA sequencing., Results: Among these 23 samples, 5 different point mutations in G6PD gene were identified, and they constituted 5 genotypes. There were 7 Han and 3 Dai patients with G487A mutation, 7 cases with both intron 11 T93C and C1311T mutations, 4 cases with intron 5 636 or 637 T-->del mutation, 1 case with G871A mutation, and 1 case with G487A/T93C/C1311T mutation. Two haplotypes, 93C/1311T and 93C/1311T/487A were identified in Yunnan. A strong association was observed between C1311T and the Nla III restriction site produced by intron 11 T93C. The findings of the investigators on IVS-5 636 or 637T-->del in Chinese, on G871A in mainland of China, and on G487A in the Han people of Yunnan have not been reported previously., Conclusion: G6PD deficiency is very heterogenous in Yunnan; G487A is one of the common mutations in that province and may be of different origins. Possibly IVS-11 T93C mutation is of non-African origin. IVS-11 T93C and C1311T might jointly result in G6PD deficiency. The above data on G6PD gene mutation types could be useful for clinical diagnosis, prevention of G6PD deficiency, and researches in the origin and migration of minorities in Yunnan or other regions.

Objective: To elucidate the molecular basis of G6PD deficiency in the Han and Li nationalities in Hainan, China., Methods: Polymerase chain reaction and restriction enzyme digestion were used to screen the mutations 1388G-->A, 1360C-->T, 1024C-->T, 592C-->T, 517T-->C, 493A-->G, 487G-->A, 392G-->T and 95A-->G. Single strand conformation polymorphism analysis was used to screen the other mutations followed by DNA sequencing to characterize the mutations of the samples with abnormal SSCP bands., Results: Of the fifty-nine Han cases with G6PD deficiency, fourteen with 1388G-->A (23.7%), three with 871G-->A(5.1%), one with 835A-->T(1.7%), one with 517T-->C (1.7%), three with 392G-->T(5.1%), and four with 95A-->G(6.8%) were found. Of the thirty-two Li cases with G6PD deficiency, six with 1388G-->A(18.8%), three with 871G-->A(9.4%), and two with 95A-->G(6.3%) were found. A new mutation 835A-->G which causes the substitution of Ala for Thr at 279 in a Han case was identified and named as G6PD Haikou. The enzyme activity of the variant is about 10% of the normal and lower than the activity of the variant 835A-->T with about 40% of the normal. Analysis of the 3D model of human G6PD has revealed that the hydroxyl group of Thr at 279 is a group in maintaining the interaction of the G6PD subunits., Conclusion: The most common mutations of G6PD deficiency in Han and Li nationalities in Hainan are similar. Compared with the mutation spectrum of G6PD gene in the populations in other regions of China, the results indicate that some G6PD gene mutations are widespread in the populations of different regions in the southern part of China. The hydroxyl group of the Thr at 279 of human G6PD may be a necessary group for maintaining the interaction of the G6PD subunits and the enzyme activity.

Objective: To investigate the 1376G-->T mutation of G6PD gene in the cases of G6PD deficiency in the Han nationality and the Li nationality of Hainan, China., Methods: DNAs were extracted from the white blood cells of G6PD deficient cases by salt-out method. Allelic specific polymerase chain reaction was used to detect the 1376G-->T mutation., Results: Fifty-nine Han nationality cases and 32 Li nationality cases were analysed; the 1376G-->T mutation was found in 19 Han cases(32.2%) and 18 Li cases(56.2%)., Conclusion: 1376G-->T is a common mutation which causes G6PD deficiency in the Han nationality and the Li nationality in Hainan, China. Based on the phylogenetic tree of populations in China, these results indicate that the mutation might occur prior to the divergence of the Han and Li nationalities. It is of significance to study the mutations of G6PD gene in different nationalities in China for elucidating the origin, migration and evolution of the nationalities.

Objective: To analyze the types and distribution of glucose-6-phosphate dehydrogenase (G6PD) gene mutations in Yunnan province., Methods: By using the natural primers or mis-matched primers mediated polymerase chain reaction (PCR) followed by endonuclease digestion methods, six types of G6PD gene mutations, G1388A, G1376T, C1024T, G392T, C592T and A95G were in sixty patients with G6PD deficiency., Result: Six types of mutation were revealed: 28.3% (17 of 60) were G1388A, 11.7% (7 of 60) G1376T, 6.7% (4 of 60) C1024T, 1.7% (1 of 60) G392T, 1.7% (1 of 60) C592T, 1.7% (1 of 60) A95G and 48.3% (29 of 60) unidentified. C592T mutation is the first report in Yunnan people., Conclusions: The G6PD gene in Yunnan people is heterogeneous. Research of G6PD gene mutation types may provide some useful data for clinical diagnosis and prevention of G6PD deficiency, and for human genetic study.

Objective: To investigate the molecular and anthropologic features of glucose-6-phosphate dehydrogenase (G6PD) deficiency and the linkage between the Nla III polymorphic site within G6PD gene and three common Chinese G6PD mutations., Methods: By using denatured gradient gel electrophoresis (DGGE) and allele specific oligonucleotide (ASO) probe dot blot hybridization, Nla III polymorphic site at minus sign13 bp upstream exon 12 within G6PD gene were screened in 54 males with normal G6PD activity and 66 G6PD deficient males., Results: Thirty-two cases with cDNA1376 (G-->T), 21 with cDNA1388 (G-->A) and 13 with cDNA95 (A-->G) were detected in the 66 G6PD deficient males. Among 54 normal controls, T at -13 bp upstream exon 12 within G6PD gene were detected in 11 cases (20.4%, without Nla III polymorphic sites) while C in 43 cases (79.6%, with Nla III polymorphic sites). All 66 G6PD deficient males were linked with C at -13 bp upstream exon 12 (with Nla III polymorphic sites)., Conclusion: The G6PD mutations cDNA1376 (G-->T), cDNA1388 (G-->A) and cDNA95 (A-->G) were linked with Nla III polymorphism. This feature will play a role in studying the complicated manifestations and anthropology of G6PD deficiency.

Objective: In order to reveal the molecular evolution and ethnic origin and immigration, on the basis of G6PD G487A mutation, and estimate the incidence of the mutation in Bai, Dai, and Han people in Yunnan and Guangxi., Method: G6PD deficient individuals were studied by using PCR-RE, PCR-SSCP and DNA sequencing., Results: The G487A mutation in Bai and Dai people was identified for the first time and its relative incidences were 6/66 and 1/52, respectively. The individuals of G6PD G487A mutation showed the features of G6PD clinic type III. No G487A was found in Han people in Guangxi(0/46)., Conclusion: The G6PD G487A mutation revealed in some ethnic groups of China implies that different national minorities of Chinese may be origined from a common ancestor.

Objective: To study the types of glucose-6-phosphate dehydrogenase (G6PD) gene mutation in North China and to find the relationship between the mutations and population emigrants., Methods: DNA samples from 22 unrelated patients(21 male and 1 female) with G6PD deficiency from Tianjin area in North China were studied for the three common mutations by using both mismatched primers mediated polymerase chain reaction/ restriction enzyme analysis and dideoxy fingerprinting methods., Results: Eight patients(8/22, 36.4%) were found to have the mutation R459L(1376G-->T), and seven patients (7/22, 31.8%) were found to have the mutation R463H (1388G-->A). In three of (3/7) other seven patients (7/22), H32R(95A-->G) were detected; one patient (1/3) was identified to have H32R (95A-->G) mutation., Conclusion: The three common mutations in South China are also found in North China. However, most of the patients in Tianjin are also the later generations of Southern emigrants.

Objective: To detect three common G6PD gene point mutations, G1388A, G1376T and A95G, in Guangdong province and to estimate the approximate frequencies., Methods: Using the amplification refractory mutation system(ARMS) method to detect G1388A and G1376T and the newly developed ARMS to identify A95G mutation., Results: Ninety cases of G6PD deficient male were examined, and 42 of them were identified to be G1388A(46.7%), 14 G1376T(15.6%), and 12 A95G(13.3%). Totally 68 out of the 90 cases(75.6%) were of the three common mutations. The rest belong to rare or undefined mutations., Conclusion: ARMS method can be used in detecting common mutations of G6PD gene with the benefits of simple, fast, economic, and accurate.

Objective: To identify three common G6PD gene mutations in Yi minority in Yunnan province., Methods: G6PD gene mutation of 28 samples from G6PD deficient patients was detected by using amplification refractory mutation system(ARMS). Samples were collected from Yi minority in Yunnan province., Results: In 28 cases, 22 were identified to be three known mutations, including 12 of 1388(G-->A), 9 of 1376(G-->T) and 1 of 95(A-->G). The remaining 6 cases were undefined., Conclusions: This is the first report of G6PD gene G1376T and A95G mutations in Yi minority. The three common mutations account for 78.6% of the 28 cases.

Objective: To use amplification refractory mutation system(ARMS) method to detect the G6PD cDNA C1311T mutation, estimate its frequency in a normal south Chinese population, and investigate whether IVS-11 C93T mutation is the cause of certain G6PD deficient cases., Methods: DNA sequencing was used to confirm the C1311T and the IVS-11 C93T mutations. ARMS was set up to detect the C1311T and to estimate its frequency., Results: Three cases of C1311T mutation were found in 40 G6PD deficient samples. The optimal condition for ARMS was established. Using this method, 19 cases of C1311T were detected in 103 normal men, and the frequency of this polymorphic mutation was estimated to be 18.4% in southern Chinese population. Four cases with G6PD deficiency were demonstrated to be C at the IVS-11 93 position. At the same time, a case of IVS-11 93 C-->T was found in a normal man., Conclusions: ARMS is a simple, time-saving, and reliable method for detecting known G6PD gene point mutation. The frequency of C1311T in a normal south Chinese population is 18.4%. IVS-11 C93T might be another polymorphic site of the G6PD gene, and it is not the cause of enzyme deficiency in certain G6PD deficient cases.

Objective: Fifty-six G6PD-deficient male individuals of the Guangxi origin were investigated at the molecular level to understand the basis of G6PD deficiency., Methods: PCR amplified the exons 2,5,6,7,8, 10,11,12 and 13 of the G6PD gene respectively, combined with ASO dot blot hybridization and restriction enzymes digestion methods to screen 10 common Chinese G6PD mutations., Results: Six mutations were detected, namely cDNA1376(G-->T)(25.0%), cDNA1388(G-->A)(16.1%), cDNA95(A-->G)(19.8%), cDNA592 (C-->T)(7.1%), cDNA1024(C-->T)(1.8%) and cDNA392(G-->T) (1.8%). The unknown mutations accounted for 28.6%. cDNA1311(C-->T),cDNA1360(C-->T),cDNA835(A-->T), cDNA493(A-->G) and cDNA487(G-->A) were not discovered among the cases., Conclusion: The findings may provide some useful informations for physician scientists to get insight into the pathogenesis of G6PD deficiency so as to improve clinical diagnosis, prevention and cure of G6PD deficiency. The results may be of significance in anthropology and geneties as well.

Objective: To study the effect of G6PD deficiency on membrane phospholipid structures., Methods: Treatment of intact erythrocyte with phospholipase A2 was used for the membrane phospholipid asymmetry. Membrane phospholipid was determined by high performance liquid chromatography (HPLC)., Results: The contents of phosphatidylethanolamine (PE), phosphatidylcholine (PC), phosphatidylserine (PS) were 5.18 +/- 1.86, 1.51 +/- 0.58 and 5.70 +/- 1.91 mg/mg membrane protein, respectively, in G6PD deficient erythrocytes and were 5.59 +/- 1.16, 2.92 +/- 0.50 and 6.64 +/- 0.92 mg/mg membrane protein, respectively, in normal erythrocytes. In normal erythrocytes, 17.6% +/- 2.5% PE and 57.9% +/- 4.7% PC were present and no PS was found on the outer leaflet of the membrane. While in G6PD deficient ones, 30.2% +/- 5.7% PE, 53.8% +/- 7.4% PC and 27.2% +/- 5.8% PS were present on the outer leaflet., Conclusion: Membrane phospholipid changes in G6PD deficient erythrocytes might be one of the factors causing hemolysis.

Objective: To analyze G6PD gene mutation in 168 Cantonese G6PD deficient male infants., Methods: PCR products were amplified directly from dried blood spots on filter paper using 7 pairs of special primers followed by digestion with a restriction enzyme., Results: Of the 168 samples, 72(42.8%) were G6PD 1376 G-->T mutation, 35 (20.8%) were G6PD 1388 G-->A, 30(17.9%) were G6PD 95 AG, 6(3.6%) were G6PD 392 G-->T, and 3(1.8%) were G6PD 1024 C-->T. No G6PD 493 A-->G and 487 G-->A mutation were found, and 22(13.1%) were not defined., Conclusion: 1. The three G6PD mutations at 1376, 1388 and 95 were common in Cantonese. 2. Dried blood spots collected on filter paper provide an easy way of sample collection, storage and transport for the epidemiological study of inherited disease.

Objective: To analyze the G6PD gene mutations in several nationalities from Yunnan province., Methods: G6PD gene mutation of 29 samples from G6PD deficient patients were analyzed by using common and mis-matched primering PCR followed by restriction enzyme digestion. The samples were collected from Yunnan province among several nationalities., Results: In these 29 cases, 20 were of two known mutations, including 19 of 1388 (G-->A) and 1 of 1376 (G-->T) mutation. The 7 cases of national minorities, including 3 of Yi ([symbol: see text]), 2 of Bai ([symbol: see text]), 1 of Dai ([symbol: see text]), and 1 of Naxi ([symbol: see text]), were all 1388 mutations. No report of the G6PD gene 1388 mutation in these ethnic groups was published excepting for Dai., Conclusion: The mutation probably had been prevalent before the emergence of these national minorities. The approximate rate of G6PD gene mutations provided in Yunnan province may, in some respects, help to interpret the molecular evolution.

Objective: To explore the biochemical mechanism of G6PD deficiency., Methods: A fluorescence polarization technique with 1,6-dipheny 1,1,3,5-heratciene (DPH) as a fluorescent probe was used to determine the fluidity of erythrocyte membrane lipid from G6PD deficient patients and normal controls. The polarization of fluorescence was measured between 0 degrees and 90 degrees., Results: The fluorescence polarization parameter (rho) was high in G6PD deficient erythrocytes as compared with normal controls, indicating a decrease in erythrocyte membrane lipid fluidity. The determination of lipoperoxidation of erythrocyte membrane by fluorescence spectrometry also showed that the content of MDA was significantly higher in G6PD deficient erythrocytes than in normal controls. The percentage of unsaturated fatty acid C18:1 and C18:2 in total phosphalipids and phosphatidyl serine was decreased in G6PD deficient erythrocytes., Conclusion: The decrease of erythrocyte membrane lipid fluidity is related to the lipid peroxidant, and the oxidative damage of erythrocyte membrane phosphalipids in G6PD deficient patients might be an important factor of hemolysis.

Hereditary glucose-6-phosphate dehydrogenase (G6PD) in red blood cell was one of the most common genetic diseases in South China. The research data of G6PD gene showed that at least 6 point mutations were responsible for various G6PD variants in Chinese. For developing a rapid, sensitive, and effective method to detect point mutation, we applied single-strand conformation (SSCP) analysis for detection of mutation in exon 2 of G6PD gene of 20 cases of G6PD variants. Four of them were found that mobility shift band in one of two single strands DNA is slower than other individuals. PCR direct sequencing for these 4 samples were given a base substitution (T to D) at nucleotide 95 of cDNA. The results indicated that this technique is very simple, sensitive, and useful over other methods of detecting point mutation. The experimental conditions of PCR-SSCP and features of this mutation were discussed at the same time.