ObjectivesTo investigate the clinical value of the melting curve analysis-based PCR assay for the clinical genetic diagnosis and prenatal diagnosis of β-thalassemia.Methods A total of 451 peripheral blood samples,including 372 cases with β-thalassemia phenotypes and 79 cases without β-thalassemia phenotypes,were collected by Liuzhou Municipal Maternity and Child Healthcare Hospital between January 2011 and August 2011.Moreover,another 84 cases,including 16 fetal villi samples (10 - 13 weeks),64 amniotic fluid samples (16 -24 weeks ) and 4 umbilical cord blood samples (above 17 weeks),whose parents were β-thalassemia carriers,were also collected for this assay between June 2011 and September 2011.A double-blind test was done to compare the detection reliability of the melting curve analysis-based assay (24 β-thalassemia mutations can be detected) with PCR-RDB probe assay (17 β-thalassemia mutations can be detected ) and DNA sequencing using these samples.The wildtype,mutant and total concordance rates of the genotyping results were calculated separately among the melting curve analysis based assay,PCR-RDB probe assay and DNA sequencing.Results Among the 451 peripheral blood samples,thirteen mutations and nineteen genotypes were obtained by using melting curve analysis-based assay.447 samples had the same detection results and 4 samples had different detection results by comparing melting curve analysis-based assay with PCR-RDB probe assay,thus,the concordance rate of the sample detection result was 99.1％ (447/451),and the concordance rate of the allele detection result was 99.6％ (898/902).DNA sequencing results of the 4 samples showed that 3 samples had the same genotyping result with melting curve analysis-based assay,and 1 sample had the same genotyping result with PCR-RDB probe assay.A rare β-globin mutation which was not included by melting curve analysis-based assay was not detected.Thus,the genotypes of 450 samples were detected accurately by melting curve analysis-based assay,and the concordance rate of the sample detection between the melting curve assay and DNA sequencing assay was 99.8％ (450/451).Among 84 fetal villi,amniotic fluid,and umbilical cord blood samples,8 mutation types and 18 genotypes were obtained by using melting curve analysis-based assay.All the samples have the same detection results by comparing melting curve analysis-based assay with PCR-RDB probe assay and DNA sequencing,so the concordance rate of the genotyping results was 100％ among the melting curve analysis-based assay,PCR-RDB probe assay and DNA sequencing.Conclusions The melting curve analysis-based PCR assay can detect multiple unknown samples simultaneously,and detect multiple mutations accurately.It is very useful for the genetic diagnosis and prenatal diagnosis of β-thalassemia.

Frequency-domain electricity properties of four objects, including bullfrog skin, bullfrog muscle, triply distilled water and 0.9% NaCl, were tested in the range of 100Hz-10MHz using home-made electrode and measuring system. The experimental results showed that the resistance of 0.9% NaCl decreased dramatically, that the amplitude frequency characteristics of bullfrog's muscle and skin were similar, but that of triply distilled water did not change significantly. The frequency dependence of 0.9% NaCl showed that the electrode had great influence on the measuring system, so a new equivalent circuit model based on the electrode system was needed. These findings suggest that the new five-parameter equivalent circuit model, which embodies considerations on the interaction between electrodes and tissues, is a reasonable equivalent circuit for studying the electrical characteristics of biological materials.
Animals ; Electric Capacitance ; Electric Impedance ; Electricity ; Electrodes ; Muscle, Skeletal ; physiology ; Rana esculenta ; Skin ; Sodium Chloride

Objective To study genotypes in nonsyndromic hearing loss (NSHL ) patients from Guangxi Zhuang Autonomous Region hearing speech rehabilitation center using DNA microarray in combination with Sanger sequencing .Methods Deaf patients received routine physical and otorhinolaryngoloical examinations as well as pure tone autiometry .Brainstem auditory evoked potential test was performed in uncooperative children .Blood samples were obtained from a total of 136 patients ,male 81 ,female 55 ,age from one year five month to seventeen ,having nonsyndromic hearing loss .Genomic DNA was extracted and then 9 hot mutation spots in 4 susceptibility genes were detected by DNA microarray .GJB2 and SLC26A was further detected by Sanger sequencing in the patients with negative results and heterozygotes .Results Among the 136 patients with nonsyndromic hearing loss ,20 cases were positive for GJB2 gene ,SLC26A4 gene or mitochondrial 12SrRNA gene mutations .There were 14 .71% (20/136)patients were positive for hot mutation spots in the deafness related genes ,25% (34/136)patients carried muta‐tions of deafness related genes using DNA microarray in combination with Sanger sequencing .Six SLC26A4 rare mutations (c .259G> T ,c .754C> T ,c .1229C> T ,c .1548_1549insC ,c .1705+5A>G and c .2086C> T) were de‐tected by Sanger sequencing .c .235delC was the most common mutation in GJB2 gene .c .919-2A>G ,c .754C> T and c .1229C> T were the common mutations in SLC26A4 gene .The mutation rate of GJB2 and SLC26A4 was 38 . 24% .and 58 .82% ,respectively .Conclusion Prevalent deafness-associated gene mutations in the nine loci studied were less frequently detected in nonsyndromic hearing loss patients from Guangxi Zhuang Autonomous Region hear‐ing speech rehabilitation center .It can improve the detection rate of deafness gene mutations by using gene microar‐ray in combination with Sanger sequencing .GJB2 and SLC26A4 are the common causative genes .

OBJECTIVETo screen for carriers of SMN1 gene mutation, which underlies spinal muscular atrophy (SMA), in 4931 pregnant women from Liuzhou region of Guangxi, and to determine the carrier rate.

METHODSCombined denaturing high-performance liquid chromatography (DHPLC) and multiple PCR techniques were used to detect the copy number of SMN1 gene. The carrier frequency was calculated. The spouse of the carrier was also screened, and prenatal diagnosis was provided to the couples who were both positive.

RESULTSAmong the 4931 pregnant women, 61 were found to harbor only one copy of the SMN1 gene, which yielded a carrier rate of 1.2%. Subsequent testing has identified 1 fetus carrying homozygous deletions of the SMN1 gene.

CONCLUSIONThe carrier rate of SMA mutation in Liuzhou region is slightly lower than that of other regions of southern China. DHPLC can effectively screen the carriers of SMA mutation and provide a basis for genetic counseling and prenatal diagnosis.

OBJECTIVE: To determine the composition and distribution of beta-thalassemia-associated genotypes in Liuzhou area of Guangxi, China. METHODS: From January to December 2017, 13 847 individuals who came for premarital examination, maternity examination or health check were recruited with informed consent. The subjects were analyzed by reverse dot blotting (RDB) for 17 common beta-thalassemia-associated variants among the Chinese population. Individuals with inconsistent results by blood test, electrophoresis, and RDB were subjected to Sanger sequencing to detect rare variants of the beta globin gene. RESULTS: In total 2098 individuals were found to harbor beta-thalassemia-associated variants, which included 2075 heterozygotes (98.90%), 12 compound heterozygotes (0.57%) and 11 homozygotes (0.52%). CD41-42 (48.43%) and CD17 (31.45%) were the most common variants. Three hundred and thirty eight-individuals were found to also carry heterozygous variants of the alpha globin gene, with the most common types being --SEA/aa, -a3.7/aa, aCSa/aa, -a4.2/aa. Through Sanger sequencing, rare genotypes such as beta-32/betaN, betaCD41-42/betaIVS-II-5 and betaCD30/betaN were detected. CONCLUSION Liuzhou area has a high incidence of beta-thalassemia, but with a complex variant spectrum and clinical phenotypes different from other regions. Genetic counseling and prenatal diagnosis for the carrier population is crucial for the reduction of the related birth defects. Our result may provide valuable information for the prevention and control of beta-thalassemia in this area.
China ; Female ; Genetic Counseling ; Genetic Variation ; Genotype ; Humans ; Mutation ; Pregnancy ; Prenatal Diagnosis ; alpha-Globins ; genetics ; beta-Globins ; genetics ; beta-Thalassemia ; diagnosis ; genetics

OBJECTIVETo evaluate the clinical value of multicolor melting curve analysis(MMCA) for detecting genetic mutations in G6PD deficiency.

METHODSA total of 402 peripheral blood samples(256 males and 146 females) were collected from suspected patients or their relatives at the Prenatal Diagnosis Center of Liuzhou Maternal and Child Health Hospital between March 2012 and May 2012. The samples were screened by G6PD/6PGD quantitative ratio testing. The reliability of the assay was evaluated by multiplex probe melting curve assay(which can detect 16 G6PD mutations) and DNA sequencing through a double blind study.

RESULTSOne hundred seventy cases with G6PD/6PGD ratio < 1.0 and 232 cases with G6PD/6PGD ratio ≥ 1.0 were detected by the enzymological method. DNA sequencing has identified 182 wild type samples, 151 hemizygous mutation samples, 5 female homozygous mutation samples, 54 female heterozygous mutation samples and 10 female double heterozygous mutation samples. Multicolor melting curve analysis has detected 185 wild type samples, 148 hemizygous mutation samples, 5 female homozygous mutation samples, 55 female heterozygous mutation samples and 9 female double heterozygous mutation samples. The specificity and sensitivity of G6PD gene mutation detection by multicolor melting curve analysis were 100%(182/182) and 98.6%(217/220), respectively. The positive predictive value and negative predictive value were 99.5%(216/217) and 98.4%(182/185), respectively, and the Youden's index was 0.986. The concordance rate of the sample detection between the melting curve assay and DNA sequencing was 99.0%(398/402). Twenty-one different genotypes were detected by the multicolor melting curve analysis and 24 different genotypes were detected by DNA sequencing. Four samples containing mutations(c.196T>A or c.406C>T) were not detected by multicolor melting curve analysis, which can be attributed to different technical settings of the two methods.

CONCLUSIONMulticolor melting curve analysis for G6PD gene mutation detection is a simple, rapid, sensitive and specific method, which can be used for clinical diagnosis of G6PD deficiency.

Adolescent ; Adult ; Child ; Child, Preschool ; Female ; Glucosephosphate Dehydrogenase ; genetics ; Humans ; Infant ; Infant, Newborn ; Male ; Mutation ; Polymerase Chain Reaction ; methods ; Sequence Analysis, DNA

BACKGROUND:Digital three-dimensional reconstruction technology is gradually applied to orthopedic diseases with the advantages of visualization,accuracy and non-invasiveness,but there is less evidence-based support for its use in artificial hip hemiarthroplasty for intertrochanteric fractures of the femur in the elderly. OBJECTIVE:To investigate the application value and economic effects of digital three-dimensional reconstruction techniques in artificial hip hemiarthroplasty of intertrochanteric fractures of the femur in the elderly. METHODS:One hundred and thirty elderly patients with intertrochanteric femur fractures admitted to Zunyi First People's Hospital from January 2019 to December 2022 were selected and randomly divided into a control group(n=65)and an observation group(n=65).Artificial hip hemiarthroplasty was performed in both groups.The control group adopted the film template measurement method for manual preoperative planning while the observation group adopted a digital three-dimensional reconstruction technique.Preoperative planning and intraoperative actual application of prosthesis compliance rate,fibrinogen,D-dimer,bilateral femoral eccentric distance difference,bilateral lower limb length difference,Harris hip function score,visual analog scale score,excellent and good rate of hip function,complications,and hospitalization cost were observed in both groups. RESULTS AND CONCLUSION:(1)The proportion of acetabular side and femoral side prosthesis in grade 0(fully compliant)was higher in the observation group than that in the control group(P<0.05).(2)Fibrinogen and D-dimer levels in the observation group were lower than those in the control group 3 days after surgery(P<0.05).(3)The difference in bilateral femoral eccentric distance and the difference in bilateral lower limb length in the observation group were smaller than those in the control group immediately after surgery(P<0.05).The differences in Harris and visual analog scale scores were not significantly different between the two groups preoperatively,6 and 12 months postoperatively(P>0.05).There was no significant difference in excellent and good rate of hip function between the two groups 12 months postoperatively(P>0.05).(4)There was no significant difference in the complication rate between the two groups(P>0.05).The hospitalization cost of the observation group was higher than that of the control group(P<0.05).(5)It is indicated that digital three-dimensional reconstruction technology applied in artificial hip hemiarthroplasty of intertrochanteric femoral fracture in the elderly can not only accurately determine the prosthesis type before surgery,but also accurately reconstruct the bilateral lower limbs offline,but its hospitalization cost is high.