OBJECTIVES: To observe and analyze the confirmed cases of paternity testing, and to explore the mutation rules of STR loci. METHODS: The mutant STR loci were screened from 20 723 confirmed cases of paternity testing by Goldeneye 20A system．The mutation rates, and the sources, fragment length, steps and increased or decreased repeat sequences of mutant alleles were counted for the analysis of the characteristics of mutation-related factors. RESULTS: A total of 548 mutations were found on 19 STR loci, and 557 mutation events were observed. The loci mutation rate was 0.07‰-2.23‰. The ratio of paternal to maternal mutant events was 3.06:1. One step mutation was the main mutation, and the number of the increased repeat sequences was almost the same as the decreased repeat sequences. The repeat sequences were more likely to decrease in two steps mutation and above. Mutation mainly occurred in the medium allele, and the number of the increased repeat sequences was almost the same as the decreased repeat sequences. In long allele mutations, the decreased repeat sequences were significantly more than the increased repeat sequences. The number of the increased repeat sequences was almost the same as the decreased repeat sequences in paternal mutation, while the decreased repeat sequences were more than the increased in maternal mutation. CONCLUSIONS There are significant differences in the mutation rate of each locus. When one or two loci do not conform to the genetic law, other detection system should be added, and PI value should be calculated combined with the information of the mutate STR loci in order to further clarify the identification opinions.
Alleles ; DNA Mutational Analysis/methods* ; Family ; Genetic Loci ; Humans ; Male ; Microsatellite Repeats ; Mutation ; Mutation Rate ; Paternity

OBJECTIVETo explore the necessity of large-scale screening of mitochondria DNA (mtDNA) A1555G mutation for prevention of aminoglycoside antibiotic induced deafness in newborns.

METHODSOne thousand blood filter samples were collected from neonates born in July 2008 in Shenzhen. DNA was extracted with Chelex-100 Resin and amplified by PCR. The mtDNA A1555G mutation was determined by denaturing high-performance liquid chromatography(DHPLC) for PCR products. The positive frequency was calculated.

RESULTSThe mitochondrial DNA A1555G mutation was detected in 2 cases of 1000 neonates. The frequency of mutation was 0.2%.

CONCLUSIONThere is a high frequency of mtDNA A1555G mutation in neonates, the large-scale screening of mtDNAA1555G mutation in newborns might detect the individuals sensitive to aminoglycoside antibiotic, which is helpful to guide a rational medication for newborns and the maternal relatives at high-risk. Furthermore, it might be useful to prevent aminoglycoside antibiotic induced deafness.

Base Sequence ; DNA Mutational Analysis ; methods ; DNA, Mitochondrial ; genetics ; Female ; Humans ; Infant, Newborn ; Male ; Polymerase Chain Reaction

High resolution melting (HRM), based on melting curve analysis, requires not only saturating dyes that fluoresce in the presence of double-stranded DNA, but also higher resolution detection equipment. The melting curve is a novel method for sequence matching, genotyping and mutation scanning. The technology is simple, accurate, rapid, closed-tube, low-cost, and high-throughput, which make it gain more and more applications. This review article presents the basic principles, key factors and both the advantage and limitations of HRM. The potential application is discussed in the study of molecular identity of traditional Chinese medicine.
DNA Mutational Analysis ; methods ; Drugs, Chinese Herbal ; classification ; Genotyping Techniques ; methods ; Medicine, Chinese Traditional ; Nucleic Acid Denaturation

OBJECTIVETo study the prediction performance evaluation with five kinds of bioinformatics software (SIFT, PolyPhen2, MutationTaster, Provean, MutationAssessor).

METHODSFrom own database for genetic mutations collected over the past five years, Chinese literature database, Human Gene Mutation Database, and dbSNP, 121 missense mutations confirmed by functional studies, and 121 missense mutations suspected to be pathogenic by pedigree analysis were used as positive gold standard, while 242 missense mutations with minor allele frequency (MAF)>5% in dominant hereditary diseases were used as negative gold standard. The selected mutations were predicted with the five software. Based on the results, the performance of the five software was evaluated for their sensitivity, specificity, positive predict value, false positive rate, negative predict value, false negative rate, false discovery rate, accuracy, and receiver operating characteristic curve (ROC).

RESULTSIn terms of sensitivity, negative predictive value and false negative rate, the rank was MutationTaster, PolyPhen2, Provean, SIFT, and MutationAssessor. For specificity and false positive rate, the rank was MutationTaster, Provean, MutationAssessor, SIFT, and PolyPhen2. For positive predict value and false discovery rate, the rank was MutationTaster, Provean, MutationAssessor, PolyPhen2, and SIFT. For area under the ROC curve (AUC) and accuracy, the rank was MutationTaster, Provean, PolyPhen2, MutationAssessor, and SIFT.

CONCLUSIONThe prediction performance of software may be different when using different parameters. Among the five software, MutationTaster has the best prediction performance.

Computational Biology ; methods ; DNA Mutational Analysis ; methods ; Gene Frequency ; Humans ; Mutation, Missense ; genetics ; Polymorphism, Single Nucleotide ; genetics ; Reproducibility of Results ; Software

DNA Mutational Analysis ; methods ; Ellis-Van Creveld Syndrome ; diagnosis ; Female ; Humans ; Male ; Pregnancy ; Prenatal Diagnosis ; methods ; Proteins ; genetics ; Ultrasonography, Prenatal

OBJECTIVETo establish a comprehensive and simple assay using denaturing high performance liquid chromatography (DHPLC) for the diagnosis of most common mutations and deletions of α-thalassemia gene in Southeast Asians and Southern Chinese.

METHODSThis assay has included a duplex polymerase chain reaction (PCR) followed by DHPLC analysis. An improved PCR was also performed followed by DHPLC analysis. With this assay, a blinded study of 160 samples was screened for three common mutations and three common deletions.

RESULTSThe duplex PCR-DHPLC combined with the improved PCR-DHPLC analysis has detected all mutations and the wild-type allele. The results were consistent with those by the original methods.

CONCLUSIONThis molecular assay may be used for the diagnosis of α-thalassemia patients from this geographical region. The method is accurate, rapid, semi-automatic and cost-effective, which makes it suitable for large-scale screening.

Chromatography, High Pressure Liquid ; methods ; DNA Mutational Analysis ; methods ; Gene Order ; Genotype ; Humans ; alpha-Globins ; genetics ; alpha-Thalassemia ; diagnosis ; genetics

OBJECTIVE: Using simultaneous multi-gene mutation screening to survey the molecular epidemiological basis of 355 patients with nonsyndromic hearing loss of Inner Mongolia Autonomous region, we can identify the causes of their deafness,and verify the new method for simultaneous multi-gene mutation screening. METHOD: Three hundred and fifty-five patients with severe non-syndromic deafness from Inner Mongolia Autonomous regior were included in the study. The SNPscan technology was used for screening the 115 spots mutations in three common deafness-related genes(GJB2, SLC26A4, MT-12S rRNA) of patients with nonsyndromic hearing loss of Inner Mongolia Autonomous region. RESULT: In 355 patients, there were 89 cases of deafness caused by mutatior (25.07%). 53 patients with the GJB2 mutations were found(14.93%), including 24 cases of homozygous mutations (6.76%), 29 patients (8.17%) of compound heterozygous mutations, and 3 cases (0.85%) of single heterozygous mutations. 33 patients with the SLC26A4 mutations were found (9.30%), including 15 cases of homozygous mutations (4.23%),18 patients (5.07%) of compound heterozygous mutations, and 5 cases (1.41%) of single heterozygous mutations. mtDNA12S rRNA A1555G mutation was found in 6 patients (1.69%). mtDNA12S rRNA 1494C>T mutation was not found. CONCLUSION SNPscan technology allows accurate, rapid and cost-effective diagnostic screening in patients with hearing loss for etiology investigation. The SNPscan technology can serve as a good diagnostic tool for large-scale genetic testing for hereditary deafness and should be widely applied.
China ; Connexins ; DNA Mutational Analysis ; methods ; Deafness ; genetics ; Genetic Testing ; methods ; Heterozygote ; Homozygote ; Humans ; Mutation ; Polymorphism, Single Nucleotide ; RNA, Ribosomal

OBJECTIVE: To explore and analyze the mutations of 15 Short Tandem Repeat (STR) loci using Identifiler system in paternity identification. METHODS: 2712 cases of paternity testing were carried out using Identifiler PCR Amplification Kit. RESULTS: Of the 2362 paternity testing cases, mutations of single locus were observed in 51 cases. The mutation loci included D8S1179, D21S11, D7S820, CSF1PO, D3S1358, D13S317, D16S539, D2S1338, D19S433, vWA, D18S51, D5S818 and FGA, with the D21S11 locus having a highest mutation rate (0.369%). Thirty-six of the STR mutations were from paternal source, 7 from maternal source, and the rest (9) were undeterminable. The mutation rates at D21S11 were highest (0.369%). CONCLUSION Mutations of STR loci are relatively common in human genome. Therefore, retesting of additional relatively stable STR loci with lower mutation rates is necessary when one or two loci exclusions are encountered in paternity testing.
Alleles ; DNA Fingerprinting/methods* ; DNA Mutational Analysis/methods* ; Forensic Medicine/methods* ; Gene Frequency ; Genetics, Population ; Humans ; Mutation ; Paternity ; Polymerase Chain Reaction/methods* ; Tandem Repeat Sequences/genetics*

OBJECTIVETo assess the accuracy and reliability of the nest-PCR-sequence specific primer(SSP) method in HLA-A site genotyping of single blastomeres retrieved from human pre-implantation embryos.

METHODSBy nest PCR on HLA-A exon 2, the success rate of first-round amplification was estimated for single blastomeres. Based on the first-round amplification, the HLA-A genotype of every single blastomeres was analyzed by commercially available PCR-SSP kits.

RESULTSThe amplification of HLA-A exon 2 were performed to 120 blasotmeres retrieved from in vitro fertilization(IVF) surplus embryos donated by 10 couples. The average success rate of family 1-5 and 6-10 was 78.2%(43/55) and 93.8%(61/65), respectively. And 86.7%(104/120) in total. Eighty blastomeres were further tested by nest-PCR-SSP, among which 11 blastomeres failed to HLA-A exon 2 amplification and then failed to genotyping while the other 69 blastomeres succeed in HLA-A exon 2 amplification and succeed in genotyping. Except for 6 blastomeres that were uncertain for allele lost because of parents' homozygosity, the left 63 blastomeres had accurate HLA genotyping. Among these 63 blastomeres, 59 blastomeres had genotypes confirmed from their parents(93.6%), 3 blastomeres lost one of parents' alleles(4.8%), and only one blastomere had two more than parents' alleles(1.6%).

CONCLUSIONThe above research results indicated that based on the successful first round amplification of single blastomeres, nest-PCR-SSP strategy offers a convenient and reliable option for HLA genotyping on single blastomeres, which is a key process in pre-selecting HLA-identical sibling for allogeneic cord blood cell transplantation.

Base Sequence ; Blastomeres ; metabolism ; DNA ; analysis ; DNA Fingerprinting ; methods ; DNA Mutational Analysis ; Female ; HLA Antigens ; analysis ; HLA-A Antigens ; analysis ; genetics ; Histocompatibility Antigens Class I ; analysis ; genetics ; Humans ; Male ; Polymerase Chain Reaction ; methods ; Single Person

This study was aimed to investigate the suitability of FVIII gene BclI (intron 18)and HindIII (intron 19) site polymorphism for genetic diagnosis of patients with hemophilia A (HA) and their families, and for detection of carriers. The FVIII gene bclI (intron 18) and HindIII (intron 19) site polymorphism on the X chromosome of 8 patients with HA and 45 families members were detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), the pedigree of patients was drawn by means of PediDraw software online provided by China Genetic Counseling Network. The results indicated that combination detection of BclI and HindIII sites could provide the diagnosis information for 5 out of 8 HA families with diagnostic ratio of 62.57%, especially 2 HA families were accompanied by mutation of 2 sites. Besides, the definite diagnosis for 6 out of 11 suspicious carriers in 8 families could be made with diagnostic ratio of 54.5%. It is concluded that the combination detection of BclI and HindIII sites for analysis of HA patient family can elevate the diagnostic rate of HA patients and carriers.
DNA Mutational Analysis ; Factor VIII ; genetics ; Female ; Hemophilia A ; diagnosis ; genetics ; Humans ; Introns ; Male ; Pedigree ; Polymerase Chain Reaction ; methods ; Polymorphism, Genetic