RESUMO
OBJECTIVE@#To establish a simple, fast and economical technique for multiplex-typing SNPs and to explore its potential forensic application.@*METHODS@#Five Y-SNP loci (IMS-JST164520, IMS-JST021354, IMS-JST003305, M119 and M134) were selected and the allele specific primers of each locus were designed with the universal reporter primers tailed at their 5' end. Alleles of these loci were amplified first by allele specific primers, then amplified by universal reporter primers tagged by fluorescent dye.@*RESULTS@#A fluorescent-multiplex PCR system of the five Y-SNP loci was established. The typing results showed that two different colors of product peaks denoted two different alleles of a SNP locus, and the fragment sizes of alleles among different SNP loci were different. The haplotype diversity of these five loci was estimated to be 0.8655 in Wuhan Han population.@*CONCLUSION@#The multiplex-typing SNPs based on the universal reporter primers is a simple, fast, and economical technique, and may have good application value in forensic medicine.
Assuntos
Feminino , Humanos , Masculino , Alelos , Povo Asiático/genética , China/etnologia , Cromossomos Humanos Y/genética , Primers do DNA , Genética Forense , Frequência do Gene , Marcadores Genéticos , Genética Populacional , Haplótipos , Técnicas de Amplificação de Ácido Nucleico/métodos , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
OBJECTIVE@#To study the application of PCR-SSCP in forensic mtDNA typing.@*METHODS@#Primers flanking the mtDNA HV-I and HV-II regions were designed. By PCR-SSCP techniques, 70 family trios and 140 unrelated Wuhan Han individuals were investigated and analyzed.@*RESULTS@#In 70 family trios, the SSCP profiles in region HV-I and HV-II of children were not same to that of their fathers in 98.57% and 97.13% respectively but were identical with their mothers. In 140 unrelated Wuhan Han individuals, 21 haplotypes were found in HVI, GD = 0.9556; 16 haplotypes were found in HVII, GD = 0.9356.@*CONCLUSION@#PCR-SSCP technique may be useful in forensic mtDNA typing, especially for screening the suspects.
Assuntos
Humanos , Impressões Digitais de DNA/métodos , Primers do DNA , DNA Mitocondrial/genética , Genética Forense/métodos , Haplótipos , Linhagem , Reação em Cadeia da Polimerase/métodos , Polimorfismo Conformacional de Fita Simples , Análise de Sequência de DNARESUMO
DNA methylation is a post-replication modification that is predominantly found in cytosines of the dinucleotide sequence CpG. Epigenetic information is stored in the distribution of the modified base 5-methylcytosine. DNA methylation profiles represent a more chemically and biologically stable source of molecular diagnostic information than RNA or most proteins. Recent advances attest to the great promise of DNA methylation markers as powerful future tools in the clinic. In the past decade, DNA methylation analysis has been revolutionized by two technological advances--bisulphite modification of DNA and methylation-specific polymerase chain reaction (MSP). The methylation pattern of human genome is space-time specific, sex-specific, parent-of-origin specific and disease specific, providing us an alternative way to solve forensic problems.
Assuntos
Humanos , Sequência de Bases , Ilhas de CpG/genética , DNA/sangue , Impressões Digitais de DNA/métodos , Metilação de DNA , Epigênese Genética , Medicina Legal/métodos , Marcadores Genéticos , Genoma Humano , Paternidade , Reação em Cadeia da Polimerase/métodosRESUMO
OBJECTIVE@#To establish a new method for single nucleotide polymorphism (SNP) typing based on allele specific PCR: fragment length discrepant allele specific PCR (FLDAS-PCR), and study the influence on specific extension by introducing a mismatch at the third or fourth 3'-terminal base of allele specific primers.@*METHODS@#For SNP loci rs759117 and rs760887, two allele specific forward primers, with different length and a mismatch introduced at the third or fourth 3'-terminal base, and a public reverse primer were designed for SNP typing. The genotyping of SNP was determined by the two allele specific fragments different in size after polyacrylamide gel and silver staining.@*RESULTS@#The different homozygote genotypes comprised a single band with different size respectively, and the heterozygote genotypes comprised two bands. Typing results were completely consistent with those by direct sequencing. Non-specific primer extension was decreased remarkably after introducing a mismatch at the third or fourth 3'-terminal base of allele specific primers, and the stringency of PCR reaction was cut down.@*CONCLUSION@#FLDAS-PCR is a simple, rapid and efficient new method for SNP typing. During FLDAS-PCR, specific primers with a mismatch at the third or fourth 3'-terminal base have more power to identify two alleles.
Assuntos
Humanos , Alelos , Pareamento Incorreto de Bases/genética , DNA/genética , Primers do DNA , Eletroforese em Gel de Poliacrilamida , Genótipo , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo de Nucleotídeo Único , Sensibilidade e Especificidade , Análise de Sequência de DNA/métodosRESUMO
<p><b>OBJECTIVE</b>To establish a novel method for the multiplex analysis of the methylation and single nucleotide polymorphism (SNP).</p><p><b>METHODS</b>The imprinted SNP rs220028 was chosen as a model. Genomic DNA, after being digested with methylation sensitive restriction enzyme, were typed by mutagenically separated PCR (MS-PCR). The polymorphism of restriction site was excluded by PCR-RFLP.</p><p><b>RESULTS</b>By post-digestion MS-PCR, the methylated allele was detected selectively, the maternal origin of which was confirmed by pedigree analysis; A=0.5085, G=0.4915,PIC=0.3749.</p><p><b>CONCLUSION</b>The multiplex analysis of methylation markers and SNP can be achieved by post-digestion MS-PCR. The imprinted SNP locus rs220028 is a potentially useful marker in screening Prader-Willi/Angelman syndrome.</p>
Assuntos
Humanos , Metilação de DNA , Enzimas de Restrição do DNA , Metabolismo , Marcadores Genéticos , Genética , Reação em Cadeia da Polimerase , Métodos , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Nowadays, the injury in human mitochondrial DNA (mtDNA) is well known to accumulate in various tissues with age. It's significant to further investigate and then apply it to estimation of the age at parenchymas.
Assuntos
Humanos , Envelhecimento/fisiologia , Pareamento Incorreto de Bases/genética , Dano ao DNA/fisiologia , Fragmentação do DNA/genética , DNA Mitocondrial/fisiologia , Deleção de Genes , Reação em Cadeia da PolimeraseRESUMO
This article review the application of chi-square test of various data handling methods and exact test in Hardy-Weinberg equilibrium testing of human genetic marker in population genetics. The importance of HWE-exact test in multiallelic system was emphasized, especially in the study of forensic VNTR and STR typing.
Assuntos
Humanos , Alelos , Distribuição de Qui-Quadrado , Medicina Legal , Frequência do Gene , Genética Populacional/métodos , Genótipo , Funções Verossimilhança , Modelos Genéticos , Modelos EstatísticosRESUMO
OBJECTIVE@#PGM1 genotyping by PCR-SSCP analysis.@*METHODS@#Amplified genome DNA from 156 unrelated Han individuals living in Wuhan, PCR products for exon 4 and exon 8 of PGM1 were then analyzed by SSCP to detect the genotypes.@*RESULTS@#2 alleles and 3 genotypes were detected in exon 4 and 8 respectively. The discrimination power was 0.7318. PCR-SSCP analysis was suitable for determination of PGM1 genotypes from old blood and semen stains.@*CONCLUSION@#PGM1 system typed by PCR-SSCP is useful for forensic identification.
Assuntos
Humanos , Alelos , Povo Asiático/genética , China , DNA/genética , Frequência do Gene , Genótipo , Fosfoglucomutase/genética , Reação em Cadeia da Polimerase/métodos , Polimorfismo Genético , Polimorfismo Conformacional de Fita Simples , Sensibilidade e EspecificidadeRESUMO
OBJECTIVE@#The genetic polymorphism of two STR loci, D20S85 and D6S477, were studied in 280 unrelated Chinese individuals in Wuhan.@*METHODS@#The PCR amplified products were analyzed by PAGE and silver staining.@*RESULTS@#10 and 9 alleles were observed in these two STR loci, and the discriminating power (DP) were 0.9085 and 0.9127 respectively. No deviations from Hardy-Weinberg equilibrium were found. The two STR loci had been successfully applied to individual identification and paternity testing.@*CONCLUSION@#The results demonstrated that the two loci were useful for forensic identification.
Assuntos
Humanos , Alelos , Povo Asiático/genética , China , Medicina Legal , Frequência do Gene , Polimorfismo Genético , Sequências de Repetição em Tandem/genéticaRESUMO
Objective To produce the standard allelic ladder by using the cloning technique. Methods After the amplification and separation of the STR alleles, they were purified and then connected with T-vectors directly. The combinants were transfected into the component E. coli DH5? cells follwed by cloning and plasmid purification. The allelic ladder were then produced by re-amplifying the recombinant plasmid DNA. Results The allelic ladder made in this way can be produced in a lager amount and can be stored in a relatively long period. Conclusion The results demonstrated that the standard allelic ladder generated in this way is more practical in forensic scienc application. This technique in useful for preparation of domestic STR kits.
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Since the information supplied by the paternity testing of alleged parents was less than that of standard triplet parentage testing,so the paternity index (PI) calculating methods of standard triplet parentage testing was not suitable for calculating the PI value of alleged parents.In order to establish a more precise method for calculating PI value of alleged parents with STR typing results,the first thing is to summarize the standard triplet PI calculating formulas according to the Essen Mller theory.These formulas are 1/p,1/2p,1/p+q,1/2p+2q.This article reports a new PI calculating method in case of paternity testing of alleged parents.Compared with other methods,the new method for calculating Y value either considering random man and random female or considering the alleged father(mother)and random female(man).
RESUMO
Objective The purpose of this paper is to study PGM1 genotyping by PCR RFLP.Method 300 unrelated individuals of Han were genotyped using PCR RFLP. The target amplificaton products of extron 4 and 8 of PGM1 gene were digested by Bgl II and Nla III respectively.The digested DNA fragments were typed by PAGE.Result This PGM1 RFLP system can discriminate 9 genotypes with Dp of 0 7450 in Han population.Compared with conventional PAGIEF, 1+2- and 1-2+ cant be differentiated and the rare genotypes also cant be detected by this method.The advantage of this method was PGM1 genotyping successfully in bloodstains stored for 25 years and with 0 1ng genomic DNA.PGM1 RFLP method is useful for forensic identification.