OBJECTIVE: To develop the single-tube one-step methylation variable position (MVP) analysis technology-single-tube post-digestion PCR-melting curve analysis (PDP-MCA). METHODS: Based on differentially methylated region (DMR) reported previously as the model, a set of primers with different melting temperatures of products in the two sides of MVP were designed. By using the FastDigest methylation-sensitive restriction enzyme (MSRE), DNA digestion, multiplex amplification, MCA detection and MCA profiles were performed in a single reaction tube. Same samples (peripheral venous blood, semen, and vaginal fluid, 5 samples each type) were tested by single-tube one step MVP and traditional MSRE-PCR MCA technology. To verify the feasibility of this method, the results were compared with that of the traditional technology. The MCA/HRM profiles of different samples were analyzed and compared. RESULTS: When the melting temperature of the fragments had a differential of 2 degrees C, the MCA melting peaks separated well, and MCA detection after multiplex amplification was successful. The single-tube PDP-MCA assay was developed, which integrated multiple reactions (digestion, amplification and detection) into one tube. By this method, the sample-specific profiles and data were analyzed in 2 h, which is similar to that of the traditional method. The rapid classifications of the samples were also realized. CONCLUSION Multiplex MVPs can be analyzed in a single closed-tube. The single-tube PDP-MCA technology is a simple, fast, and automatable method. It can be used for detection of DNA methylation variations.
DNA/isolation & purification* ; DNA Methylation/genetics* ; DNA Primers/genetics* ; Humans ; Multiplex Polymerase Chain Reaction/standards* ; Nucleic Acid Denaturation

Forensic entomotoxicology is a branch of forensic medicine, which applies entomology, toxicology and other related studies to solve the poisoning cases. It has an obvious advantage in the investigation on poisoning death. Based on the expounding definition and research of entomotoxicology, this paper reviews research progress and application value in some aspects of forensic medicine, such as the effects of drugs/toxins on the growth and development of sarcosaphagous insects and the qualitative and quantitative analysis of the drugs/toxins in the poisoned body tissue.
Animals ; Death ; Entomology/methods* ; Forensic Medicine/methods* ; Humans ; Insecta ; Postmortem Changes

Stain identification has long been a task in forensic biology. The identification of semen stain, one of the most common human stains, can provide crucial information for crime scene reconstruction and forensic investigation. Traditional detection of semen stain depends largely on the microscopic identification of spermatozoa, enzyme activity-based methods or antigen-antibody reactions. These morphological, proteinological and zymological approaches, however, are apparently inadequate in identifying tiny, admixed, degraded or contaminated samples. With the development of transcriptomics and epigenetics, many semen-specific mRNA markers, such as protamine-1 (PRM1) and -2 (PRM2), have been applied to semen and semen stain identification. Messenger RNA profiling shows great promise in identifying tissues as demonstrated by the recognition of specific markers. Further more, studies on tis-sue-specific differential DNA methylation will provide a scrumptious way of identifying difficult samples.
DNA Methylation ; Forensic Medicine ; methods ; Genetic Markers ; Humans ; Male ; RNA, Messenger ; analysis ; Semen

Humans ; Precursor Cell Lymphoblastic Leukemia-Lymphoma/diagnosis* ; Death, Sudden/etiology*

Objective To detect 715 bp sequence of 28S rRNA in sarcosaphagous flies, and to identify their common species for solving the problem of morphological identification, as well as providing technical support for postmortem interval (PMI) estimation. Methods Twenty-nine common sarcosaphagous flies were collected in Luoyang and classified by morphological characteristics. The DNA was extracted from the fly's legs by Chelex-100 method and then the fragments of 28S rRNA were amplified and sequenced. The results were compared with twenty-eight corresponding fly species of GenBank and EMBL databases. All the sequences were analyzed by MEGA7.0 software, and sequence alignment was performed by the searching in BLAST. The nucleotide composition was analysed, and the intraspecific and interspecific ge-netic distance and phylogenetic tree were established. Results Twenty-nine sarcosaphagous flies were classified into 6 species of 5 genera, 3 families by morphological characteristics. In the obtained 715 bp sequence of 28S rRNA , the comparison result of online BLAST showed that the similarity was 100％. Five species were well clustered by a phylogenetic tree. Between different groups, the interspecific and intraspecific differences ranged from 0.007 to 0.045 and 0 to 0.001, respectively. Conclusion The 28S rRNA target gene sequences shows a good identification capability, which can be a new genetic marker for the identification of sarcosaphagous flies.

Medical tangles caused by the death of women and infants in perinatal period are very normal in the forensic appraisal. The author collected and analyzed 49 cases of these tangles from many aspects, such as sex and age of the dead, hospital,information of autopsy, fault of medical action and so on,and discovered the normal causes of death, medical action's effects and the causes of tangle. It would be useful to the forensic appraisal, settlement and prevention of these medical tangles.
Adult ; Asphyxia Neonatorum/mortality* ; Autopsy ; Cause of Death ; Female ; Forensic Pathology ; Humans ; Infant, Newborn ; Male ; Malpractice/legislation & jurisprudence* ; Maternal Mortality ; Postpartum Hemorrhage/mortality* ; Pregnancy ; Respiratory Tract Diseases/mortality* ; Retrospective Studies ; Young Adult

Alu family is the primate specific short interspersed repetitive elements (SINEs). Its abundance and diversity distribution in genome, high methylation level and polymorphic for insertion make them ideally suitable as tools in forensic applications. The application of A4 lu sequence in forensic genomics, include DNA quantitation, race determination, species and gender identification, personal identification, paternity testing and whole-genome amplification. The principles and characteristics of these Alu-based techniques are also summarized. The prospect of Alu as forensic molecular marker is discussed as well.
Alu Elements/genetics* ; Base Sequence ; Chromosomes, Human/genetics* ; DNA/genetics* ; DNA Methylation ; Forensic Genetics/methods* ; Genetic Markers ; Genome, Human ; Humans ; Polymerase Chain Reaction ; Polymorphism, Genetic/genetics* ; Sensitivity and Specificity ; Sequence Analysis, DNA

This study was to investigate the antiviral effects of a hot water soluble extract S-03 isolated from Isatis indigotica root on different subtypes of influenza A and B viruses in MDCK cell cultures, using plaque reduction, immunofluorescence and hemo-agglutination inhibition (HAD) assays. Chemical analysis of the extract S-03 showed that it contained high proportion of polysaccharides. The antiviral effects in vitro showed that the S-03 had no effect on different influenza viruses if the drug was used before virus adsorption, but S-03 showed obvious activities against influenza viruses if treatment after virus adsorption or direct reaction of drug and virus before virus adsorption. Hemagglutination inhibition assay showed that S-03 inhibited HA activities of different human influenza viruses (inhibition concentration ranged from 3.12 to 25 mg/mL), avain influenza viruses (inhibition concentration ranged from 25 to 50 mg/mL). The antiviral effects of S-03 on different influenza A and B viruses in vitro might be through the inhibition of the HA to prevent infection.
Animals ; Cells, Cultured ; Dogs ; Fluorescent Antibody Technique ; Hemagglutination Inhibition Tests ; Influenza A virus ; drug effects ; Influenza B virus ; drug effects ; Isatis ; chemistry ; Plant Extracts ; pharmacology ; Plant Roots

OBJECTIVESTo confirm previous whole-genome scan results of mapping type 2 diabetes susceptibility genes in chromosome 1 in Northern Chinese Han population by conducting a new genome scan with both an enlarged number of type 2 diabetes families and a new set of microsatellite markers.

METHODSA genome scan method was applied. After multiplexed PCR, electrophoreses, genescan and genotyping analysis, size informations for all loci were obtained, and a further study was done using both parametric and non-parametric linkage analysis to calculate the P-values and Z-values of these loci.

RESULTSA total of 34 microsatellite markers distributed within 5 regions along chromosome 1 were surveyed, and 12,000 genotypes were screened. Evidence of linkage with diabetes was identified for 8 of the 34 loci (all the P-values of the 8 loci distributed in 3 regions were lower than 0.05, and the highest Z-value was 2.17). Interestingly, all the 5 markers at the P terminal 1p36.3-1p36.23 region, spanning a long range of 16.9 cM, suggested to be linked with the disease. The results of the other two regions were not consistent with the previous ones.

CONCLUSIONSThe study results have confirmed those gained in the previous genome-wide scan. The fact that all 5 loci at the P terminal region displayed linkage with diabetes suggests that more than 1 susceptibility gene may reside in this region.

Asian Continental Ancestry Group ; Chromosome Mapping ; Chromosomes, Human, Pair 1 ; Diabetes Mellitus, Type 2 ; genetics ; Ethnic Groups ; Genetic Linkage ; Genetic Predisposition to Disease ; Genetic Testing ; Genotype ; Humans ; Microsatellite Repeats ; genetics

OBJECTIVES: To analyze the relationship among electrical conductivity （EC）, total volatile basic nitrogen （TVB-N）, which is an index of decomposition rate for meat production, and postmortem interval （PMI）. To explore the feasibility of EC as an index of cadaveric skeletal muscle decomposition rate and lay the foundation for PMI estimation. METHODS: Healthy Sprague-Dawley rats were sacrificed by cervical vertebrae dislocation and kept at 28 ℃. Muscle of rear limbs was removed at different PMI, homogenized in deionized water and then skeletal extraction liquid of mass concentration 0.1 g/mL was prepared. EC and TVB-N of extraction liquid were separately determined. The correlation between EC (x₁) and TVB-N (x₂) was analyzed, and their regression function was established. The relationship between PMI (y) and these two parameters were studied, and their regression functions were separately established. RESULTS: The change trends of EC and TVB-N of skeletal extraction liquid at different PMI were almost the same, and there was a linear positive correlation between them. The regression equation was x₂=0.14x₁-164.91(R²=0.982). EC and TVB-N of skeletal muscle changed significantly with PMI, and the regression functions were y=19.38x₁³-370.68x₁²+2 526.03 x₁-717.06(R²=0.994), and y=2.56x₂³-48.39x₂²+330.60x₂-255.04(R²=0.997), respectively. CONCLUSIONS EC and TVB-N of rat postmortem skeletal muscle show similar change trends, which can be used as an index for decomposition rate of cadaveric skeletal muscle and provide a method for further study of late PMI estimation.
Animals ; Autopsy ; Electric Conductivity ; Forensic Pathology ; Muscle, Skeletal/pathology* ; Nitrogen ; Postmortem Changes ; Rats ; Rats, Sprague-Dawley ; Time Factors