OBJECTIVES: To observe and analyse the Amelogenin allelic loss in parent-child identification cases, and to explore the type and mechanism of Amelogenin allelic loss as well as its influence on gender identification and solutions. METHODS: After the detection by SiFaSTR™ 23plex DNA identification system, samples had the characteristics of the peak area of Amelogenin X was the same as the one of adjacent heterozygote or lower than one half of adjacent homozygote in females while Amelogenin X loss was observed in males were selected. X chromosome STR （X-STR） typing and Amelogenin X sequencing were performed. The samples with Amelogenin Y loss in males were confirmed by the detection of Y chromosome STR typing and sex-determining region of Y （SRY）. The type and rate of Amelogenin allelic loss were confirmed and calculated, and the mechanism and influence of this variation were also analysed. RESULTS: Amelogenin X allelic loss was observed in one male sample, the mutation in primer-binding region was confirmed by sequencing. The suspected Amelogenin X allelic loss was observed in four female samples, but the mutation in primer-binding region was confirmed by sequencing in only one sample. Amelogenin Y allelic loss was observed in seven male samples, SRY positive cases was detected in five of them, and two were SRY negative. Y-STR type was detected in four cases of the five SRY positive cases, which was not detected in the two SRY negative cases. The rate of Amelogenin allelic loss was about 0.029%. CONCLUSIONS Amelogenin X allelic loss does not affect the gender identification, but Amelogenin Y allelic loss may cause wrong gender identification. Thus, Y-STR or SRY should be detected for gender confirmation. When Y-STR genotypes are not detected in a "male" whose SRY detection is also negative, then the chromosome karyotype analysis and sex differentiation related genes test should be taken to further confirm the gender.
Amelogenin/genetics* ; DNA/genetics* ; Female ; Humans ; Loss of Heterozygosity/genetics* ; Male ; Sex Determination Analysis

OBJECTIVE: To explore the change rules of peak area ratio of STR loci to Amelogenin (AMEL) locus (STR/AMEL), a sex-determining gene in DNA degradation, and to evaluate the application of STR/AMEL value in the estimation of DNA degradation degree. METHODS: DNA was extracted from iliopsoas, and the variations of STR/AMEL value (Penta E/AMEL, Penta D/AMEL, FGA/AMEL) were analyzed after the artificial degradation was made by DNase I, and the changes of these three ratios of the iliopsoas naturally degraded in an outdoor environment were also analyzed. The regression curves were analyzed using the periods of DNA degradation and outside the body as the independent variable (x) and the STR/AMEL value as the dependent variable (y) and three curve equations under two conditions were established. RESULTS: Both under the conditions of artificial and natural degradation, STR/AMEL value had a negative relationship with the degradation time. The relationship between STR/AMEL and degradation time can be well simulated by the cubic function. R2 was over 0.99 under controlled degradation condition and over 0.86 under natural degradation condition. CONCLUSION The STR/AMEL value (Penta E/AMEL, Penta D/AMEL, FGA/AMEL) is negatively related with the DNA degradation degree, which follows mathematical regression models strictly, and it might be applied to evaluate the DNA degradation degree.
Amelogenin/genetics* ; DNA Damage/genetics* ; DNA Primers ; Humans ; Microsatellite Repeats ; Regression Analysis ; Time Factors

OBJECTIVES: To establish multiplex system of 16 miniSTR loci, and explore its application value for the degraded materials in forensic medicine. METHODS: The multiplex system of 16 miniSTR loci was established using a six-dye fluorescence labeling technology and its application value in forensic medicine was assessed. RESULTS: A six-dye fluorescence labeling miniSTR amplification kit was developed, which enabled 15 autosomal STR loci, Amelogenin locus and DYS391 to be typed simultaneously. This method showed good specificity and could provide stable and accurate typing results with a sensitivity of 50 pg. This system also provided a good test result for the normal biological sample of actual cases. CONCLUSIONS The multiplex system of 16 miniSTR loci has application value for degraded and trace materials with the advantages of high sensitivity and database compatibility, which can be used for forensic casework.
Amelogenin ; DNA Fingerprinting ; DNA Primers ; Forensic Medicine/methods* ; Microsatellite Repeats/genetics* ; Polymerase Chain Reaction

Alleles ; Amelogenin/genetics* ; Chromosomes, Human, Y ; Humans ; Male ; Sex Determination Analysis

OBJECTIVES: To recognize the possibility of Y fragment deletion of Amelogenin gene intuitively and simply according to the genotyping graphs. METHODS: By calculating the ratio of total peak height of genotyping graphs, the statistics of equilibrium distribution between Amelogenin and D3S1358 loci, Amelogenin X-gene and Amelogenin Y-gene, and different alleles of D3S1358 loci from 1 968 individuals was analyzed after amplified by PowerPlex^® 21 detection kit. RESULTS: Sum of peak height of Amelogenin X allele was not less than 60% that of D3S1358 loci alleles in 90.8% female samples, and sum of peak height of Amelogenin X allele was not higher than 70% that of D3S1358 loci alleles in 94.9% male samples. CONCLUSIONS The result of genotyping after amplified by PowerPlex^® 21 detection kit shows that the possibility of Y fragment deletion should be considered when only Amelogenin X-gene of Amelogenin is detected and the peak height of Amelogenin X-gene is not higher than 70% of the total peak height of D3S1358 loci.
Alleles ; Amelogenin/genetics* ; Asian People/genetics* ; DNA Fingerprinting/methods* ; Female ; Genotype ; Humans ; Male ; Mutation ; Polymerase Chain Reaction/methods* ; Population Groups

OBJECTIVETo investigate the types and frequencies of variants in Amelogenin gene in Chinese population and to explore the mutations' influence to the sex test.

METHODSThe Amelogenin gene of 8850 unrelated Chinese individuals was typed with PowerPlex 16 system. The samples with abnormal typing results were calculated directly, validated with different primer sets, Y-STR typing and sequencing.

RESULTSTwo samples with X chromosomal Amelogenin (AMELX) allelic dropout and 2 samples with Y chromosomal Amelogenin (AMELY) allelic dropout were observed in male individuals, the total rate of mutation was 0.045% and the rate in the male was 0.085%. Two types of point mutation which may result in null allele were observed in the primer binding region of the plostq AMELX alleles, and the mutation rate in the male was 0.042%. The mutation rate of AMELY allele was also 0.042%. One sample failed to amplify 10 Y-STR loci out of 12 loci, which could be speculated that large interstitial deletion of the Y chromosome encompassing the AMELY and other Y-STR loci occurred.

CONCLUSIONAMELX or AMELY allelic dropout may occur due to the mutation of Amelogenin gene, which may interfere with the sex test and induce wrong gender identification.

Alleles ; Amelogenin ; genetics ; Asian Continental Ancestry Group ; genetics ; DNA ; analysis ; Gene Frequency ; Humans ; Mutation ; Polymerase Chain Reaction ; methods ; Population Groups ; genetics

There are two kinds of amelogenin gene mutation, including mutation in primer-binding region of amelogenin gene and micro deletion of Y chromosome encompassing amelogenin gene, and the latter is more common. The mechanisms of mutation in primer-binding region of amelogenin gene is nucleotide point mutation and the mechanism of micro deletion of Y chromosome encompassing amelogenin gene maybe non-allelic homologous recombination or non-homologous end-joining. Among the population worldwide, there is a notably higher frequency of amelogenin gene mutations in Indian population, Sri Lanka population and Nepalese population which reside within the Indian subcontinent. Though amelogenin gene mutations have little impact on fertility and phenotype, they might cause incorrect result in gender identification. Using composite-amplification kit which including autosomal STR locus, amelogenin gene locus and multiple Y-STR locus, could avoid wrong gender identification caused by amelogenin gene mutation.
Alleles ; Amelogenin/genetics* ; Asian People/genetics* ; Chromosome Aberrations ; Chromosomes, Human, Y/genetics* ; Humans ; India ; Male ; Microsatellite Repeats ; Nepal ; Polymerase Chain Reaction ; Sequence Deletion ; Sri Lanka

OBJECTIVETo extracted DNA from ancient human teeth dated 3000 years ago unearthed in Xi'an and determine the genders for the individuals.

METHODSThirty five ancient human teeth were studied. A 'Reverse-root-canal' technique and a Chelex-100 solution were used to extract the DNA. Specific primers for Amelogenin gene were designed for PCR amplification.

RESULTSGenomic DNA was successfully extracted from 30 samples, for which 8 were determined to be males and 22 were females.

CONCLUSIONThe 'Reverse-root-canal' technique may be used for extracting DNA from ancient human teeth. Genetics method can supplement physical anthropology for determination of sex for ancient samples.

Amelogenin ; genetics ; China ; DNA ; analysis ; genetics ; isolation & purification ; Female ; History, Ancient ; Humans ; Male ; Paleodontology ; Polymerase Chain Reaction ; Sex Determination Analysis ; Tooth ; chemistry

OBJECTIVEUsing nested polymerase chain reaction (PCR) to perform preimplantation gender diagnosis.

METHODSOne (or two) lymphocyte and blastomere (n=50/group) were collected and prepared under the following conditions: (1) water only (H(2)O); (2) freeze-thaw liquid nitrogen, then boiling; (3) potassium hydroxide/dithiotheriol, heated to 65 degree centigrade, followed by acid neutralization (KOH). Cells were analyzed by PCR using nested primers amplification with amelogenin gene.

RESULTSThe amplification rate and allele dropout (ADO) rate for male lymphocytes by the three methods were 83%, 94%, 95% and 24%, 12%, 4%, respectively. Using two cells per reaction did not increase the amplification rate for the KOH method.

CONCLUSIONThe KOH method for DNA preparation is superior to the other methods evaluated. Dual blastomere biopsy and independent blastomere analysis may improve preimplantation diagnostic reliability.

Amelogenin ; Blastocyst ; cytology ; metabolism ; Blastomeres ; cytology ; metabolism ; DNA ; genetics ; Dental Enamel Proteins ; genetics ; Female ; Genotype ; Humans ; Lymphocytes ; cytology ; metabolism ; Male ; Polymerase Chain Reaction ; methods ; Sex Determination Analysis ; methods

OBJECTIVE: To establish a multiplex STR genotyping method for autosomal STR and Y-STR loci in forensic biological practice. METHODS: Widely used autosomal STR loci and Y-STR loci were selected. A set of PCR primers was designed, and a 5-dye fluorescent labeled STR multiplex PCR reagent kit was developed. RESULTS: A kit was developed which can simultaneously detect 15 autosomal STR loci, 10 Y-STR loci, and an Amelogenin. CONCLUSION The 15 autosomal STR plus 10 Y-STR kit in combination with capillary electrophoresis method was used to STR genotyping with accurate and reliable results. The new one-step testing kit can potentially be widely used in forensic cases and DNA databank in the future.
Amelogenin ; Chromosomes, Human, Y/genetics* ; DNA Primers ; Databases, Nucleic Acid ; Forensic Genetics/methods* ; Genotype ; Genotyping Techniques/instrumentation* ; Humans ; Indicators and Reagents ; Microsatellite Repeats ; Multiplex Polymerase Chain Reaction