Sex determination is the most important step in personal identification in forensic investigations. DNA-based sex determination analysis is comparatively more reliable than the other conventional methods of sex determination analysis. Advanced technology like real-time polymerase chain reaction (PCR) offers accurate and reproducible results and is at the level of legal acceptance. But still there are situations like chimerism where an individual possess both male and female specific factors together in their body. Sex determination analysis in such cases can give erroneous results. This paper discusses the phenomenon of chimerism and its impact on sex determination analysis in forensic investigations.
Logistic regression was applied to develop a morphometric sexing method of two closely related stork species that were previously sexed through amplification of the CHD gene. Tarsus length (TL) and bill length (BL) measurements were recorded from captive populations of adult Milky Stork (Mycteria cinerea) (n = 60) and Painted Stork (Mycteria leucocephala) (n = 58) at Zoo Negara Malaysia. Despite having monomorphic plumages, both stork species exhibited normal sexual size dimorphism in which males were significantly larger than females in the tested variables. Based on logistic regression analysis, BL correctly classified the sex of sampled individuals from Painted and Milky stork with an overall predicted accuracy of 94.8 and 90.0%, respectively. However, TL measurements generated a lower predicted accuracy level of 86.2% and a same accuracy level of 90% on the sex classification of individuals from Painted and Milky stork, respectively. By comparing the measurements of both species, only the average BL measurements of the Milky storks were significantly lower than that of Painted storks (t-test, P80.001). The logistic regression equation in this study may serve as a simple and more practical option for sexing Milky and Painted storks for their breeding and conservation programmes.
Matched MeSH terms: Sex Determination Analysis/methods*; Sex Determination Analysis/veterinary*
Individual identification is an important and challenging task in forensic investigation. Lip print on drinking glass or cigarette butt found at crime scenes may link to a suspect. The aim of this study was to determine the differences in lip print between sexes or races, differences in lip measurement between sexes or races and determine a way to estimate sex and race by using lip print or lip measurements for main races in Malaysia. A total of 134 subjects (67 males and 67 females) of Malay, Chinese and India were recruited from Universiti Kebangsaan Malaysia Campus Kuala Lumpur (UKMKKL), Malaysia. Lip prints were taken by using a lipstick and a transparent cellophane tape. Lip measurements were taken by using electronic digital callipers. Lip prints were classified according to Tsuchihashi classification. Statistical analysis indicated that there was a significant difference in lip print between sexes (p < 0.001) but not in races (p > 0.05). Width of oral opening and the height of lower lip both indicated significant differences between sexes (p < 0.001) while the height of upper lip and lower lip each indicated significant differences between races (p < 0.05). However, there was no significant interaction between sexes and races for all lip measurements. Formulae for sex and race determination were calculated with Classification Tree when there was significant difference between every comparison. Tables of accuracy percentage and performance evaluation for method in categorizing sex or race by using lip print or lip measurement were made. For validation of method in sex determination based on the formulae formed, accuracy in females is 90% and 65% in males. Therefore, overall percentage of accuracy in sex determination was 77.5%. This study can provide a preliminary idea about the use of lip prints in sex or race determination among Malaysian population.
In this study, we first identified male-specific SNP markers using restriction site-associated DNA sequencing, and further developed a PCR-based sex identification technique for Charybdis feriatus. A total of 296.96 million clean reads were obtained, with 114.95 and 182.01 million from females and males. After assembly and alignment, 10 SNP markers were identified being heterozygous in males but homozygous in females. Five markers were further confirmed to be male-specific in a large number of individuals. Moreover, two male-specific sense primers and a common antisense primer were designed, using which, a PCR-based genetic sex identification method was successfully developed and used to identify the sex of 103 individuals, with a result of 49 females and 54 males. The presence of male-specific SNP markers suggests an XX/XY sex determination system for C. feriatus. These findings should be helpful for better understanding sex determination mechanism, and drafting artificial breeding program in crustaceans.
Matched MeSH terms: Sex Determination Analysis/methods*
Chelonian exhibit temperature dependent sex determination, and ex situ incubation of eggs in conservation hatcheries may render a gender bias. The gender of juvenile Painted terrapins (Batagur borneoensis) produced at a conservation hatchery in Malaysia was determined by endoscopy of the gonads. Circulating reproductive hormones (testosterone, progesterone and estradiol) were profiled for 31 juveniles and nine captive-reared non-breeding adult terrapins. Endoscopy revealed a gender bias of 96.8% (30/31) females. Testosterone levels in the juvenile females (2.49 ± 1.29) were significantly lower than that of the adult females (12.20 ± 4.29), and lower than values in the juvenile male (9.36) and adult males (27.60, 35.62). The progesterone levels in the juvenile females (107.12 ± 68.68) were significantly higher than that of the adult females (51.13 ± 24.67), but lower than values in the juvenile male (33.27) and adult males (3.43, 8.51). Estrogen levels were significantly lower in the juvenile females (1.57 ± 1.35) compared to the adult females (77.46 ± 53.45). Negative correlations were observed between levels of progesterone and testosterone, and progesterone and estrogen. A positive correlation was noted between estrogen and testosterone. The present study constitutes the first attempt to determine the gender and reproductive hormone profiles of juvenile Painted terrapins produced by ex situ incubation, and captive non-breeding adults. Endoscopy of the gonads is a useful techniques for gender determination among juvenile turtles, while the use of testosterone as a gender biomarker warrants further investigation.
Matched MeSH terms: Sex Determination Analysis/veterinary*
A new proteomics technology has been implemented to study the protein repertoires of developing oocytes of giant grouper (Epinephelus lanceolatus). Knowledge of the chemical composition and physiochemical properties of vitellogenin (Vtg) is necessary to interpret the functional and biological properties attributed during ovulation. Vtg, as a biomarker indicator in sex determination, has been analyzed to determine the sex and maturational status of fish in the absence of the gonad tissue. A male giant grouper was induced by 2 mg/kg of 17ß-estradiol (E2), and blood was sampled at days 0, 1, 3, 5, and 10. SDS-PAGE 1D electrophoresis was used to analyze Vtg protein, and Vtg identification was done with 4800 Plus MALDI TOF/TOF™ mass spectrophotometer (Applied Biosystems/MDS SCIEX, USA). Meanwhile, MS/MS de novo sequencing identified the proteins by matching sequences of tryptic peptides to the known sequences of other species. Vtg was confirmed by MASCOT at 95% significant level, and molecular mass was 187 kDa. Protein resolved on SDS-PAGE as a double band of approximately the same mass as determined with MALDI-TOF. The N-terminal sequences and identification of Vtg were also determined. The potential of using MS methods to understand the structure and function of Vtg is discussed.
Matched MeSH terms: Sex Determination Analysis/methods; Sex Determination Analysis/veterinary*
Sex determination of livestock is performed to achieve the objectives of livestock breeding programmes. Techniques for sex determination have evolved from karyotyping to detecting Y-specific antigens and recently to the polymerase chain reaction (PCR), which appears to be the most sensitive, accurate, rapid and reliable method to date. In this study, a PCR-based sex determination method for potential application in goat breeding programmes was developed. Primers were designed to amplify a portion of the X amelogenin gene (Aml-X) on the X chromosome to give a 300 bp product and Sry gene on the Y chromosome to give a 116 bp product. PCR optimization was performed using DNA template extracted from a whole blood sample of Jermasia goats (German Fawn x Katjang) of both sexes. It was possible to identify the sex chromosomes by amplifying both male- and female-specific genes simultaneously in a duplex reaction with males yielding two bands and females yielding one band. The Aml-X primer set, which served as an internal control primer, did not interfere with amplification of the Y-specific sequence even when a low amount of DNA (1 ng) was used. The duplex reaction subjected to a blind test showed 100% (14/14) concordance, proving its accuracy and reliability. The primer sets used were found to be highly specific and were suitable for gender selection of goats.
Matched MeSH terms: Sex Determination Analysis/methods*
Amelogenin paralogs on Chromosome X (AMELX) and Y (AMELY) are commonly used sexing markers. Interstitial deletion of Yp involving the AMELY locus has previously been reported. The combined frequency of the AMELY null allele in Singapore and Malaysia populations is 2.7%, 0.6% in Indian and Malay ethnic groups respectively. It is absent among 541 Chinese screened. The null allele in this study belongs to 3 Y haplogroups; J2e1 (85.7%), F* (9.5%) and D* (4.8%). Low and high-resolution STS mapping, followed by sequence analysis of breakpoint junction confirmed a large deletion of 3 to 3.7-Mb located at the Yp11.2 region. Both breakpoints were located in TSPY repeat arrays, suggesting a non-allelic homologous recombination (NAHR) mechanism of deletion. All regional null samples shared identical breakpoint sequences according to their haplogroup affiliation, providing molecular evidence of a common ancestry origin for each haplogroup, and at least 3 independent deletion events recurred in history. The estimated ages based on Y-SNP and STR analysis were approximately 13.5 +/- 3.1 kyears and approximately 0.9 +/- 0.9 kyears for the J2e1 and F* mutations, respectively. A novel polymorphism G > A at Y-GATA-H4 locus in complete linkage disequilibrium with J2e1 null mutations is a more recent event. This work re-emphasizes the need to include other sexing markers for gender determination in certain regional populations. The frequency difference among global populations suggests it constitutes another structural variation locus of human chromosome Y. The breakpoint sequences provide further information to a better understanding of the NAHR mechanism and DNA rearrangements due to higher order genomic architecture.
Matched MeSH terms: Sex Determination Analysis/methods
Multi-wall carbon nanotubes (MWCNTs) were modified to design a new DNA biosensor. Functionalized MWCNTs were equipped with gold nanoparticles (GNPs) (~15nm) (GNP-MWCNTCOOH) to construct DNA biosensors based on carbon-paste screen-printed (SPE) electrodes. GNP attachment onto functionalized MWCNTs was carried out by microwave irradiation and was confirmed by spectroscopic studies and surface analysis. DNA biosensors based on differential pulse voltammetry (DPV) were constructed by immobilizing thiolated single-stranded DNA probes onto GNP-MWCNTCOOH. Ruthenium (III) chloride hexaammoniate [Ru(NH3)6,2Cl(-)] (RuHex) was used as hybridization redox indicator. RuHex and MWCNT interaction was low in compared to other organic redox hybridization indicators. The linear response range for DNA determination was 1×10(-21) to 1×10(-9)M with a lower detection limit of 1.55×10(-21)M. Thus, the attachment of GNPs onto functionalized MWCNTs yielded sensitive DNA biosensor with low detection limit and stability more than 30days. Constructed electrode was used to determine gender of arowana fish.
Matched MeSH terms: Sex Determination Analysis/methods*