Internal validation of the GA118-24B Genetic Analyzer, a stable capillary electrophoresis system for forensic DNA identification.

The GA118-24B Genetic Analyzer (hereafter, "GA118-24B") is an independently developed capillary electrophoresis instrument. In the present research, we designed a series of validation experiments to test its performance at detecting DNA fragments compared to the Applied Biosystems 3500 Genetic Analyzer (hereafter, "3500"). Three commercially available autosomal short tandem repeat multiplex kits were used in this validation. The results showed that GA118-24B had acceptable spectral calibration for three kits. The results of accuracy and concordance studies were also satisfactory. GA118-24B showed excellent precision, with a standard deviation of less than 0.1 bp. Sensitivity and mixture studies indicated that GA118-24B could detect low-template DNA and complex mixtures as well as the results generated by 3500 in parallel experiments. Based on the experimental results, we set specific analytical and stochastic thresholds. Besides, GA118-24B showed superiority than 3500 within certain size ranges in the resolution study. Instead of conventional commercial multiplex kits, GA118-24B performed stably on a self-developed eight-dye multiplex system, which were not performed on 3500 Genetic Analyzer. We compared our validation results with those of previous research and found our results to be convincing. Overall, we conclude that GA118-24B is a stable and reliable genetic analyzer for forensic DNA identification.

Validation of the PowerPlex®35GY System: a novel eight-dye STR multiplex kit on the Spectrum Compact CE System.

The PowerPlex® 35GY System (Promega, USA) is an advanced eight-dye multiplex STR kit, incorporating twenty-three autosomal STR loci, eleven Y chromosome STR loci, one sex determining marker Amelogenin, and two quality indicators. This multiplex system includes 20 CODIS loci and up to 15 mini-STR loci with sizing values less than 250 bases. In this study, validation for PowerPlex® 35GY System was conducted following the guidelines of SWGDAM, encompassing sensitivity, precision, accuracy, concordance, species specificity, stutter, mixture, stability, and degraded DNA. The results from experiments demonstrated that the PowerPlex® 35GY System could effectively amplify DNA samples, with complete allele detection achieved at 125 pg. Moreover, over 90% of alleles from minor contributors were detected at a mixed ratio of 1:4. Additionally, the system was found to yield full profiles even in the presence of hematin, humic acid, and indigo. The PowerPlex® 35GY System demonstrated superior performance in the sensitivity and degraded DNA studies compared to a six-dye STR kit. Hence, it is evident that the PowerPlex® 35GY System is well-suited for forensic practice, whether in casework or for database samples. These findings provide strong support for the efficacy and reliability of the PowerPlex® 35GY System in forensic applications.

Boosting Smoking Cessation Intervention Utilization in Chinese Healthcare Providers: A Randomized Controlled Trial of the 'WeChat WeQuit' Medical Education Program.

INTRODUCTION: In China, standard smoking cessation practices are rarely used by healthcare service providers (HSPs). WeChat, a popular social media app, has been widely used in China. METHODS: In this single-blind, randomized trial, undertaken in China with 8-week interventions and follow-up to 34 weeks, 1887 HSPs were randomly selected to the intervention (n=942) or control group (n=945) from Oct 2020 to Oct 2021. The intervention group received regular smoking cessation training program messages from the professional team for 8 weeks and followed for 34 weeks. The control group received thanks messages for 8 weeks, and follow-up to 34 weeks. Both groups received a hard copy of the manual after randomization. The primary outcome measure was the utilization rate of behavioral and pharmacotherapy interventions for smoking patients from 9 to 34 weeks. This trial is registered at ClinicalTrials.gov (number NCT03556774). RESULTS: HSPs in the intervention group demonstrated a better overall utilization rate of smoking cessation at 20-week follow-up compared to the control group (35.54% vs 31.41%, p=0.036). Additionally, both groups showed a significant increase in the adoption of various components of the 5A's model - including "Assess", "Assist: set a quit date", "Assist: recommend cessation program", "Assist: provide information", "Assist: recommend medication", and "Arrange" - at the 9-week follow-up relative to baseline. Notably, at the 20-week follow-up, the intervention group reported significantly enhanced utilization rates for all these components, except "Assist: set a quit date". CONCLUSIONS: The "Wechat Wequit" training program effectively enhanced smoking cessation intervention adoption among Chinese HSPs. IMPLICATION: 'WeChat WeQuit' training program was effective in increasing the provision of effective tobacco cessation interventions by Chinese-speaking HSPs to patients with cigarette smoking, which could provide valuable insights into bridging the gap between need and services for smoking cessation in China.

A preliminary exploration for co-detecting RNA virus and STR type on capillary electrophoresis in forensic practice.

RNA virus infection such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection shows severe respiratory symptoms on human and could be an obvious individual characteristic for investigations in forensic science. As for biological samples suspected to contain RNA virus in forensic casework, it requires respective detection of viral RNA and human DNA: reverse transcriptase polymerase chain reaction and DNA type (short tandem repeat [STR] analysis). Capillary electrophoresis (CE) has been shown to be a versatile technique and used for a variety of applications, so we preliminarily explored the co-detection of RNA virus and STR type on CE by developing a system of co-detecting SARS-CoV-2 and STR type under ensuring both the efficiency of forensic DNA analysis and safety of the laboratory. This study investigated the development and validation of the system, including N and ORF1ab primer designs, polymerase chain reaction amplification, allelic ladder, CE detection, thermal cycling parameters, concordance, sensitivity, species specificity, precision, and contrived and real SARS-CoV-2 sample studies. Final results showed the system could simultaneously detect SARS-CoV-2 and STR type, further indicating that CE has possibilities in the multi-detection of RNA viruses/STR type to help to prompt individual characteristics (viral infection) and narrow the scope of investigation in forensic science.

Preliminary study on genetic factors related to Demirjian's tooth age estimation method based on genome-wide association analysis.

The age determination of individuals, especially minors, is critical in forensic research. In forensic practice, dental age estimation is one of the most commonly used methods for determining age as teeth are easy to preserve and relatively resistant to environmental factors. Tooth development is affected and regulated by genetic factors; however, these are not incorporated into current commonly used tooth age inference methods, leading to unreliable results. Here, we established a Demirjian and a Cameriere tooth age estimation-based methods suitable for use in children in southern China. By using the difference between the inferred age and the actual age (MD) as the phenotype, we identified 65 and 49 SNPs related to tooth age estimation from 743,722 loci among 171 children in southern China through a genome-wide association analysis (p<0.0001). We also conducted a genome-wide association study on dental development stage (DD) using the Demirjian tooth age estimation method and screened two sets of SNP sites (52 and 26) based on whether age difference was considered. The gene function enrichment analysis of these SNPs found that they were related to bone development and mineralization. Although SNP sites screened based on MD seem to improve the accuracy of tooth age estimation, there is little correlation between these SNPs and an individual's Demirjian morphological stage. In conclusion, we found that individual genotypes can affect tooth age estimation, and based on different phenotypic analysis models, we have identified some novel SNP sites related to tooth age inference and Demirjian's tooth development stage. These studies provide a reference for subsequent phenotypic selection based on tooth age inference analysis, and the results could possibly be used in the future to make forensic age estimation more accurate.

Application of Microhaplotypes in Sibling Kinship Testing.

OBJECTIVES: To investigate the efficacy of different numbers of microhaplotype (MH) loci and the introduction of different reference samples on the identification of full sibling, half sibling and differentiation between full sibling and half sibling kinships, and to explore the effect of changing mutation rate on sibling testing. METHODS: First, a family map involving three generations was established, and four full sibling identification models, five half sibling identification models and five models distinguishing full and half siblings were constructed for different reference samples introduced. Based on the results of the previous study, two sets of nonbinary SNP-MH containing 34 and 54 loci were selected. Based on the above MH loci, 100 000 pairs of full sibling vs. unrelated individuals, 100 000 pairs of half sibling vs. unrelated individuals and 100 000 pairs of full sibling vs. half sibling were simulated based on the corresponding sibling kinship testing models, and the efficacy of each sibling kinship testing model was analyzed by the likelihood ratio algorithm under different thresholds. The mutant rate of 54 MH loci was changed to analyze the effect of mutation rate on sibling identification. RESULTS: In the same relationship testing model, the systematic efficacy of sibling testing was positively correlated with the number of MH loci detected. With the same number of MH loci, the efficacy of full sibling testing was better than that of uncle or grandfather when the reference sample introduced was a full sibling of A, but there was no significant difference in the identification efficacy of the four reference samples introduced for full sibling and half sibling differentiation testing. In addition, the mutation rate had a slight effect on the efficacy of sibling kinship testing. CONCLUSIONS: Increasing the number of MH loci and introducing reference samples of known relatives can increase the efficacy of full sibling testing, half sibling testing, and differentiation between full and half sibling kinships. The level of mutation rate in sibling testing by likelihood ratio method has a slight but insignificant effect on the efficacy.

A novel set of short microhaplotypes based on non-binary SNPs for forensic challenging samples.

Short tandem repeats (STRs) are the most widely used genetic markers in forensic application, but they are not ideal genetic markers for the analysis of forensic challenging samples such as highly degraded or unbalanced mixed samples because of their relatively large amplicons and stutter peaks. In this study, we developed a set of short microhaplotypes based on non-binary SNPs with molecular extent sizes no longer than 60 bases and genotyped 100 unrelated individuals from northern Han groups. Our results showed this panel has similar discrimination power to STR kits, as the combined random match probability (CMP) reached 1.396 × 10-22 and mean effective number of alleles (Ae) was 3.59. The cumulative probability of exclusion for duos (CPE-duos) was 0.999919 and the cumulative probability of exclusion for trios (CPE-trios) was 0.9999999987, suggesting this panel could be applied for forensic personal identification and parentage testing independently. Population differentiation in 26 populations from the 1000 Genomes Project indicated this panel could distinguish populations from Africa, East Asia, South Asia, America, and Europe. These microhaplotypes based on non-binary SNPs have short amplicons, good discrimination power, no stutter artifacts, and have great potential in detection of highly degraded and unbalanced mixtures for personal identification, paternity testing, and ancestry inference.

An 18 Multi-InDels panel for analysis of highly degraded forensic biological samples.

DNA genotyping from trace and highly degraded biological samples is one of the most significant challenges of forensic DNA identification. There is a lack of simple and effective methods for genotyping highly degraded samples. In this study, a multiple loci insertion/deletion polymorphisms (Multi-InDels) panel was designed for detecting 18 autosomal Multi-InDels through capillary electrophoresis (CE) with amplicon sizes no longer than 125 bp. Studies of sensitivity, degradation, and species specificity were performed and a population study was carried out using 192 samples from Han populations in Hunan province in the south of China. The combined random match probability (CMP) of these 18 Multi-InDels was 3.23 × 10-12 and the cumulative probability of exclusion (CPE) was 0.9989, suggesting this panel could be used independently for human identification and could provide efficient supporting information for parentage testing. Complete profiles were obtained from as low as 62.5 pg of total input DNA after increasing the number of PCR cycles. Moreover, all alleles were detected from artificially highly degraded DNA after 80 min of boiling water bath treatment. This 18 Multi-InDels panel is simple, fast, and effective for the forensic analysis of highly degraded DNA.

Developmental validation of the STRscan-17LC kit: a 6 Dye STR kit enhanced stability and ability to detect degraded samples.

Genotyping of short tandem repeat (STR) markers is the basic method of forensic science. Enhanced technologies are needed to meet the requirements of databasing and casework samples. The STRscan-17LC kit is a 6 Dye STR kit which amplifies 16 STR loci: D3S1358, TPOX, D16S539, vWA, D2S1338, CSF1PO, D19S433, D7S820, FGA, D8S1179, D5S818, D13S317, D18S51, TH01, D12S391, and D21S11 and the sex-determinant locus amelogenin. This kit is designed for better tolerance to PCR inhibitors and analysis of mildly degraded samples with all fragments smaller than 330 bases. In this study, the STRscan-17LC kit is validated according to the SWGDAM (Scientific Working Group on DNA Analysis Methods) guidelines, including PCR-based studies, sensitivity, precision and accuracy, inhibitors, species specificity, DNA mixture studies, population, and concordance studies. The validation results suggest that the STRscan-17LC kit is a useful tool for forensic application.

Development and validation of novel 8-dye short tandem repeat multiplex system for forensic applications.

DNA profiling of short tandem repeats (STRs) is the primary method for genotyping forensic samples. However, degraded DNA and trace samples are still major problems for commercial 5- or 6-dye STR kits. In order to improve the performance of this method, we developed a novel 8-dye STR multiplex system containing 18 autosomal loci (D3S1358, D1S1656, TPOX, D16S539, vWA, D6S1043, D2S1338, CSF1PO, D19S433, D7S820, FGA, D8S1179, D5S818, D13S317, TH01, D21S11, D12S391, and PentaD) and the sex-determining locus Amelogenin, with all fragments smaller than 330 bases. Validation was carried out as recommended by the Scientific Working Group on DNA Analysis Methods. The results showed that complete profiles were obtainable when the input DNA was as low as 0.0625 ng. Full profiles were obtained even in the presence of inhibitors such as humic acid (< 300 ng/µl), hematin (< 100 µM), and indigo (0.01%). The 8-dye STR multiplex system also showed good performance in the detection degraded DNA samples. These results indicate that the 8-dye STR multiplex system is suitable for human DNA genotyping, including for difficult forensic materials.

Genetic characterization of 19 X-STRs in Sierra Leone population from Freetown.

A total of 550 individuals (265 males and 285 females) from Sierra Leone, a west-African coastal country, were genotyped using the Microreader™ 19X ID System kit. No significant deviations from the Hardy-Weinberg equilibrium were observed. A total of 250 alleles were identified with corresponding allele frequencies spanning from 0.0012 to 0.6762. PIC of the loci ranged from 0.4615 to 0.9481. The CPE, CPDF, and CPDM were 0.9999997856, 0.999999999999999999995774, and 0.999999999998997, respectively. The highly combined MECKruger, MECKishida, MECDesmarais, and MECDesmarais Duo were achieved as 0.99999992508, 0.999999999990802, 0.999999999990836, and 0.99999998412, respectively. Genetic comparisons revealed that genetic homogeneity existed in similar ethno origin or geographic origin populations. This is a pioneering genetic investigation using the Microreader™ 19X ID System kit in the population of Sierra Leone.

Current status and potential role of circular RNAs in neurological disorders.

Given the importance of non-coding RNAs in modulating normal brain functions and their implications in the treatment of neurological disorders, non-coding RNA-based diagnostic and therapeutic strategies have shown great clinical potential. Circular RNAs (circRNAs) have emerged as potentially important players in this field. Recent studies have indicated that circRNAs might play vital roles in Alzheimer's disease, Parkinson's disease, ischemic brain injury, and neurotoxicity. However, the mechanisms of action of circRNAs have not been fully characterized. We aimed to review recent advances in circRNA research in the brain to provide new insights on the roles of circRNAs in neurological disorders.

Intrapuparial stage aging and PMI estimation based on the developmental transcriptomes of forensically important Aldrichina grahami (Diptera: Calliphoridae) gene expression.

Background: The expression profiles of differentially expressed genes (DEGs) during pupal development have been demonstrated to be vital in age estimation of forensic entomological study. Here, using forensically important Aldrichina grahami (Diptera: Calliphoridae), we aimed to explore the potential of intrapuparial stage aging and postmortem interval (PMI) estimation based on characterization of successive developmental transcriptomes and gene expression patterns. Methods: We collected A. grahami pupae at 11 successive intrapuparial stages at 20 °C and used the RNA-seq technique to build the transcriptome profiles of their intrapuparial stages. The DEGs were identified during the different intrapuparial stages using comparative transcriptome analysis. The selected marker DEGs were classified and clustered for intrapuparial stage aging and PMI estimation and then further verified for transcriptome data validation. Ultimately, we categorized the overall gene expression levels as the dependent variable and the age of intrapuparial A. grahami as the independent variable to conduct nonlinear regression analysis. Results: We redefined the intrapuparial stages of A. grahami into five key successive substages (I, II, III, IV, and V), based on the overall gene expression patterns during pupal development. We screened 99 specific time-dependent expressed genes (stage-specific DEGs) to determine the different intrapuparial stages based on comparison of the gene expression levels during the 11 developmental intrapuparial stages of A. grahami. We observed that 55 DEGs showed persistent upregulation during the development of intrapuparial A. grahami. We then selected four DEGs (act79b, act88f, up and ninac) which presented consistent upregulation using RT-qPCR (quantitative real-time PCR) analysis, along with consideration of the maximum fold changes during the pupal development. We conducted nonlinear regression analysis to simulate the calculations of the relationships between the expression levels of the four selected DEGs and the developmental time of intrapuparial A. grahami and constructed fitting curves. The curves demonstrated that act79b and ninac showed continuous relatively increasing levels. Conclusions: This study redefined the intrapuparial stages of A. grahami based on expression profiles of developmental transcriptomes for the first time. The stage-specific DEGs and those with consistent tendencies of expression were found to have potential in age estimation of intrapuparial A. grahami and could be supplementary to a more accurate prediction of PMI.

A preliminary study on identification of the blood donor in a body fluid mixture using a novel compound genetic marker blood-specific methylation-microhaplotype.

Blood-containing mixtures are frequently encountered at crime scenes involving violence and murder. However, the presence of blood, and the association of blood with a specific donor within these mixtures present significant challenges in forensic analysis. In light of these challenges, this study sought to address these issues by leveraging blood-specific methylation sites and closely linked microhaplotype sites, proposing a novel composite genetic marker known as "blood-specific methylation-microhaplotype". This marker was designed to the detection of blood and the determination of blood donor within blood-containing mixtures. According to the selection criteria mentioned in the Materials and Methods section, we selected 10 blood-specific methylation-microhaplotype loci for inclusion in this study. Among these loci, eight exhibited blood-specific hypomethylation, while the remaining two displayed blood-specific hypermethylation. Based on data obtained from 124 individual samples in our study, the combined discrimination power (CPD) of these 10 successfully sequenced loci was 0.999999298. The sample allele methylation rate (Ram) was obtained from massive parallel sequencing (MPS), which was defined as the proportion of methylated reads to the total clustered reads that were genotyped to a specific allele. To develop an allele type classification model capable of identifying the presence of blood and the blood donor, we used the Random Forest algorithm. This model was trained and evaluated using the Ram distribution of individual samples and the Ram distribution of simulated shared alleles. Subsequently, we applied the developed allele type classification model to predict alleles within actual mixtures, trying to exclude non-blood-specific alleles, ultimately allowing us to identify the presence of blood and the blood donor in the blood-containing mixtures. Our findings demonstrate that these blood-specific methylation-microhaplotype loci have the capability to not only detect the presence of blood but also accurately associate blood with the true donor in blood-containing mixtures with the mixing ratios of 1:29, 1:19, 1:9, 1:4, 1:2, 2:1, 7:1, 8:1, 31:1 and 36:1 (blood:non-blood) by DNA mixture interpretation methods. In addition, the presence of blood and the true blood donor could be identified in a mixture containing four body fluids (blood:vaginal fluid:semen:saliva = 1:1:1:1). It is important to note that while these loci exhibit great potential, the impact of allele dropouts and alleles misidentification must be considered when interpreting the results. This is a preliminary study utilising blood-specific methylation-microhaplotype as a complementary tool to other well-established genetic markers (STR, SNP, microhaplotype, etc.) for the analysis in blood-containing mixtures.

Utilization of intraoperative indocyanine green fluorescence imaging to identify vascular anatomy in severe pleural adhesions in uniportal video-assisted thoracoscopic surgery: a case report.

Background: Extensive and dense pleural adhesion is a serious challenge in video-assisted thoracoscopic surgery (VATS), in which identification of vessels and their anatomical spaces is difficult. Once critical vessel is damaged while dissecting adhesion in VATS, leading to fatal hemorrhage, the surgeon will have to switch to thoracotomy. This is the first report of a case in which intraoperative indocyanine green (ICG) fluorescence imaging was used to identify critical vessels in severe pleural adhesions in uniportal VATS. Case Description: The patient (67-year-old male) with an 8-year history of tuberculosis and severe mixed ventilation dysfunction underwent a standardized wedge resection due to chest computed tomography (CT) scan that revealed a 2.6-cm nodule in the right upper lung. Intraoperatively, the superior vena cava and azygos vein were successfully identified and safely dissected using ICG fluorescence imaging in the presence of extensive and dense pleural adhesions. The chest drainage tube was removed on postoperative day (POD) 3, and patient was released from hospital on POD 5. The patient recovered well and no complication was observed in the follow-up. Conclusions: The ICG fluorescence imaging is used to illustrate the vessels and help to dissect them safely, which is a feasible, visualizable, and user-friendly method in severe pleural adhesions in uniportal VATS.

Research on the Potential Biomarkers of Mild Traumatic Brain Injury: a Systematic Review and Bibliometric Analysis.

Reliable diagnostic methods for mild traumatic brain injury (mTBI) are lacking, and many researchers continue to search for objective biomarkers that can both define and detect mTBI. Although much research has been conducted in this field, there have not been many bibliometric studies. In this study, we aim to analyze the development over the last two decades in scientific output relating to the diagnosis of mTBI. To do this, we extracted documents from Web of Science, PubMed, and Embase and performed descriptive analysis (number of publications, primary journals, authors, and countries/regions), trend topics analysis, and citation analysis for papers across the globe, with a particular focus on molecular markers. One thousand twenty-three publications spanning 390 journals were identified on Web of Science, PubMed, and Embase for the period from 2000 to 2022. The number of publications increased every year (from 2 in 2000 to 137 in 2022). Of all the publications we analyzed, 58.7% had authors from the USA. Our analysis shows that molecular markers are the most studied markers in the field of mTBI diagnostics, accounting for 28.4% of all publications, and that the number of studies focused on this specific aspect has increased sharply in the past 5 years, indicating that molecular markers may become a research trend in the future.

Combining the third molar mineralization to further improve the accuracy of the Kvaal's method in dental age estimation of subadults in northern China.

The morphological changes based on deposition of secondary dentin and mineralization of the third molar have been proven to be related to chronological age. However, Kvaal's method on the theory of deposition of secondary dentin was controversial with respect to dental age estimation in the recent research. The aim of this study was to combine the parameters of Kvaal's method with relatively high correlation coefficients and mineralization stages of the third molar to improve the accuracy of predicting the dental age of subadults in northern China. A total of 340 digital orthopantomograms of subadults aged from 15 to 21 years were analysed. A training group was used to test the accuracy of the original Kvaal's method and to establish novel methods for subadults in northern China. A testing group was used to compare the accuracy of the newly established methods with the Kvaal's original method and with published method specifically used in northern China. To increase the feasibility of our estimation model, we combined the mineralization of the third molar to build a combined specific formula. The results showed that the combined specific model increased the coefficient of determination to 0.513, and the standard error of the estimate was reduced to 1.482 years. We concluded that the combined specific model based on the deposition of secondary dentin and mineralization of the third molar could improve the accuracy of dental age assessment of subadults in northern China. Key Points: The decrease in the dental pulp cavity based on deposition of secondary dentin is a useful variable for assessing age.A total of 340 orthopantomographs were used in this research, including 278 in training groups and 62 in testing groups.Original Kvaal's method underestimated the dental age for subadults in northern China.The equation of combined specific method constructed in our study was proved more suitable to calculate dental age for subadults in northern China.

Minimally Invasive Carinal Reconstruction Using Bronchial Flap and Omental Flap Reinforcement.

Carinal reconstruction and omental flap harvesting are traditionally performed through open approaches. We report a case in which carinal reconstruction with bronchial flap and omental flap reinforcement was performed using minimally invasive approaches. The omental flap was harvested laparoscopically and wrapped around the anastomosis, which reduced the risk of airway anastomosis complications. Noncircumferential resection and reconstruction used bronchial flap, which made it easier to perform under video-assisted thoracoscopic surgery conditions. Minimally invasive carinal reconstruction with bronchial flap and omental reinforcement after neoadjuvant treatment can be safely performed.

Comparing the accuracy of Demirjian and Nolla methods and establishing a new method for dental age estimation in northeastern Chinese children.

Dental age estimation plays an important role in the field of clinic medicine and forensic medicine. The Demirjian and Nolla methods are common scoring methods for dental age estimation but there was no research about the comparison of accuracy of these two methods in northeastern Chinese children. Hence, in this study, we compared the accuracy of these two methods to explore more suitable method for our studied population. We collected 535 orthopantomograms from northern Chinese children aged from 6 to 15 years and divided them into training dataset and testing dataset according to the ratio of 7:3. The dental age of training dataset were estimated using Demirjian and Nolla methods, respectively. The results suggested that the mean differences of these two methods were 0.24 and -0.40 years, and mean absolute difference were 0.65 and 0.59 years. Then to further improve the accuracy of dental age assessment, the new improved formulas and dental age conversion tables were established after analyzing the relationship between the sum scores based on Nolla method and chronology age in training dataset. According to the new method used in testing dataset, the minimum value of mean difference (0.00) and mean absolute difference (0.49) were obtained, which are largely smaller than that of Demirjian and Nolla methods. The new developed method and dental age conversion scales may be more suitable dental age estimation method for northeastern Chinese children.

The application of short and highly polymorphic microhaplotype loci in paternity testing and sibling testing of temperature-dependent degraded samples.

Paternity testing and sibling testing become more complex and difficult when samples degrade. But the commonly used genetic markers (STR and SNP) cannot completely solve this problem due to some disadvantages. The novel genetic marker microhaplotype proposed by Kidd's research group combines the advantages of STR and SNP and is expected to become a promising genetic marker for kinship testing in degraded samples. Therefore, in this study, we intended to select an appropriate number of highly polymorphic SNP-based microhaplotype loci, detect them by the next-generation sequencing technology, analyze their ability to detect degraded samples, calculate their forensic parameters based on the collected 96 unrelated individuals, and evaluate their effectiveness in paternity testing and sibling testing by simulating kinship relationship pairs, which were also compared to 15 STR loci. Finally, a short and highly polymorphic microhaplotype panel was developed, containing 36 highly polymorphic SNP-based microhaplotype loci with lengths smaller than 100 bp and A e greater than 3.00, of which 29 microhaplotype loci could not reject the Hardy-Weinberg equilibrium and linkage equilibrium after the Bonferroni correction. The CPD and CPE of these 29 microhaplotype loci were 1-2.96E-26 and 1-5.45E-09, respectively. No allele dropout was observed in degraded samples incubated with 100°C hot water for 40min and 60min. According to the simulated kinship analysis, the effectiveness at the threshold of 4/-4 reached 98.39% for relationship parent-child vs. unrelated individuals, and the effectiveness at the threshold of 2/-2 for relationship full-sibling vs. unrelated individuals was 93.01%, which was greater than that of 15 STR loci (86.75% for relationship parent-child vs. unrelated individuals and 81.73% for relationship full-sibling vs. unrelated individuals). After combining our 29 microhaplotype loci with other 50 short and highly polymorphic microhaplotype loci, the effectiveness values at the threshold of 2/-2 were 82.42% and 90.89% for relationship half-sibling vs. unrelated individuals and full-sibling vs. half-sibling. The short and highly polymorphic microhaplotype panel we developed may be very useful for paternity testing and full sibling testing in degraded samples, and in combination with short and highly polymorphic microhaplotype loci reported by other researchers, may be helpful to analyze more distant kinship relationships.