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In addition to commonly used short tandem repeats (STRs) and single nucleotide polymorphisms (SNPs), insertion and deletion polymorphisms (InDels) have considerable potential in the field of forensic genetics because they combine desirable characteristics of both STRs and SNPs. In the present study, the SifaInDel 45plex system was designed to amplify 45 InDel markers, including 27 autosomal InDels (A-InDels), 16 X chromosome InDels (X-InDels) and two Y chromosome InDels (Y-InDels), simultaneously in a single PCR procedure and then detect products by capillary electrophoresis (CE). We also optimized the PCR conditions for the novel panel and performed several validation studies including repeatability/reproducibility, concordance, accuracy, sensitivity, stability, species specificity and population genetics. The results confirmed that full profiles could be obtained from ≥62.5 pg of input DNA and from a series of challenging samples encountered in routine casework. The SifaInDel 45plex panel could tolerate different concentrations of inhibitors, such as ≤50 μM hematin, ≤20 ng/μL nigrosine and ≤8000 ng/μL urea. In a population investigation, for the 27 A-InDels, the combined power of discrimination (CPD) exceeded 0.999999, and the combined power of exclusion in duos (CPED) and trios (CPET) was 0.955118 and 0.997754, respectively. For the 16 X-InDels, the combined PDMale and PDFemale was computed as 0.999845 and 0.999998, respectively, and the combined mean exclusion chance in father/daughter or mother/son duos (MECDuo) and mean exclusion chance in standard trios involving daughters (MECTrio) was 0.976220 and 0.998163, respectively. In addition, the two Y-InDels could play a role in correctly determining gender. Overall, the established SifaInDel 45plex panel is a well-performing, reliable and robust multiplex system that stands out for combining a considerable number of A-indels, X-indels and Y-indels based on a CE platform. The population study results also demonstrated that the SifaInDel 45plex panel could be a valid complementary approach for human identification and complex kinship analysis.

Growing evidence suggests that somatic hypermutational status and programmed cell death-1 overexpression are potential predictive biomarkers indicating treatment benefits from immunotherapy using immune checkpoint inhibitors. However, biomarker-matched trials are still limited, and many of the genomic alterations remain difficult to target. To isolate the potential somatic hypermutational tumor from microsatellite instability low/microsatellite stability (MSI-L/MSS) cases, we employed two commercial kits to determine MSI and forensic short tandem repeat (STR) alternations in 250 gastrointestinal (GI) tumors. Three types of forensic STR alternations, namely, allelic loss, Aadd, and Anew, were identified. 62.4% (156/250) of the patients with GI exhibited STR alternation, including 100% (15/15) and 60% (141/235) of the microsatellite high instability and MSI-L/MSS cases, respectively. 30% (75/250) of the patients exhibited STR instability with more than 26.32% (26.32%-84.21%) STR alternation. The cutoff with 26.32% of the STR alternations covered all 15 MSI cases and suggested that it might be a potential threshold. Given the similar mechanism of the mutations of MSI and forensic STR, the widely used forensic identifier STR kit might provide potential usage for identifying hypermutational status in GI cancers.

We aimed to analyze factors related to human immunodeficiency virus (HIV) infection among 15 to 25-year-old rural unmarried Yi adolescents in Liangshan Yi Autonomous Prefecture from 2011 to 2016, and to investigate the relationship among the HIV-related risk factors.Three sites (Butuo, Zhaojue, and Ganluo counties) with a Yi population were selected from 2011 to 2016 in Liangshan prefecture. We used questionnaires to investigate demographic and behavioral characteristics and performed HIV testing. Logistic regression analysis and multiple correspondence analysis (MCA) were used to analyze the data.A total of 6311 qualified rural unmarried Yi adolescents were involved in this study, with an HIV infection rate of 4.18% (264/6311). Logistic regression analysis indicated that males (OR = 1.56, 95% CI = 1.17-2.08), illiteracy (OR = 2.70, 95% CI = 1.79-4.08), primary school education or below (OR = 1.62, 95% CI = 1.12-2.36), history of out-migrating for work (OR = 1.58, 95% CI = 1.18-2.13), and injecting drug user (IDU) (OR = 6.42, 95% CI = 3.06-13.46) contributed to the risk of HIV infection. MCA revealed a strong interrelationship between the following risk factors and HIV infection: aged 23 to 25 years, IDU, history of out-migrating for work, and ever having sexual behaviors.HIV infection among 15 to 25-year-old rural unmarried Yi adolescents in Liangshan is a serious issue. MCA and logistic regression analysis can be used to assess the relationship between risk factors for HIV infection.

Aim: Due to the difference in epidemiology and outcomes between eastern and western populations with Type 2 diabetes mellitus (T2DM), an important challenge is determining how useful the outcomes from diabetes models based on western populations are for eastern patients. Consequently, the principal aim of this study was to develop and validate a Health Policy Model (Chinese Outcomes Model for T2DM [COMT]) for supporting Chinese medical and health economic studies. Methods: The model is created to simulate a series of important complications of T2DM diabetes based on the latest Risk Equations for Complications of Type 2 Diabetes, which was adjusted by adding the adjustment regulator to the linear predictor within the risk equation. The validity of the model was conducted by using a total of 171 validation outcomes from seven studies in eastern populations and ten studies in western populations. The simulation cohorts in the COMT model were generated by copying each validation study’s baseline characteristics. Concordance was tested by assessing the difference between the identity (45°) line and the best-fitting regression of the scatterplots for the predicted versus observed outcomes. Results: The slope coefficients of the best-fitting regression line between the predicted and corresponding observed actual outcomes was 0.9631 and the R 2 was 0.8701. There were major differences between western and eastern populations. The slope and R 2 of predictions were 0.9473 and 0.9272 in the eastern population and 1.0566 and 0.8863 in the western population, which showed more perfect agreement with the observed values in the eastern population than the western populations. The subset of macro-vascular and micro-vascular outcomes in the eastern population showed an identical tendency (the slope coefficient was close to 1), and mortality outcomes showed a slight tendency toward overestimation (the slope coefficient was close to 0.9208). Some degree of underprediction of macro-vascular and micro-vascular end points and overprediction of mortality end point was found in the western population. Conclusion: The COMT diabetes model simulated the long-term patient outcomes observed in eastern Asian T2DM patients with prediction accuracy. This study supports the COMT as a credible tool for Chinese healthcare decision makers. Further work is necessary to incorporate new local data to improve model validity and credibility.

In present study, forensic parameters based on the population data of nineteen autosomal short tandem repeats (STRs) loci were estimated in Chinese Han populations from three riverfront areas of Yangtze Basin in Eastern China. The combined power of discrimination (CPD) were 0.9999999999999999999999883, 0.9999999999999999999999846 and 0.9999999999999999999999863, respectively. The combined power of exclusion (CPE) were 0.999999992248957, 0.999999989800400 and 0.999999989799819, respectively. Furthermore, the genetic relationships among our studied populations and 32 previously investigated populations residing in other areas of China were also analyzed based on pairwise genetic distances (FST). The multidimensional scaling (MDS) plot and phylogenetic tree indicated that our studied populations were majorly clustered with the Han populations along the Yangtze Basin and populations to the north of it. In summary, our results supported that the 19 autosomal STR loci, which were polymorphic and informative in forensic identification, could also be applied in population genetics studies.

We evaluated HIV prevalence among unmarried youth in rural areas of Liangshan Prefecture in southwest China over the period of 2011-2015, and the interactions between behaviors associated with HIV risk. We used HIV sentinel surveillance (HSS) data for our analysis. We used questionnaires to investigate information among unmarried youth aged 15 to 25 years and performed HIV testing. Multivariate logistic regression and path analysis were applied to explore associations between behavioral factors related with HIV infection. A total of 9536 unmarried youth were enrolled in this study, of whom 2.75% (262/9536) tested positive for HIV infection. Multivariate logistic regression showed that male gender, Yi ethnicity, illiteracy, out-migrating for work, needle sharing, and engaging in high-risk sexual behaviors were risk factors. Path analysis revealed that needle sharing had the greatest effect on HIV infection (0.267), followed by drug use (0.171), out-migrating for work (0.079), and high-risk sexual behavior (0.064). The epidemic of HIV infection among unmarried youth in rural areas of Liangshan Prefecture is severe, and prevention efforts should focus on high-risk groups such as youth who are male, Yi, illiterate, and out-migrating for work, and those who engage in high-risk sexual behaviors and needle sharing.

Hairs are commonly encountered at crime scenes. Determining the biological origins of hairs is important for case investigations. However, the DNA extraction of rootless hair shafts are quite difficult. In this study, an enhanced DNA amplification methodology is revealed, in combination with a self-developed system of 5 × PCR Reaction Master Mix IV, to directly amplify the mtDNA 12S ribosomal RNA (12S rRNA) gene from rootless hair shafts collected from 12 common species (human, chicken, duck, cattle, sheep, dog, mouse, pig, horse, cat, rabbit and pigeon) without DNA extraction. The amplicons were subsequently sequenced on Pyromark Q48 (QIAGEN, Hilden, Germany). The results illustrated that the direct amplification system could efficiently amplify the targeted mtDNA genes and pyrosequencing technology could further readily differentiate the sample origin of the rootless hair shafts. Data analysis of the 12S rRNA was equipped to an automatic analysis module. This study provides a new solution for species identification of rootless hair shafts.

Over the last decade, massively parallel sequencing (MPS) has been rapidly developed, bringing many advantages to forensic genomics. It could simultaneously target multiple genetic markers including short tandem repeats (STRs), insertion and deletion polymorphisms (InDels), and single nucleotide polymorphisms (SNPs). In this study, we developed an assay which can simultaneously sequence 22 Y-STRs and 26 Y-SNPs on the MiSeq FGx™ Forensic Genomics System (Illumina, Inc., San Diego, USA). The primers of the markers were designed using the online DesignStudio (Illumina). Subsequently, DNA libraries were established with the TruSeq Amplicon method. For the pilot study, we sequenced 25 unrelated Chinese Han males. A total of 125 sequence-based alleles were observed at the 22 Y-STR loci, with the average depth of coverage (DoC) ranging from 314.9 x to 637.7 × . For the concordant study, the sequencing results of these samples were fully consistent with genotypes called via capillary electrophoresis (CE). For the SNPs, a preferable performance was observed with the average DoC of 836.5 × . However, inter-locus imbalance was observed at some of the markers, which needs substantial improvement of the primer pool. In general, this customized Y-chromosome STR and SNP panel on the MiSeqFGx™ Forensic Genomics System can provide an effective solution for these markers on Y chromosome, which may allow more forensically relevant information to be obtained from limited forensic materials, such as those from sexual assault and rape cases. Besides, it could also facilitate the forensic practices including paternity testing, genetic determination of pedigree and inter-population migration integration.

MicroRNA (miRNA) expression profiling is gaining interest in the forensic community because the intrinsically short fragment and tissue-specific expression pattern enable miRNAs as a useful biomarker for body fluid identification. Measuring the quantity of miRNAs in forensically relevant body fluids is an important step to screen specific miRNAs for body fluid identification. The recent introduction of massively parallel sequencing (MPS) has the potential for screening miRNA biomarkers at the genome-wide level, which allows both the detection of expression pattern and miRNA sequences. In this study, we employed the Ion Personal Genome Machine(®) System (Ion PGM™ System, Thermo Fisher) to characterize the distribution and expression of 2588 human mature miRNAs (miRBase v21) in 5 blood samples and 5 saliva samples. An average of 1,885,000 and 1,356,000 sequence reads were generated in blood and saliva respectively. Based on miRDong, a Perl-based tool developed for semi-automated miRNA distribution designations, and manually ascertained, 6 and 19 miRNAs were identified respectively as potentially blood and saliva-specific biomarkers. Herein, this study describes a complete and reliable miRNA workflow solution based on Ion PGM™ System, starting from efficient RNA extraction, followed by small RNA library construction and sequencing. With this workflow solution and miRDong analysis it will be possible to measure miRNA expression pattern at the genome-wide level in other forensically relevant body fluids.

TMEM16A is the molecular basis of calcium-activated chloride channels and shows Ca(2+)-dependent gating. It is critical to understand how the Ca(2+) sensors dynamically control the gate of TMEM16A. However, the detailed mechanism by which the calcium ions bind and open the channel is still obscure. In this study, the authors confirmed that there are two Ca(2+) sensors which cooperatively couple together in TMEM16A. Our data show that mutations at both Ca(2+)-sensitive domains, E447Y and E702Q-E705Q, weaken the Ca(2+) affinity for TMEM16A channel. The EC50 for WT, E447Y, and E702Q-E705Q are 0.53 ± 0.11, 14.5 ± 0.3, and 26.5 ± 3.6 μM, respectively. The triple mutation, including both of the Ca(2+) sensors, E447Y-E702Q-E705Q, with EC50 as 55.6 ± 5.1 μM, results in much further right-shifted dose response curve than the single sensor's mutations (E447Y, E702Q-E705Q) do, which indicates that there is a cooperation between the two Ca(2+)-sensitive domains. We also found that the divalent cations, both Ca(2+) and Sr(2+), share common mechanism of gating the TMEM16A.

Monozygotic (MZ) twins, considered to be genetically identical, cannot be distinguished from one another by standard forensic DNA testing. A recent study employed whole genome sequencing to identify extremely rare mutations and reported that mutation analysis could be used to differentiate between MZ twins. Compared with nuclear DNA, mitochondrial DNA (mtDNA) has higher mutation rates; therefore, minor differences theoretically exist in MZ twins' mitochondrial genome (mtGenome). However, conventional Sanger-type sequencing (STS) is neither amenable to, nor feasible for, the detection of low-level sequence variants. The recent introduction of massively parallel sequencing (MPS) has the capability to sequence many targeted regions of multiple samples simultaneously with desirable depth of coverage. Thus, the aim of this study was to assess whether full mtGenome sequencing analysis can be used to differentiate between MZ twins. Ten sets of MZ twins provided blood samples that underwent extraction, quantification, mtDNA enrichment, library preparation, and ultra-deep sequencing. Point heteroplasmies were observed in eight sets of MZ twins, and a single nucleotide variant (nt15301) was detected in five sets of MZ twins. Thus, this study demonstrates that ultra-deep mtGenome sequencing could be used to differentiate between MZ twins.

The She people, with a population of about 710,000, are the largest minority in Fujian province. They live primarily in the mountainous areas of Fujian Province and also present in Zhejiang, Jiangxi, Guangdong, Anhui, Hunan and Taiwan. Currently, Y-STR analysis has well characterized and become an essential tool in forensic and population genetics. In the present study, we investigated genetic and phylogenetic relationship between the She population, which included 53 unrelated She males living in the Fujian Province and typed for 26 Y-STR loci, and 13 reference populations. We found that the She population has the closest relationship with the Miao population, followed by the populations from south China.

The growth arrest special 5 (GAS5) is known to be involved in various cancers. However, its expression regulation remains unclear. Polymorphisms in the promoter region of GAS5 may affect its expression and be associated with cancer susceptibility. In this research, we first evaluated the association of a 5-base pair indel polymorphism (rs145204276) in the promoter region of GAS5 with hepatocelluar carcinoma (HCC) susceptibility in Chinese populations. Logistic regression analysis showed that the deletion allele of rs145204276 significantly increased the risk of HCC in two independent case control sets (1034 HCC and 1054 controls). Further genotype-phenotype association analysis revealed that the deletion allele was markedly correlated with higher expression of GAS5 in HCC tissues. The luciferase activity analysis in an in vitro reporter gene system suggested that the deletion allele improved an increased expression of GAS5 in three hepatoma cell lines. Intriguingly, overexpression of GAS5 displayed an anti-apoptosis effect in HCC cell lines, GAS5 knockdown could partially revert this anti-apoptosis effect, suggesting that GAS5 may act as a proto-oncogene in HCC, in contrast with its inhibitory role in other cancers. Further pyrosequencing revealed that the genotypes of rs145204276 were associated with methylation status of GAS5 promoter region. Taken together, our findings provided evidence that rs145204276 may contribute to hepatocarcinogenesis by affecting methylation status of the GAS5 promoter and subsequently its transcriptional activity thus serving as a potential therapy target for HCC.

In this study, a new STR 25-plex typing system, including 23 autosomal STRs (D1S1656, D2S1338, D2S441, D3S1358, D5S818, D6S1043, D7S820, D8S1179, D10S1248, D12S391, D13S317, D16S539, D18S51, D19S433, D21S11, D22S1045, CSF1PO, FGA, Penta D, Penta E, TH01, TPOX, vWA) and a Y-STR locus of DYS391 and amelogenin, was developed. The included 24 STRs belonged to the main international DNA databases (CODIS, ISSL, ESS-extended, UCL, GCL and NCIDD) except D6S1043 (specially chosen for Chinese population). Developmental validation indicated that the STR 25-plex typing system was reproducible, accurate, sensitive and robust. The sensitivity testing of the system was such that a full profile was obtainable even with 125pg of human DNA. Specificity testing was demonstrated by the lack of cross-reactivity with a variety of commonly encountered animal species and microbial pool. For the stability testing, full profiles can been obtained with humic acid concentration≤60ng/μL and hematin

While the QuikChange site-directed mutagenesis method and its later modifications are extremely useful and simple, they suffer from several drawbacks. Here, we propose a new method, named LFEAP mutagenesis (Ligation of Fragment Ends After PCR) for creating various mutations in plasmid by leveraging three existing concepts: inverse PCR, single primer PCR, and sticky-end assembly. The first inverse PCR on the target plasmid yielded linearized DNA fragments with mutagenic ends, and a second single primer PCR resulted in complementary single-stranded DNA fragments with the addition of overhangs at the 5′ end of each strand. The resulting single strands were then annealed to produce double-stranded DNA with free 5′ single-stranded DNA tails. These products with compatible sticky ends were efficiently assembled into a circular, mutagenized plasmid. With this strategy, multiple simultaneous changes (up to 15) and mutations in large plasmids (up to 50 kb) were achieved with high efficiency and fidelity. LFEAP mutagenesis is a versatile method that offers significant advantages for introducing large and multiple changes in plasmid DNA.

Background Yi people make up about 50% of the population in Liangshan Prefecture, Sichuan Province, China, but accounted for 88.07% of new HIV cases in the prefecture from 2011 to 2013. This study evaluated HIV prevalence in pregnant women of Liangshan Prefecture using HIV sentinel surveillance (HSS) data over the period of 2009 to 2015. Methods Xichang, Zhaojue County, and Butuo County were selected as HSS sites. We investigated the temporal trends in HIV prevalence in these areas, and the association between demographic and behavioral characteristics and risk of HIV infection. Results Data on a total of 2797 pregnant women in Xichang and 3983 pregnant women in Zhaojue and Butuo was collected for the period 2009 to 2015. There was a fluctuating HIV prevalence among pregnant women of Xichang, with a rate of 0.75% in 2015 (χ2trend = 2.27, P = 0.13). HIV prevalence among pregnant women of Zhaojue and Butuo was consistently high, varying between 3.4% (9/267, 2011) and 10.3% (82/796, 2012) over the period of 2010 to 2015 (χ2trend = 0.12, P = 0.73). In Xichang, we found that Yi ethnicity (OR = 11.37, 95% CI = 2.92-44.25) and a husband who used drugs (OR = 32.13, 95% CI = 5.33-193.67) were significantly associated with HIV risk in pregnant women. For Zhaojue and Butuo, we observed that pregnant women had a higher risk of HIV infection when they were over 30 years old (OR = 1.72, 95% CI = 1.17-2.52), when they had a higher number of previous births, when their husbands had a history of migrating for work (OR = 1.56, 95% CI = 1.16-2.08), and when they had a history of other sexually transmitted infections (OR = 2.19, 95% CI = 1.16-2.08). Compared to those with a primary school education or below, pregnant women with a secondary or high school education level had a lower risk of HIV infection (OR = 0.28, 95% CI = 0.09-0.89). Conclusion Our results indicate that there is a serious HIV epidemic among pregnant Yi women, especially for those with less education, more past births, or a husband with a history of out-migrating for work or STD infection.

Purpose To compare the axial length (AL), anterior chamber depth (ACD) and intraocular lens power (IOLP) of IOLMaster and Ultrasound in normal, long and short eyes. Methods Seventy-four normal eyes (≥ 22 mm and ≤ 25 mm), 74 long eyes (> 25 mm) and 78 short eyes (< 22 mm) underwent AL and ACD measurements with both devices in the order of IOLMaster followed by Ultrasound. The IOLP were calculated using a free online LADAS IOL formula calculator. Results The difference in AL and IOLP between IOLMaster and Ultrasound was statistically significant when all three groups were combined. The difference in ACD between IOLMaster and Ultrasound was statistically significant in the normal group (P

PURPOSE: To investigate the wound healing changes of clear corneal incisions performed with a femtosecond laser or a steel bevel knife in cataract surgery using spectral-domain optical coherence tomography. METHODS: Prospectively, consecutive patients undergoing either femtosecond laser–assisted cataract surgery (femtosecond laser group) using the LenSx laser (Alcon Laboratories, Inc., Fort Worth, TX) or traditional cataract surgery using steel blades (control group) were enrolled. The incidence of posterior wound gape, Descemet's membrane detachment, posterior wound retraction, and the inner and outer corneal incision thicknesses were assessed. RESULTS: Compared to the control group, the femtosecond laser group had significantly lower incidences of posterior wound gape at 1 day ( P = .012) and Descemet's membrane detachment at 1 week ( P = .030), 1 month ( P = .048), and 3 months ( P = .048), and a significantly higher incidence of posterior wound retraction at 1 month ( P < .001) and 3 months ( P < .001) postoperatively. The inner and outer corneal incision thicknesses decreased over time in both groups. CONCLUSIONS: The clear corneal incisions created by femtosecond laser demonstrated a lower incidence of posterior wound gape and Descemet's membrane detachment. However, the higher posterior wound retraction incidence, which indicated that there was wound remodeling, was apparent. [ J Refract Surg. 2018;34(1):17–22.]

Sudden cardiac death (SCD) describes the unexpected natural death from a cardiac cause within a short time period. Compelling evidence suggests the involvement of host genetic factors in SCD etiology. Identification of genetic variations predisposed to SCD enables genetic testing that may contribute to SCD diagnosis and risk stratification. Previous studies have suggested that dysregulation of pro-alpha2 chain of type I collagen, encoded by collagen type I alpha 2 chain ( COL1A2 ) gene, was involved in cardiac disorders such as myocardial infarction, hypertrophic cardiomyopathy and atherosclerosis. By using a candidate-gene-based approach, we evaluated the association of a 7-base pair (7-bp) indel polymorphism (rs3917) in the 3′UTR of COL1A2 with the risk of SCD in a Chinese population (79SCD cases and 328 controls). Logistic regression analysis showed that the deletion allele of rs3917 significantly increased the risk of SCD [odds ratio (OR)=1.82; 95% confidence interval (CI)=1.08–3.06; P =0.0159]. Further genotype-phenotype association analysis revealed that the deletion allele was markedly correlated with lower expression of COL1A2 in human myocardium tissues. The luciferase activity analysis in an in vitro reporter gene system suggested that rs3917 could regulate COL1A2 expression through interrupting the binding of miR-296-3p with COL1A2 in an allele-dependent manner, which in turn confer SCD risk. Our data provided initial evidence that rs3917 was highly relevant to SCD susceptibility, and this indel may become a potential marker for molecular diagnosis and genetic counseling of SCD. The replication of our studies and further functional studies are needed to validate our findings.

Background Insulin therapy should be strongly considered in newly diagnosed type 2 diabetic (T2D) patients with severe hyperglycemia. However, whether insulin be continued or patients switched to an oral antidiabetic drug (OAD) after short-term intensive insulin therapy (ITT) is not clear. Methods After ITT for 10–14 days, 47 patients were randomized into either a glargine (n = 21) or metformin-based OAD (n = 26) group for a 6-month intervention. After these 6 months, patients in the glargine group were switched to metformin-based OAD therapy, whereas treatment in the OAD group was unchanged. Patients were followed-up for another 6 months. Results At the end of the 6-month intervention, HbA1c was similarly reduced in both the glargine (from 11.81 ± 1.70% to 6.48 ± 0.79%; P

Second generation sequencing offers the opportunity to analyse forensic DNA samples and obtain massively parallel coverage of targeted fragments. 124 SNPs included in HID-Ion Ampliseq™ panel for human identification were evaluated on the Ion torrent PGM Sequencer System this time. Reference sample of 9948 and blood samples of 45 unrelated individuals were involved for study. Concordance study was performed with reference sample of 9948 between NGS and Sanger sequencing. Full concordance were obtained, however, 3 SNP loci (rs7520386, rs214955 and rs4530059) were detected with F MAR (%) values above 60% for heterozygote. In the SGS data analysis, F MAR values for accurate heterozygotes should be range from 50% to 60%, for homozygotes or Y-SNP should be above 90%. SNPs of rs7520386, rs4530059, rs214955, rs1523537, rs2342747, rs576261 and rs12997453 were recognized as poorly performing loci, either with allelic imbalance or with lower coverage. For the genetic analysis, 8 SNPs (rs2056277, rs1886510, rs740910, rs717302, rs251934, rs938283, rs729172 and rs733164) with lower polymorphism in HAN population (PIC −23 with the 90 auto-SNPs in HAN population of China. And when using the Variant Caller plug-in, variants close to target sites of rs1490413, rs12997453, rs727811, rs6955448, rs1015250, rs10776839, rs2342747 and rs430046 were observed. This preliminary study has demonstrated considerable potential of Ion Torrent PGM as a low to medium throughput SGS platform for SNP detection and HID-Ion Ampliseq™ panel as a commercial tool for human identification in HAN population from China.

The identification of the biological origin of a specimen is necessary in some forensic casework. Mitochondrial DNA (mtDNA) has become a useful tool in forensic identification because of the high number of copies and the lack of recombination. In this study, we amplified and sequenced a 100-bp fragment of the mtDNA 12S ribosomal RNA (12S rRNA) gene from 17 different species including human, monkey, chicken, duck, cattle, sheep, dog, SD rat, mouse, goose, pig, horse, cat, rabbit, donkey, pigeon, and deer. The amplified products were sequenced for the specimen identification by pyrosequencing technique. The results showed that the specimen origin could be accurately determined by only 12 bases. Through pyrosequencing the 12S rRNA gene, a new method for discrimination of 17 species was developed.

Renal tubular epithelial cell proliferation and transepithelial cyst fluid secretion are key features in the progression of polycystic kidney disease (PKD). As the role of the apical renal sodium-glucose cotransporters in these processes is not known, we tested whether phlorizin inhibits cyst growth and delays renal disease progression in a rat model of PKD. Glycosuria was induced by subcutaneous injection of phlorizin in male heterozygous (Cy/+) and wild-type Han:SPRD rats. Phlorizin induced immediate and sustained glycosuria and osmotic diuresis in these rats. Cy/+ rats treated with phlorizin for 5 weeks showed a significant increase in creatinine clearance, a lower 2-kidneys/body weight ratio, a lower renal cyst index, and reduced urinary albumin excretion as compared with vehicle-treated Cy/+ rats. Measurement of Ki67 staining found significantly lower cell proliferation in dilated tubules and cysts of Cy/+ rats treated with phlorizin, as well as a marked inhibition of the activated MAP kinase pathway. In contrast, the mTOR pathway remained unaltered. Phlorizin dose dependently inhibited MAP kinase in cultured tubular epithelial cells from Cy/+ rats. Thus, long-term treatment with phlorizin significantly inhibits cystic disease progression in a rat model of PKD. Hence, induction of glycosuria and osmotic diuresis (glycuresis) by renal sodium-glucose cotransporters inhibition could have a therapeutic effect in polycystic kidney disease.

The study aimed to analyze the relationship between metabolic variables and estimated glomerular filtration rate (eGFR) and explore the potential risk factors for a mildly reduced eGFR in a community-based population.Cross-sectional study in 643 adults without a history of kidney disease whose eGFR levels were greater than 60 mL/min/1.73 m(2) according to the Chronic Kidney Disease Epidemiology Collaboration equation (CKD-EPI). Anthropometric measurements, blood pressure, fasting lipid profile and levels of fasting and post-load glucose, insulin, serum creatinine and uric acid (UA) were tested. The eGFR was calculated, and the correlations between eGFR and each variable were analyzed.The subjects were divided into two groups by using 90 mL/min/1.73 m(2) as the cut-off value of the eGFR. In the lower eGFR group, the age, systolic blood pressure (SBP), diastolic blood pressure (DBP), body mass index (BMI), waist circumference (WC), total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), fasting plasma glucose (FPG), 2 h post-load plasma glucose (2 h-PG) levels and UA were significantly increased, and the incidences of hypertension, diabetes, obesity, hypertriglyceridemia and hypercholesterolemia were also higher (P0.05). A multiple linear stepwise regression analysis showed that the WC, SBP, FPG and UA were independently correlated with the eGFR after adjusting for the other covariables.The WC, SBP, FPG and UA were closely related to the eGFR in the subjects whose eGFR levels were greater than 60 mL/min/1.73 m(2). The increased WC, SBP, FPG and UA may be the main risk factors for a mildly reduced eGFR.

Sequencing validation of assemble with various overhangs. (a)-(g) various overhang sizes; (h) overhang designed as 5â ˛ end of G/C; (i) overhang designed as 5â ˛ end of A/T. Overhang regions were marked by red dashed line rectangles. (DOCX 2777Â kb)

Comparisons of AFEAP cloning with common DNA assembly methods. (DOCX 23Â kb)

Sequencing validation of number of fragments characterization. The join sites were shown as S1 to S13, and number of fragments for assembly was shown as 2 + V to 12 + V. The overhang sequences were shown. V: vector backbone. (DOCX 11884 kb)

PCR thermocycling conditions. (DOCX 13Â kb)

Supporting Information. (DOCX 32Â kb)

Primers used in this work. (DOCX 32Â kb)

PCR product reannealing conditions. (DOCX 14Â kb)

Sequencing validation of plasmid sizes characterization. Five join sites are S1, S2, S3, S4, and S5. (a) 11.5 kb plasmid; (b) 19.6 kb plasmid; (c) 28 kb plasmid; (d) 34.6 kb plasmid. The overhang sequences were shown. (DOCX 3815 kb)

PCR conditions. (DOCX 14Â kb)

In this study, a total of 484 unrelated healthy individuals and 359 two-generation families in the Han population in Shanghai, China were successfully analyzed with the Investigator HDplex Kit (Qiagen, Hilden, Germany). Hardy-Weinberg equilibrium tests demonstrated no significant deviation from expected values (p > 0.05) for the 12 autosomal STRs included in this kit. A total of 173 alleles were detected in 12 autosomal STR loci; the largest number of alleles (32) was detected for the SE33 locus. The combined power of discrimination was 0.99999999992. The combined mean exclusion chance in duo cases was 0.99982, whereas the combined mean exclusion chance in trio cases was 0.9999986. The average mutation rate across all loci was 0.0023 (95% confidence intervals: 0.0013-0.0039). These results suggest that the 12 autosomal STR loci can provide highly informative polymorphic data for paternity testing and forensic identification in the Han population in Shanghai, China.

China has repeatedly been the subject of genetic studies to elucidate its prehistoric and historic demography. While some studies reported a genetic distinction between Northern and Southern Han Chinese, others showed a more clinal picture of small differences within China. Here, we investigated the distribution of Y chromosome variation along administrative as well as ethnic divisions in the mainland territory of the People's Republic of China, including 28 administrative regions and 19 recognized Chinese nationalities, to assess the impact of recent demographic processes. To this end, we analyzed 37,994 Y chromosomal 17-marker haplotype profiles from the YHRD database with respect to forensic diversity measures and genetic distance between groups defined by administrative boundaries and ethnic origin. We observed high diversity throughout all Chinese provinces and ethnicities. Some ethnicities, including most prominently Kazakhs and Tibetans, showed significant genetic differentiation from the Han and other groups. However, differences between provinces were, except for those located on the Tibetan plateau, less pronounced. This discrepancy is explicable by the sizeable presence of Han speakers, who showed high genetic homogeneity all across China, in nearly all studied provinces. Furthermore, we observed a continuous genetic North-South gradient in the Han, confirming previous reports of a clinal distribution of Y chromosome variation and being in notable concordance with the previously observed spatial distribution of autosomal variation. Our findings shed light on the demographic changes in China accrued by a fast-growing and increasingly mobile population.

Kir2.1 plays key roles in setting rest membrane potential and modulation of cell excitability. Mutations of Kir2.1, such as D172N or E299V, inducing gain-of-function, can cause type3 short QT syndrome (SQT3) due to the enlarged outward currents. So far, there is no clinical drug target to block the currents of Kir2.1. Here, we identified a novel blocker of Kir2.1, styrax, which is a kind of natural compound selected from traditional Chinese medicine. Our data show that styrax can abolish the inward and outward currents of Kir2.1. The IC50 of styrax for WT, D172N and E299V are 0.0113 ± 0.00075, 0.0204 ± 0.0048 and 0.0122 ± 0.0012 (in volume), respectively. The results indicate that styrax can serve as a novel blocker for Kir2.1.

Utilizing massively parallel sequencing (MPS) technology for SNP testing in forensic genetics is becoming attractive because of the shortcomings of STR markers, such as their high mutation rates and disadvantages associated with the current PCR-CE method as well as its limitations regarding multiplex capabilities. MPS offers the potential to genotype hundreds to thousands of SNPs from multiple samples in a single experimental run. In this study, we designed a customized SNP panel that includes 273 forensically relevant identity SNPs chosen from SNPforID, IISNP, and the HapMap database as well as previously related studies and evaluated the levels of genotyping precision, sequence coverage, sensitivity and SNP performance using the Ion Torrent PGM. In a concordant study of the custom MPS-SNP panel, only four MPS callings were missing due to coverage reads that were too low (

Aim The aim of the study was to investigate the interchangeability and reliability of macular perfusion measurements using optical coherence tomography angiography. Methods A prospective cross-sectional observational study. Healthy adult Chinese subjects were recruited. Macular perfusion parameters were automatically analysed by software included in a spectral-domain optical coherence tomography system. The vessel density (VD) of the whole, parafovea, superior-hemi, inferior-hemi, fovea, temporal, superior, nasal and inferior quadrants as well as the foveal avascular zone (FAZ) and choroidal capillary VD (CCVD) were quantified. Results A total of 51 eyes in 27 subjects were included (8 men and 19 women, mean age 24±4 years ). Significant differences in VD of all quadrants (all p Conclusions FAZ and CCVD are interchangeable between the 3×3 mm and 6×6 mm macular scan sizes. The VD differences between the two different scan sizes are not clinically meaningful. The macular perfusion parameters presented good but not perfect reliability, which should be acknowledged in clinical practice.

Insertion/deletion (InDels) markers can serve as a useful supporting tool to short tandem repeat (STR) typing systems for human identification. The Qiagen DIPplex Investigator kit, which contains 30 biallelic autosomal InDels and amelogenin, has been developed for forensic use. To estimate the genetic diversity of the 30 markers in Han Chinese individuals living in Zhejiang and to further evaluate their applicability in forensic science, 246 unrelated Han Chinese from Zhejiang were genotyped at these loci. No significant departures from Hardy-Weinberg equilibrium were observed at these loci in these participants. The combined power of discrimination was over 0.99999999 and the combined probability of exclusion was over 0.9901. Results demonstrated that the 30 InDel markers could be used as a supporting tool for the human identification of specific Han Chinese individuals from Zhejiang. The genetic differences and phylogenetic relationships among Han Chinese from Zhejiang, Han Chinese from five other areas, nine minority ethnic groups, as well as two other East Asian populations were also investigated. Two InDel markers, HLD39 and HLD40, showed significant allele-frequency differences between Han Chinese from Zhejiang and ethnic minorities. Further analysis can be used to evaluate their role in forensic science.

Transformation of macrophages into foam cells plays a critical role in the pathogenesis of atherosclerosis. The aim of this study was to determine the expression and biological roles of microRNA (miR)-150 in the formation of macrophage foam cells and to identify its functional target(s). Exposure to 50 μg/ml oxidized low-density lipoprotein (oxLDL) led to a significant upregulation of miR-150 in THP-1 macrophages. Overexpression of miR-150 inhibited oxLDL-induced lipid accumulation in THP-1 macrophages, while knockdown of miR-150 enhanced lipid accumulation. apoA-I- and HDL-mediated cholesterol efflux was increased by 66% and 43%, respectively, in miR-150-overexpressing macrophages relative to control cells. In contrast, downregulation of miR-150 significantly reduced cholesterol efflux from oxLDL-laden macrophages. Bioinformatic analysis and luciferase reporter assay revealed adiponectin receptor 2 (AdipoR2) as a direct target of miR-150. Small interfering RNA-mediated downregulation of AdipoR2 phenocopied the effects of miR-150 overexpression, reducing lipid accumulation and facilitating cholesterol efflux in oxLDL-treated THP-1 macrophages. Knockdown of AdipoR2 induced the expression of proliferator-activated receptor gamma (PPARγ), liver X receptor alpha (LXRα), ABCA1, and ABCG1. Moreover, pharmacological inhibition of PPARγ or LXRα impaired AdipoR2 silencing-induced upregulation of ABCA1 and ABCG1. Taken together, our results indicate that miR-150 can attenuate oxLDL-induced lipid accumulation in macrophages via promotion of cholesterol efflux. The suppressive effects of miR-150 on macrophage foam cell formation are mediated through targeting of AdipoR2. Delivery of miR-150 may represent a potential approach to prevent macrophage foam cell formation in atherosclerosis.