AIM:To estimate if nanopore targeted sequencing(NTS)could identify pathogens causing postoperative endophthalmitis and further determine the feasibility of clinical application of NTS.METHODS:A total of 55 patients(55...AIM:To estimate if nanopore targeted sequencing(NTS)could identify pathogens causing postoperative endophthalmitis and further determine the feasibility of clinical application of NTS.METHODS:A total of 55 patients(55 eyes)with postoperative endophthalmitis were retrospectively included in this study with their medical records.Intraocular fluid samples were examined by NTS and microbial culture.All included patients had undergone examinations including measurement of best corrected visual acuity(BCVA)and intraocular pressure(IOP),slit-lamp biomicroscopy,and indirect ophthalmoscopy;additionally,they underwent B-ultrasound,anterior segment photography,and fundus photography if necessary.RESULTS:Among 55 patients with postoperative endophthalmitis,the age was 65.25±15.04y and there were 30 female(54.54%)patients.Forty-one(74.54%)vitreous humor samples and fourteen(25.45%)aqueous humor samples were sent for both NTS and microbial culture.NTS had a notable higher detection rate than microbial culture in detecting pathogens(90.91%vs 38.18%,χ^(2)=33.409,P<0.001).NTS exhibited high sensitivity of pathogen detection in both microbial culture positive and negative samples(100%and 85.29%,respectively).In 16 of 21(76.19%)patients who showed culture-positivity,their results corresponded with those of NTS.Moreover,in two patients(9.52%),NTS showed a better species resolution than microbial culture;in three patients(14.28%),NTS identified additional pathogens.As for fungus,the positive detection rate of NTS was significantly higher than that of microbial culture(20%vs 3.64%,χ^(2)=7.066,P=0.008).Also,NTS could detect multi-infection by bacteria and fungi than microbial culture(32.73%vs 0,χ^(2)=21.522,P<0.001).NTS could detect bacteria as well as fungi simultaneously within 48h in all patients.Meanwhile,NTS had a shorter detection time than microbial culture(1.13±0.34 vs 2.67±0.55d,Z=-9.218,P<0.001).After the NTS results were obtained,15 patients received additional intravitreal/intracameral anti-infection treatment.At follow-up,there was a statistically significant improvement in the visual acuity relative to the baseline(Z=−5.222,P<0.001).CONCLUSION:NTS can provide rapid identification and highly sensitive detection of pathogens among patients with postoperative endophthalmitis,which can guide anti-infection treatment and improve visual prognosis.展开更多
Genotyping by Target Sequencing(GBTS)technology,known for its flexibility,high efficiency,high throughput,and low cost,has been increasingly employed in molecular breeding.However,there is still limited study on the d...Genotyping by Target Sequencing(GBTS)technology,known for its flexibility,high efficiency,high throughput,and low cost,has been increasingly employed in molecular breeding.However,there is still limited study on the design and development of high-throughput genotyping tools in watermelon.In this study,we identified 112000 high quality SNPs by analyzing the resequencing data of 43 cultivated watermelon accessions.11921 and 6094 SNPs were selected for developing two sets of watermelon liquid-phase chips with different marker densities,named Watermelon 10K and 5K,respectively.Furthermore,the SNPs and Indels of most mapped gene/QTLs for many agronomic important traits in watermelon were also integrated into the two chips for foreground selection.These chips have been tested using GBTS technology in various applications in watermelon.The genotyping of 76 accessions by Watermelon 5K liquid-phase chip showed an average detection rate of 99.28%and 81.78%for cultivated and wild watermelon accessions,respectively.This provided enough markers information for GWAS and two significant QTLs,ssc1.1 and ssc1.2,associated with soluble sugar content were detected.Furthermore,BSA-seq analysis for non-lobed leaf and dwarf traits were validated by liquid-phase chips,and the candidate region was consistent with our previous studies.Additionally,we precisely introduced the Cldw1 and Clbl genes into an elite inbred line WT2 using Watermelon 5K for assisted selection,resulting in the development of three new germplasm with good plant architecture.As a high-throughput genotyping liquid-phase SNP array,the Watermelon 10K and 5K chips will greatly facilitate functional studies and molecular breeding in watermelon.展开更多
Different newborn screening(NBS) programs have been practiced in many countries since the 1960 s. It is of considerable interest whether next-generation sequencing is applicable in NBS. We have developed a panel of 46...Different newborn screening(NBS) programs have been practiced in many countries since the 1960 s. It is of considerable interest whether next-generation sequencing is applicable in NBS. We have developed a panel of 465 causative genes for 596 early-onset, relatively high incidence, and potentially actionable severe inherited diseases in our Newborn Screening with Targeted Sequencing(NESTS) program to screen 11,484 babies in 8 Women and Children’s hospitals nationwide in China retrospectively. The positive rate from preliminary screening of NESTS was 7.85%(902/11,484). With 45.89%(414/902) follow-up of preliminary positive cases, the overall clinically confirmative diagnosis rate of monogenic disorders was 12.07%(50/414), estimating an average of 0.95%(7.85% × 12.07%) clinical diagnosis rate, suggesting that monogenic disorders account for a considerable proportion of birth defects. The disease/gene spectrum varied in different regions of China. NESTS was implemented in a hospital by screening 3923 newborns to evaluate its clinical application. The turn-around time of a primary report, including the sequencing period of < 7 days, was within 11 days by our automatic interpretation pipeline. Our results suggest that NESTS is feasible and cost-effective as a first-tier NBS program, which will change the status of current clinical practice of NBS in China.展开更多
Genotyping by target sequencing(GBTS)integrates the advantages of silicon-based technology(high stability and reliability)and genotyping by sequencing(high flexibility and cost-effectiveness).However,GBTS panels are n...Genotyping by target sequencing(GBTS)integrates the advantages of silicon-based technology(high stability and reliability)and genotyping by sequencing(high flexibility and cost-effectiveness).However,GBTS panels are not currently available in pigs.In this study,based on GBTS technology,we first developed a 50K panel,including 52,000 single-nucleotide polymorphisms(SNPs),in pigs,designated GBTS50K.A total of 6,032 individuals of Large White,Landrace,and Duroc pigs from 10 breeding farms were used to assess the newly developed GBTS50K.Our results showed that GBTS50K obtained a high genotyping ability,the SNP and individual call rates of GBTS50K were 0.997–0.998,and the average consistency rate and genotyping correlation coefficient were 0.997 and 0.993,respectively,in replicate samples.We also evaluated the efficiencies of GBTS50K in the application of population genetic structure analysis,selection signature detection,genome-wide association studies(GWAS),genotyped imputation,genetic selection(GS),etc.The results indicate that GBTS50K is plausible and powerful in genetic analysis and molecular breeding.For example,GBTS50K could gain higher accuracies than the current popular GGP-Porcine bead chip in genomic selection on 2 important traits of backfat thickness at 100 kg and days to 100 kg in pigs.Particularly,due to the multiple SNPs(mSNPs),GBTS50K generated 100K qualified SNPs without increasing genotyping cost,and our results showed that the haplotype-based method can further improve the accuracies of genomic selection on growth and reproduction traits by 2 to 6%.Our study showed that GBTS50K could be a powerful tool for underlying genetic architecture and molecular breeding in pigs,and it is also helpful for developing SNP panels for other farm animals.展开更多
Objective This study aimed to explore the diagnostic value of novel technique-targeted next-generation sequencing(tNGS)of bronchoalveolar lavage fluid(BALF)in pulmonary mycobacterial infections.Methods This retrospect...Objective This study aimed to explore the diagnostic value of novel technique-targeted next-generation sequencing(tNGS)of bronchoalveolar lavage fluid(BALF)in pulmonary mycobacterial infections.Methods This retrospective study was conducted on patients who underwent bronchoscopy and tNGS,smear microscopy,and mycobacterial culture of BALF.Patients with positive Mycobacterium tuberculosis(MTB)culture or GeneXpert results were classified into the tuberculosis case group.Those diagnosed with nontuberculous mycobacteria(NTM)-pulmonary disease(NTM-PD)composed the case group of NTM-PD patients.The control group comprised patients without tuberculosis or NTM-PD.Sensitivity,specificity,and receiver operating characteristic(ROC)curves were used to evaluate the diagnostic performance.Results For tuberculosis patients with positive mycobacterial culture results,the areas under the ROC curves(AUCs)for tNGS,GeneXpert,and smear microscopy were 0.975(95%CI:0.935,1.000),0.925(95%CI:0.859,0.991),and 0.675(95%CI:0.563,0.787),respectively.For tuberculosis patients with positive GeneXpert results,the AUCs of tNGS,culture,and smear microscopy were 0.970(95%CI:0.931,1.000),0.850(95%CI:0.770,0.930),and 0.680(95%CI:0.579,0.781),respectively.For NTM-PD,the AUCs of tNGS,culture,and smear-positive but GeneXpert-negative results were 0.987(95%CI:0.967,1.000),0.750(95%CI:0.622,0.878),and 0.615(95%CI:0.479,0.752),respectively.The sensitivity and specificity of tNGS in NTM-PD patients were 100%and 97.5%,respectively.Conclusion tNGS demonstrated superior diagnostic efficacy in mycobacterial infection,indicating its potential for clinical application.展开更多
46,XY disorders of sex development(DSD)is characterized by incomplete masculinization genitalia,with gonadal dysplasia and with/without the presence of Mullerian structures.At least 30 genes related to 46,XY DSD have ...46,XY disorders of sex development(DSD)is characterized by incomplete masculinization genitalia,with gonadal dysplasia and with/without the presence of Mullerian structures.At least 30 genes related to 46,XY DSD have been found.However,the clinical phenotypes of patients with different gene mutations overlap,and accurate diagnosis relies on gene sequencing technology.Therefore,this study aims to determine the prevalence of pathogenic mutations in a Chinese cohort with 46,XY DSD by the targeted nextgeneration sequencing(NGS)technology.Eighty-seven 46,XY DSD patients were enrolled from the Peking Union Medical College Hospital(Beijing,China).A total of fifty-four rare variants were identified in 60 patients with 46,XY DSD.The incidence of these rare variants was approximately 69.0%(60/87).Twenty-five novel variants and 29 reported variants were identified.Based on the American College of Medical Genetics and Genomics(ACMG)guidelines,thirty-three variants were classified as pathogenic or likely pathogenic variants and 21 variants were assessed as variants of uncertain significance.The overall diagnostic rate was about 42.5%based on the pathogenic and likely pathogenic variants.Androgen receptor{AR),steroid 5-alpha-reductase 2(SRD5A2)and nuclear receptor subfamily 5 Group A member 1(NR5A1)gene variants were identified in 21,13 and 13 patients,respectively.The incidence of these three gene variants was about 78.3%(47/60)in patients with rare variants.It is concluded that targeted NGS is an effective method to detect pathogenic mutations in 46,XY DSD patients and AR,SRD5A2,and NR5A1 genes were the most common pathogenic genes in our cohort.展开更多
Owing to the limitation of a large genome size(~13 Gb),the genetic and gene mapping studies on faba bean(Vicia faba L.)are lagging far behind those for other legumes.In this study,we selected three purified faba bean ...Owing to the limitation of a large genome size(~13 Gb),the genetic and gene mapping studies on faba bean(Vicia faba L.)are lagging far behind those for other legumes.In this study,we selected three purified faba bean lines(Yundou 8137,H0003712,and H000572)as parents and constructed two F2 populations.These two F2 populations,namely 167 F2 plants in Pop1(Yundou 8137×H0003712)and 204 F2 plants in Pop2(H000572×Yundou 8137),were genotyped using a targeted next-generation sequencing(TNGS)genotyping platform,and two high-density single nucleotide polymorphisms(SNP)genetic linkage maps of faba bean were constructed.The map constructed from Pop1 contained 5103 SNPs with a length of 1333.31 cM and an average marker density of 0.26 cM.The map constructed from Pop2 contained 1904 SNPs with a greater length of 1610.61 cM.In these two F2 populations,QTL mapping identified 98 QTLs for 14 agronomic traits related to the flowers,pods,plant types and grains.The two maps were then merged into an integrated genetic linkage map containing 6895 SNPs,with a length of 3324.48 cM.These results not only lay the foundation for fine mapping and map-based cloning of related genes,but can also accelerate the molecular marker-assisted breeding of faba bean.展开更多
Colorectal cancer(CRC)is a common malignant tumor worldwide,and its tumor microenvironment(TME)plays a crucial role in tumor progression.Neutrophil extracellular traps(NETs),as an important component of the TME,have r...Colorectal cancer(CRC)is a common malignant tumor worldwide,and its tumor microenvironment(TME)plays a crucial role in tumor progression.Neutrophil extracellular traps(NETs),as an important component of the TME,have received widespread attention in recent years.This article explores the biological functions and molecular mechanisms of NETs in CRC and their impact on disease progression,while analyzing the application of single-cell sequencing technology(SCS)in this field.The development of SCS provides a new perspective for understanding the role of NETs in CRC.By combining SCS technology,targeting key regulatory nodes of NETs is expected to reverse the immunosuppressive microenvironment and provide a theoretical basis for developing novel diagnostic biomarkers and targeted therapeutic strategies,thereby promoting the development of precision medicine in CRC and helping enhance patient prognosis.Future research should further explore the integration of SCS technology with complementary methodologies to investigate NETs and develop specific detection methods and therapeutic strategies targeting NETs to enhance early diagnosis and treatment efficacy of tumors.展开更多
Introduction: Omicron is a highly divergent variant of concern (VOCs) of a severe acute respiratory syndrome SARS-CoV-2. It carries a high number of mutations in its spike protein hence;it is more transmissible in the...Introduction: Omicron is a highly divergent variant of concern (VOCs) of a severe acute respiratory syndrome SARS-CoV-2. It carries a high number of mutations in its spike protein hence;it is more transmissible in the community by immune evasion mechanisms. Due to mutation within S gene, most Omicron variants have reported S gene target failure (SGTF) with some commercially available PCR kits. Such diagnostic features can be used as markers to screen Omicron. However, Whole Genome Sequencing (WGS) is the only gold standard approach to confirm novel microorganisms at genetically level as similar mutations can also be found in other variants that are circulating at low frequencies worldwide. This Retrospective study is aimed to assess RT-PCR sensitivity in the detection of S gene target failure in comparison with whole genome sequencing to detect variants of Omicron. Methods: We have analysed retrospective data of SARS-CoV-2 positive RT-PCR samples for S gene target failure (SGTF) with TaqPath COVID-19 RT-PCR Combo Kit (ThermoFisher) and combined with sequencing technologies to study the emerged pattern of SARS-CoV-2 variants during third wave at the tertiary care centre, Surat. Results: From the first day of December 2021 till the end of February 2022, a total of 321,803 diagnostic RT-PCR tests for SARS-CoV-2 were performed, of which 20,566 positive cases were reported at our tertiary care centre with an average cumulative positivity of 6.39% over a period of three months. In the month of December 21 samples characterized by the SGTF (70/129) were suggestive of being infected by the Omicron variant and identified as Omicron (B.1.1.529 lineage) when sequence. In the month of January, we analysed a subset of samples (n = 618) with SGTF (24%) and without SGTF (76%) with Ct values Conclusions: During the COVID-19 pandemic, it took almost more than 15 days to diagnose infection and identify pathogen by sequencing technology. In contrast to that molecular assay provided quick identification with the help of SGTF phenomenon within 5 hours of duration. This strategy helps scientists and health policymakers for the quick isolation and identification of clusters. That ultimately results in a decreased transmission of pathogen among the community.展开更多
This article reviews basic concepts, general applications, and the potential impact of next-generation sequencing （NGS） technologies on genomics, with particular reference to currently available and possible future ...This article reviews basic concepts, general applications, and the potential impact of next-generation sequencing （NGS） technologies on genomics, with particular reference to currently available and possible future platforms and bioinformatics. NGS technologies have demon- strated the capacity to sequence DNA at unprecedented speed, thereby enabling previously unimaginable scientific achievements and novel biological applications. But, the massive data produced by NGS also presents a significant challenge for data storage, analyses, and management solutions. Advanced bioinformatic tools are essential for the successful application of NGS technology. As evidenced throughout this review, NGS technologies will have a striking impact on genomic research and the entire biological field. With its ability to tackle the unsolved challenges unconquered by previous genomic technologies, NGS is likely to unravel the complexity of the human genome in terms of genetic variations, some of which may be confined to susceptible loci for some common human conditions. The impact of NGS technologies on genomics will be far reaching and likely change the field for years to come.展开更多
Gastric cancer is the second leading cause of cancerrelated deaths worldwide.Conventional cytotoxic chemotherapy has limited efficacy for metastatic gastric cancer,with an overall survival of approximately ten months....Gastric cancer is the second leading cause of cancerrelated deaths worldwide.Conventional cytotoxic chemotherapy has limited efficacy for metastatic gastric cancer,with an overall survival of approximately ten months.Recent advances in high-throughput technologies have enabled the implementation of personalized cancer therapy for high-risk patients.The use of such high-throughput technologies,including microarray and next generation sequencing,have promoted the discovery of novel targets that offer new treatment strategies for patients lacking other therapeutic options.Many molecular pathways are currently under investigation as therapeutic targets in gastric cancer,including those related to the epidermal growth factor receptor family,the mesenchymal-epithelial transition factor axis,and the phosphatidylinositol 3-kinase-AKTmammalian target of rapamycin factors.Advances in molecular diagnostic tools further support the discovery of new molecular targets.Limitations exist,however;not all patients can be tested for biomarkers,and numerous challenges hamper implementation of targeted therapy in clinical settings.Indeed,the scale of tumor genomic profiling is rapidly outpacing our ability to appropriately synthesize all the information in order to optimally refine patient care.Therefore,clinicians must continue to educate themselves regarding new tools and frameworks,and to utilize multidisciplinary team science,comprised of oncologists,geneticists,pathologists,biologists and bioinformaticians,to successfully implement this genomic approach therapeutically.展开更多
On the basis of scale invariant feature transform（SIFT） descriptors,a novel kind of local invariants based on SIFT sequence scale（SIFT-SS） is proposed and applied to target classification.First of all,the merits o...On the basis of scale invariant feature transform（SIFT） descriptors,a novel kind of local invariants based on SIFT sequence scale（SIFT-SS） is proposed and applied to target classification.First of all,the merits of using an SIFT algorithm for target classification are discussed.Secondly,the scales of SIFT descriptors are sorted by descending as SIFT-SS,which is sent to a support vector machine（SVM） with radial based function（RBF） kernel in order to train SVM classifier,which will be used for achieving target classification.Experimental results indicate that the SIFT-SS algorithm is efficient for target classification and can obtain a higher recognition rate than affine moment invariants（AMI） and multi-scale auto-convolution（MSA） in some complex situations,such as the situation with the existence of noises and occlusions.Moreover,the computational time of SIFT-SS is shorter than MSA and longer than AMI.展开更多
AIM：To make a comprehensive analysis of the potential pathogenic genes related with Leber congenital amaurosis（LCA） in Chinese.METHODS：LCA subjects and their families were retrospectively collected from 2013 to 20...AIM：To make a comprehensive analysis of the potential pathogenic genes related with Leber congenital amaurosis（LCA） in Chinese.METHODS：LCA subjects and their families were retrospectively collected from 2013 to 2015.Firstly,whole-exome sequencing was performed in patients who had underwent gene mutation screening with nothing found,and then homozygous sites was selected,candidate sites were annotated,and pathogenic analysis was conducted using softwares including Sorting Tolerant from Intolerant（SIFT）,Polyphen-2,Mutation assessor,Condel,and Functional Analysis through Hidden Markov Models（FATHMM）.Furthermore,Gene Ontology function and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses of pathogenic genes were performed followed by co-segregation analysis using Fisher exact Test.Sanger sequencing was used to validate single-nucleotide variations（SNVs）.Expanded verification was performed in the rest patients.RESULTS：Totally 51 LCA families with 53 patients and24 family members were recruited.A total of 104 SNVs（66 LCA-related genes and 15 co-segregated genes）were submitted for expand verification.The frequencies of homozygous mutation of KRT12 and CYP1A1 were simultaneously observed in 3 families.Enrichment analysis showed that the potential pathogenic genes were mainly enriched in functions related to cell adhesion,biological adhesion,retinoid metabolic process,and eye development biological adhesion.Additionally,WFS7 and STAU2 had the highest homozygous frequencies.CONCLUSION：LCA is a highly heterogeneous disease.Mutations in KRT12,CVP1A1,WFS1,and STAU2 may be involved in the development of LCA.展开更多
Iron-sulfur clusters(ISC)are essential cofactors for proteins involved in various biological processes,such as electron transport,biosynthetic reactions,DNA repair,and gene expression regulation.ISC assembly protein I...Iron-sulfur clusters(ISC)are essential cofactors for proteins involved in various biological processes,such as electron transport,biosynthetic reactions,DNA repair,and gene expression regulation.ISC assembly protein IscA1(or MagR)is found within the mitochondria of most eukaryotes.Magnetoreceptor(MagR)is a highly conserved A-type iron and iron-sulfur cluster-binding protein,characterized by two distinct types of iron-sulfur clusters,[2Fe-2S]and[3Fe-4S],each conferring unique magnetic properties.MagR forms a rod-like polymer structure in complex with photoreceptive cryptochrome(Cry)and serves as a putative magnetoreceptor for retrieving geomagnetic information in animal navigation.Although the N-terminal sequences of MagR vary among species,their specific function remains unknown.In the present study,we found that the N-terminal sequences of pigeon MagR,previously thought to serve as a mitochondrial targeting signal(MTS),were not cleaved following mitochondrial entry but instead modulated the efficiency with which iron-sulfur clusters and irons are bound.Moreover,the N-terminal region of MagR was required for the formation of a stable MagR/Cry complex.Thus,the N-terminal sequences in pigeon MagR fulfil more important functional roles than just mitochondrial targeting.These results further extend our understanding of the function of MagR and provide new insights into the origin of magnetoreception from an evolutionary perspective.展开更多
Objective:Plant-derived cytotoxic transgene expression,such as trichosanthin(tcs),regulated by recombinant adeno-associated virus(r AAV)vector is a promising cancer gene therapy.However,the cytotoxic transgene can ham...Objective:Plant-derived cytotoxic transgene expression,such as trichosanthin(tcs),regulated by recombinant adeno-associated virus(r AAV)vector is a promising cancer gene therapy.However,the cytotoxic transgene can hamper the vector production in the r AAV producer cell line,human embryonic kidney(HEK293)cells.Here,we explored micro RNA-122(miR122)and its target sequence to limit the expression of the cytotoxic gene in the r AAV producer cells.Methods:A miR122 target(122 T)sequence was incorporated into the 30 untranslated region of the tcs c DNA sequence.The firefly luciferase(fluc)transgene was used as an appropriate control.Cell line HEK293-mir122 was generated by the lentiviral vector-mediated genome integration of the mir122 gene in parental HEK293 cells.The effects of miR122 overexpression on cell growth,transgene expression,and r AAV production were determined.Results:The presence of 122 T sequence significantly reduced transgene expression in the miR122-enriched Huh7 cell line(in vitro),fresh human hepatocytes(ex vivo),and mouse liver(in vivo).Also,the normal liver physiology was unaffected by delivery of 122 T sequence by r AAV vectors.Compared with the parental cells,the miR122-overexpressing HEK293-mir122 cell line showed similar cell growth rate and expression of transgene without 122 T,as well as the ability to produce liver-targeting r AAV vectors.Fascinatingly,the yield of r AAV vectors carrying the tcs-122 T gene was increased by 77.7-fold in HEK293-mir122 cells.Moreover,the tcs-122 T-containing r AAV vectors significantly reduced the proliferation of hepatocellular carcinoma cells without affecting the normal liver cells.Conclusion:HEK293-mir122 cells along with the 122 T sequence provide a potential tool to attenuate the cytotoxic transgene expression,such as tcs,during r AAV vector production.展开更多
The advent of single-cell RNA sequencing(scRNA-seq)has provided insight into the tumour immune microenvironment(TIME).This review focuses on the application of scRNA-seq in investigation of the TIME.Over time,scRNA-se...The advent of single-cell RNA sequencing(scRNA-seq)has provided insight into the tumour immune microenvironment(TIME).This review focuses on the application of scRNA-seq in investigation of the TIME.Over time,scRNA-seq methods have evolved,and components of the TIME have been deciphered with high resolution.In this review,we first introduced the principle of scRNA-seq and compared different sequencing approaches.Novel cell types in the TIME,a continuous transitional state,and mutual intercommunication among TIME components present potential targets for prognosis prediction and treatment in cancer.Thus,we concluded novel cell clusters of cancerassociated fibroblasts(CAFs),T cells,tumour-associated macrophages(TAMs)and dendritic cells(DCs)discovered after the application of scRNA-seq in TIME.We also proposed the development of TAMs and exhausted T cells,as well as the possible targets to interrupt the process.In addition,the therapeutic interventions based on cellular interactions in TIME were also summarized.For decades,quantification of the TIME components has been adopted in clinical practice to predict patient survival and response to therapy and is expected to play an important role in the precise treatment of cancer.Summarizing the current findings,we believe that advances in technology and wide application of single-cell analysis can lead to the discovery of novel perspectives on cancer therapy,which can subsequently be implemented in the clinic.Finally,we propose some future directions in the field of TIME studies that can be aided by scRNA-seq technology.展开更多
Objective Thrombotic thrombocytopenic purpura(TTP)is a rare and fatal disease caused by a severe deficiency in the metalloprotease ADAMTS13 and is characterized by thrombotic microangiopathy.The present study aimed to...Objective Thrombotic thrombocytopenic purpura(TTP)is a rare and fatal disease caused by a severe deficiency in the metalloprotease ADAMTS13 and is characterized by thrombotic microangiopathy.The present study aimed to investigate the genes and variants associated with TTP in a Chinese population.Methods Target sequencing was performed on 220 genes related to complements,coagulation factors,platelets,fibrinolytic,endothelial,inflammatory,and anticoagulation systems in 207 TTP patients and 574 controls.Subsequently,logistic regression analysis was carried out to identify the TTP-associated genes based on the counts of rare deleterious variants in the region of a certain gene.Moreover,the associations between common variants and TTP were also investigated.Results ADAMTS13 was the only TTP-associated gene(OR=3.77;95%CI:1.82–7.81;P=3.6×10^(-4))containing rare deleterious variants in TTP patients.Among these 8 variants,5 novel rare variants that might contribute to TTP were identified,including rs200594025,rs782492477,c.T1928G(p.I643S),c.3336_3361del(p.Q1114Afs*20),and c.3469_3470del(p.A1158Sfs*17).No common variants associated with TTP were identified under the stringent criteria of correction for multiple testing.Conclusion ADAMTS13 is the primary gene related to TTP.The genetic variants associated with the occurrence of TTP were slightly different between the Chinese and European populations.展开更多
The high-tech listed firms are not only representative for mainstream of nongovernmental businesses, but also afflux new livingness into the capital market development in China. By considering our country＇s high-tech...The high-tech listed firms are not only representative for mainstream of nongovernmental businesses, but also afflux new livingness into the capital market development in China. By considering our country＇s high-tech listed firms as study object, we have disclosed different degree between diverse financial targets and business value, built a financial targets sequence model of impact on business value. The study supplies a certain extent reference to valid supervising and exalting of firms＇ financial core.展开更多
Pseudogenes are frequently encountered noncoding sequences with a high sequence similarity to their protein-coding paralogue.For this reason,their presence is often considered troublesome in molecular diagnostics.In p...Pseudogenes are frequently encountered noncoding sequences with a high sequence similarity to their protein-coding paralogue.For this reason,their presence is often considered troublesome in molecular diagnostics.In pseudoxanthoma elasticum(PXE),a disease predominantly caused by mutations in ATPbinding cassette family C member 6(ABCC6),the presence of two pseudogenes complicates the analysis of sequence data.With whole-exome sequencing(WES)becoming the standard of care in molecular diagnostics,we wanted to evaluate whether this technique is as reliable as gene-specific targeted enrichment analysis for the analysis of ABCC6.We established a PCR-based targeted enrichment and next-generation sequencing testing approach and demonstrated that the ABCC6-specific enrichment combined with the applied mapping algorithm overcomes the complication of ABCC6 pseudogene aspecificities,contrary to WES.We propose a time-and cost-efficient diagnostic strategy for comprehensive and accurate molecular genetic testing of PXE,which is highly automatable.展开更多
基金Supported by Open Project of Key Laboratory of Hubei Province(No.2023KFZZ026).
文摘AIM:To estimate if nanopore targeted sequencing(NTS)could identify pathogens causing postoperative endophthalmitis and further determine the feasibility of clinical application of NTS.METHODS:A total of 55 patients(55 eyes)with postoperative endophthalmitis were retrospectively included in this study with their medical records.Intraocular fluid samples were examined by NTS and microbial culture.All included patients had undergone examinations including measurement of best corrected visual acuity(BCVA)and intraocular pressure(IOP),slit-lamp biomicroscopy,and indirect ophthalmoscopy;additionally,they underwent B-ultrasound,anterior segment photography,and fundus photography if necessary.RESULTS:Among 55 patients with postoperative endophthalmitis,the age was 65.25±15.04y and there were 30 female(54.54%)patients.Forty-one(74.54%)vitreous humor samples and fourteen(25.45%)aqueous humor samples were sent for both NTS and microbial culture.NTS had a notable higher detection rate than microbial culture in detecting pathogens(90.91%vs 38.18%,χ^(2)=33.409,P<0.001).NTS exhibited high sensitivity of pathogen detection in both microbial culture positive and negative samples(100%and 85.29%,respectively).In 16 of 21(76.19%)patients who showed culture-positivity,their results corresponded with those of NTS.Moreover,in two patients(9.52%),NTS showed a better species resolution than microbial culture;in three patients(14.28%),NTS identified additional pathogens.As for fungus,the positive detection rate of NTS was significantly higher than that of microbial culture(20%vs 3.64%,χ^(2)=7.066,P=0.008).Also,NTS could detect multi-infection by bacteria and fungi than microbial culture(32.73%vs 0,χ^(2)=21.522,P<0.001).NTS could detect bacteria as well as fungi simultaneously within 48h in all patients.Meanwhile,NTS had a shorter detection time than microbial culture(1.13±0.34 vs 2.67±0.55d,Z=-9.218,P<0.001).After the NTS results were obtained,15 patients received additional intravitreal/intracameral anti-infection treatment.At follow-up,there was a statistically significant improvement in the visual acuity relative to the baseline(Z=−5.222,P<0.001).CONCLUSION:NTS can provide rapid identification and highly sensitive detection of pathogens among patients with postoperative endophthalmitis,which can guide anti-infection treatment and improve visual prognosis.
基金supported by the National Natural Science Foundation of China(Grant Nos.32172602,32472739)the Major Science and Technology Project of Henan Province(Grant No.221100110400)+3 种基金the Funding of Joint Research on Agricultural Varietie Improvement of Henan Province(Grant No.2022010503)the Natural Science Foundation of Henan(Grant No.242300421030)the Key Scientifc and Technological Project of Henan Province(Grant Nos.242102111124,242102111115)the Key Research and Development Program of Xinjiang Uygur autonomous region(2023B02017-2).
文摘Genotyping by Target Sequencing(GBTS)technology,known for its flexibility,high efficiency,high throughput,and low cost,has been increasingly employed in molecular breeding.However,there is still limited study on the design and development of high-throughput genotyping tools in watermelon.In this study,we identified 112000 high quality SNPs by analyzing the resequencing data of 43 cultivated watermelon accessions.11921 and 6094 SNPs were selected for developing two sets of watermelon liquid-phase chips with different marker densities,named Watermelon 10K and 5K,respectively.Furthermore,the SNPs and Indels of most mapped gene/QTLs for many agronomic important traits in watermelon were also integrated into the two chips for foreground selection.These chips have been tested using GBTS technology in various applications in watermelon.The genotyping of 76 accessions by Watermelon 5K liquid-phase chip showed an average detection rate of 99.28%and 81.78%for cultivated and wild watermelon accessions,respectively.This provided enough markers information for GWAS and two significant QTLs,ssc1.1 and ssc1.2,associated with soluble sugar content were detected.Furthermore,BSA-seq analysis for non-lobed leaf and dwarf traits were validated by liquid-phase chips,and the candidate region was consistent with our previous studies.Additionally,we precisely introduced the Cldw1 and Clbl genes into an elite inbred line WT2 using Watermelon 5K for assisted selection,resulting in the development of three new germplasm with good plant architecture.As a high-throughput genotyping liquid-phase SNP array,the Watermelon 10K and 5K chips will greatly facilitate functional studies and molecular breeding in watermelon.
基金partially supported by grants from the Ministry of Science and Technology of China(2016YFC1000306)the Beijing Municipal Science and Technology Commission Foundation(Z181100001918003)+1 种基金the Beijing Municipal Commission of Health and Family Planning Foundation(2018-21141,2020-4-1144)Beihang University&Capital Medical University Advanced Innovation Center for Big Data-Based Precision Medicine Plan(BHME-201905)。
文摘Different newborn screening(NBS) programs have been practiced in many countries since the 1960 s. It is of considerable interest whether next-generation sequencing is applicable in NBS. We have developed a panel of 465 causative genes for 596 early-onset, relatively high incidence, and potentially actionable severe inherited diseases in our Newborn Screening with Targeted Sequencing(NESTS) program to screen 11,484 babies in 8 Women and Children’s hospitals nationwide in China retrospectively. The positive rate from preliminary screening of NESTS was 7.85%(902/11,484). With 45.89%(414/902) follow-up of preliminary positive cases, the overall clinically confirmative diagnosis rate of monogenic disorders was 12.07%(50/414), estimating an average of 0.95%(7.85% × 12.07%) clinical diagnosis rate, suggesting that monogenic disorders account for a considerable proportion of birth defects. The disease/gene spectrum varied in different regions of China. NESTS was implemented in a hospital by screening 3923 newborns to evaluate its clinical application. The turn-around time of a primary report, including the sequencing period of < 7 days, was within 11 days by our automatic interpretation pipeline. Our results suggest that NESTS is feasible and cost-effective as a first-tier NBS program, which will change the status of current clinical practice of NBS in China.
基金supported by the grants from the Key R&D Program of Shandong Province,China(2022LZGC003)the China Agriculture Research System of MOF and MARA(CARS-35)+1 种基金the National Key Research and Development Project of China(2019YFE0106800)the 2115 Talent Development Program of China Agricultural University。
文摘Genotyping by target sequencing(GBTS)integrates the advantages of silicon-based technology(high stability and reliability)and genotyping by sequencing(high flexibility and cost-effectiveness).However,GBTS panels are not currently available in pigs.In this study,based on GBTS technology,we first developed a 50K panel,including 52,000 single-nucleotide polymorphisms(SNPs),in pigs,designated GBTS50K.A total of 6,032 individuals of Large White,Landrace,and Duroc pigs from 10 breeding farms were used to assess the newly developed GBTS50K.Our results showed that GBTS50K obtained a high genotyping ability,the SNP and individual call rates of GBTS50K were 0.997–0.998,and the average consistency rate and genotyping correlation coefficient were 0.997 and 0.993,respectively,in replicate samples.We also evaluated the efficiencies of GBTS50K in the application of population genetic structure analysis,selection signature detection,genome-wide association studies(GWAS),genotyped imputation,genetic selection(GS),etc.The results indicate that GBTS50K is plausible and powerful in genetic analysis and molecular breeding.For example,GBTS50K could gain higher accuracies than the current popular GGP-Porcine bead chip in genomic selection on 2 important traits of backfat thickness at 100 kg and days to 100 kg in pigs.Particularly,due to the multiple SNPs(mSNPs),GBTS50K generated 100K qualified SNPs without increasing genotyping cost,and our results showed that the haplotype-based method can further improve the accuracies of genomic selection on growth and reproduction traits by 2 to 6%.Our study showed that GBTS50K could be a powerful tool for underlying genetic architecture and molecular breeding in pigs,and it is also helpful for developing SNP panels for other farm animals.
文摘Objective This study aimed to explore the diagnostic value of novel technique-targeted next-generation sequencing(tNGS)of bronchoalveolar lavage fluid(BALF)in pulmonary mycobacterial infections.Methods This retrospective study was conducted on patients who underwent bronchoscopy and tNGS,smear microscopy,and mycobacterial culture of BALF.Patients with positive Mycobacterium tuberculosis(MTB)culture or GeneXpert results were classified into the tuberculosis case group.Those diagnosed with nontuberculous mycobacteria(NTM)-pulmonary disease(NTM-PD)composed the case group of NTM-PD patients.The control group comprised patients without tuberculosis or NTM-PD.Sensitivity,specificity,and receiver operating characteristic(ROC)curves were used to evaluate the diagnostic performance.Results For tuberculosis patients with positive mycobacterial culture results,the areas under the ROC curves(AUCs)for tNGS,GeneXpert,and smear microscopy were 0.975(95%CI:0.935,1.000),0.925(95%CI:0.859,0.991),and 0.675(95%CI:0.563,0.787),respectively.For tuberculosis patients with positive GeneXpert results,the AUCs of tNGS,culture,and smear microscopy were 0.970(95%CI:0.931,1.000),0.850(95%CI:0.770,0.930),and 0.680(95%CI:0.579,0.781),respectively.For NTM-PD,the AUCs of tNGS,culture,and smear-positive but GeneXpert-negative results were 0.987(95%CI:0.967,1.000),0.750(95%CI:0.622,0.878),and 0.615(95%CI:0.479,0.752),respectively.The sensitivity and specificity of tNGS in NTM-PD patients were 100%and 97.5%,respectively.Conclusion tNGS demonstrated superior diagnostic efficacy in mycobacterial infection,indicating its potential for clinical application.
基金This work was supported by the National Natural Science Foundation of China(No.81971375 and No.81771576)the National Key Research and Development Program of China(2016YFC0905100)+1 种基金the CAMS Innovation Fund for Medical Sciences(2016-I2M-1-002)the Nonprofit Central Research Institute Fund of the Chinese Academy of Medical Sciences(No.2017PT32020 and No.2018PT32001).
文摘46,XY disorders of sex development(DSD)is characterized by incomplete masculinization genitalia,with gonadal dysplasia and with/without the presence of Mullerian structures.At least 30 genes related to 46,XY DSD have been found.However,the clinical phenotypes of patients with different gene mutations overlap,and accurate diagnosis relies on gene sequencing technology.Therefore,this study aims to determine the prevalence of pathogenic mutations in a Chinese cohort with 46,XY DSD by the targeted nextgeneration sequencing(NGS)technology.Eighty-seven 46,XY DSD patients were enrolled from the Peking Union Medical College Hospital(Beijing,China).A total of fifty-four rare variants were identified in 60 patients with 46,XY DSD.The incidence of these rare variants was approximately 69.0%(60/87).Twenty-five novel variants and 29 reported variants were identified.Based on the American College of Medical Genetics and Genomics(ACMG)guidelines,thirty-three variants were classified as pathogenic or likely pathogenic variants and 21 variants were assessed as variants of uncertain significance.The overall diagnostic rate was about 42.5%based on the pathogenic and likely pathogenic variants.Androgen receptor{AR),steroid 5-alpha-reductase 2(SRD5A2)and nuclear receptor subfamily 5 Group A member 1(NR5A1)gene variants were identified in 21,13 and 13 patients,respectively.The incidence of these three gene variants was about 78.3%(47/60)in patients with rare variants.It is concluded that targeted NGS is an effective method to detect pathogenic mutations in 46,XY DSD patients and AR,SRD5A2,and NR5A1 genes were the most common pathogenic genes in our cohort.
基金supported by the National Key R&D Program of China(2019YFD1001300 and 2019YFD1001303)the Construction of Molecular Database of Faba Bean and Pea and Identification of Maize Germplasm Project,Ministry of Agriculture and Rural Affairs,China(19200030)+3 种基金the Yunnan Key R&D Program,China(202202AE090003)the earmarked fund for China Agriculture Research System(CARS-08)the Crop Germplasm Resources Protection(2130135)the Major Agricultural Science and Technology Program of Chinese Academy of Agricultural Sciences(CAAS-XTCX20190025)。
文摘Owing to the limitation of a large genome size(~13 Gb),the genetic and gene mapping studies on faba bean(Vicia faba L.)are lagging far behind those for other legumes.In this study,we selected three purified faba bean lines(Yundou 8137,H0003712,and H000572)as parents and constructed two F2 populations.These two F2 populations,namely 167 F2 plants in Pop1(Yundou 8137×H0003712)and 204 F2 plants in Pop2(H000572×Yundou 8137),were genotyped using a targeted next-generation sequencing(TNGS)genotyping platform,and two high-density single nucleotide polymorphisms(SNP)genetic linkage maps of faba bean were constructed.The map constructed from Pop1 contained 5103 SNPs with a length of 1333.31 cM and an average marker density of 0.26 cM.The map constructed from Pop2 contained 1904 SNPs with a greater length of 1610.61 cM.In these two F2 populations,QTL mapping identified 98 QTLs for 14 agronomic traits related to the flowers,pods,plant types and grains.The two maps were then merged into an integrated genetic linkage map containing 6895 SNPs,with a length of 3324.48 cM.These results not only lay the foundation for fine mapping and map-based cloning of related genes,but can also accelerate the molecular marker-assisted breeding of faba bean.
基金the Shandong Province Medical and Health Science and Technology Development Plan Project,No.202203030713Yantai Science and Technology Program,No.2024YD005,No.2024YD007 and No.2024YD010and Science and Technology Program of Yantai Affiliated Hospital of Binzhou Medical University,No.YTFY2022KYQD06.
文摘Colorectal cancer(CRC)is a common malignant tumor worldwide,and its tumor microenvironment(TME)plays a crucial role in tumor progression.Neutrophil extracellular traps(NETs),as an important component of the TME,have received widespread attention in recent years.This article explores the biological functions and molecular mechanisms of NETs in CRC and their impact on disease progression,while analyzing the application of single-cell sequencing technology(SCS)in this field.The development of SCS provides a new perspective for understanding the role of NETs in CRC.By combining SCS technology,targeting key regulatory nodes of NETs is expected to reverse the immunosuppressive microenvironment and provide a theoretical basis for developing novel diagnostic biomarkers and targeted therapeutic strategies,thereby promoting the development of precision medicine in CRC and helping enhance patient prognosis.Future research should further explore the integration of SCS technology with complementary methodologies to investigate NETs and develop specific detection methods and therapeutic strategies targeting NETs to enhance early diagnosis and treatment efficacy of tumors.
文摘Introduction: Omicron is a highly divergent variant of concern (VOCs) of a severe acute respiratory syndrome SARS-CoV-2. It carries a high number of mutations in its spike protein hence;it is more transmissible in the community by immune evasion mechanisms. Due to mutation within S gene, most Omicron variants have reported S gene target failure (SGTF) with some commercially available PCR kits. Such diagnostic features can be used as markers to screen Omicron. However, Whole Genome Sequencing (WGS) is the only gold standard approach to confirm novel microorganisms at genetically level as similar mutations can also be found in other variants that are circulating at low frequencies worldwide. This Retrospective study is aimed to assess RT-PCR sensitivity in the detection of S gene target failure in comparison with whole genome sequencing to detect variants of Omicron. Methods: We have analysed retrospective data of SARS-CoV-2 positive RT-PCR samples for S gene target failure (SGTF) with TaqPath COVID-19 RT-PCR Combo Kit (ThermoFisher) and combined with sequencing technologies to study the emerged pattern of SARS-CoV-2 variants during third wave at the tertiary care centre, Surat. Results: From the first day of December 2021 till the end of February 2022, a total of 321,803 diagnostic RT-PCR tests for SARS-CoV-2 were performed, of which 20,566 positive cases were reported at our tertiary care centre with an average cumulative positivity of 6.39% over a period of three months. In the month of December 21 samples characterized by the SGTF (70/129) were suggestive of being infected by the Omicron variant and identified as Omicron (B.1.1.529 lineage) when sequence. In the month of January, we analysed a subset of samples (n = 618) with SGTF (24%) and without SGTF (76%) with Ct values Conclusions: During the COVID-19 pandemic, it took almost more than 15 days to diagnose infection and identify pathogen by sequencing technology. In contrast to that molecular assay provided quick identification with the help of SGTF phenomenon within 5 hours of duration. This strategy helps scientists and health policymakers for the quick isolation and identification of clusters. That ultimately results in a decreased transmission of pathogen among the community.
基金supported by NINDS/NIH(JZ),Coldwell Foundation(JZ) and TTUHSC(JZ)
文摘This article reviews basic concepts, general applications, and the potential impact of next-generation sequencing （NGS） technologies on genomics, with particular reference to currently available and possible future platforms and bioinformatics. NGS technologies have demon- strated the capacity to sequence DNA at unprecedented speed, thereby enabling previously unimaginable scientific achievements and novel biological applications. But, the massive data produced by NGS also presents a significant challenge for data storage, analyses, and management solutions. Advanced bioinformatic tools are essential for the successful application of NGS technology. As evidenced throughout this review, NGS technologies will have a striking impact on genomic research and the entire biological field. With its ability to tackle the unsolved challenges unconquered by previous genomic technologies, NGS is likely to unravel the complexity of the human genome in terms of genetic variations, some of which may be confined to susceptible loci for some common human conditions. The impact of NGS technologies on genomics will be far reaching and likely change the field for years to come.
文摘Gastric cancer is the second leading cause of cancerrelated deaths worldwide.Conventional cytotoxic chemotherapy has limited efficacy for metastatic gastric cancer,with an overall survival of approximately ten months.Recent advances in high-throughput technologies have enabled the implementation of personalized cancer therapy for high-risk patients.The use of such high-throughput technologies,including microarray and next generation sequencing,have promoted the discovery of novel targets that offer new treatment strategies for patients lacking other therapeutic options.Many molecular pathways are currently under investigation as therapeutic targets in gastric cancer,including those related to the epidermal growth factor receptor family,the mesenchymal-epithelial transition factor axis,and the phosphatidylinositol 3-kinase-AKTmammalian target of rapamycin factors.Advances in molecular diagnostic tools further support the discovery of new molecular targets.Limitations exist,however;not all patients can be tested for biomarkers,and numerous challenges hamper implementation of targeted therapy in clinical settings.Indeed,the scale of tumor genomic profiling is rapidly outpacing our ability to appropriately synthesize all the information in order to optimally refine patient care.Therefore,clinicians must continue to educate themselves regarding new tools and frameworks,and to utilize multidisciplinary team science,comprised of oncologists,geneticists,pathologists,biologists and bioinformaticians,to successfully implement this genomic approach therapeutically.
基金supported by the National High Technology Research and Development Program (863 Program) (2010AA7080302)
文摘On the basis of scale invariant feature transform（SIFT） descriptors,a novel kind of local invariants based on SIFT sequence scale（SIFT-SS） is proposed and applied to target classification.First of all,the merits of using an SIFT algorithm for target classification are discussed.Secondly,the scales of SIFT descriptors are sorted by descending as SIFT-SS,which is sent to a support vector machine（SVM） with radial based function（RBF） kernel in order to train SVM classifier,which will be used for achieving target classification.Experimental results indicate that the SIFT-SS algorithm is efficient for target classification and can obtain a higher recognition rate than affine moment invariants（AMI） and multi-scale auto-convolution（MSA） in some complex situations,such as the situation with the existence of noises and occlusions.Moreover,the computational time of SIFT-SS is shorter than MSA and longer than AMI.
基金Supported by National Natural Science Foundation of China(No.81470642No.81271045)
文摘AIM：To make a comprehensive analysis of the potential pathogenic genes related with Leber congenital amaurosis（LCA） in Chinese.METHODS：LCA subjects and their families were retrospectively collected from 2013 to 2015.Firstly,whole-exome sequencing was performed in patients who had underwent gene mutation screening with nothing found,and then homozygous sites was selected,candidate sites were annotated,and pathogenic analysis was conducted using softwares including Sorting Tolerant from Intolerant（SIFT）,Polyphen-2,Mutation assessor,Condel,and Functional Analysis through Hidden Markov Models（FATHMM）.Furthermore,Gene Ontology function and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses of pathogenic genes were performed followed by co-segregation analysis using Fisher exact Test.Sanger sequencing was used to validate single-nucleotide variations（SNVs）.Expanded verification was performed in the rest patients.RESULTS：Totally 51 LCA families with 53 patients and24 family members were recruited.A total of 104 SNVs（66 LCA-related genes and 15 co-segregated genes）were submitted for expand verification.The frequencies of homozygous mutation of KRT12 and CYP1A1 were simultaneously observed in 3 families.Enrichment analysis showed that the potential pathogenic genes were mainly enriched in functions related to cell adhesion,biological adhesion,retinoid metabolic process,and eye development biological adhesion.Additionally,WFS7 and STAU2 had the highest homozygous frequencies.CONCLUSION：LCA is a highly heterogeneous disease.Mutations in KRT12,CVP1A1,WFS1,and STAU2 may be involved in the development of LCA.
基金supported by the National Natural Science Foundation of China(31640001 and T2350005 to C.X.,U21A20148 to X.Z.and C.X.)Ministry of Science and Technology of China(2021ZD0140300 to C.X.)+2 种基金Natural Science Foundation of Hainan Province(No.822RC703 for J.L.)Foundation of Hainan Educational Committee(No.Hnky2022-27 for J.L.)Presidential Foundation of Hefei Institutes of Physical Science,Chinese Academy of Sciences(Y96XC11131,E26CCG27,and E26CCD15 to C.X.,E36CWGBR24B and E36CZG14132 to T.C.)。
文摘Iron-sulfur clusters(ISC)are essential cofactors for proteins involved in various biological processes,such as electron transport,biosynthetic reactions,DNA repair,and gene expression regulation.ISC assembly protein IscA1(or MagR)is found within the mitochondria of most eukaryotes.Magnetoreceptor(MagR)is a highly conserved A-type iron and iron-sulfur cluster-binding protein,characterized by two distinct types of iron-sulfur clusters,[2Fe-2S]and[3Fe-4S],each conferring unique magnetic properties.MagR forms a rod-like polymer structure in complex with photoreceptive cryptochrome(Cry)and serves as a putative magnetoreceptor for retrieving geomagnetic information in animal navigation.Although the N-terminal sequences of MagR vary among species,their specific function remains unknown.In the present study,we found that the N-terminal sequences of pigeon MagR,previously thought to serve as a mitochondrial targeting signal(MTS),were not cleaved following mitochondrial entry but instead modulated the efficiency with which iron-sulfur clusters and irons are bound.Moreover,the N-terminal region of MagR was required for the formation of a stable MagR/Cry complex.Thus,the N-terminal sequences in pigeon MagR fulfil more important functional roles than just mitochondrial targeting.These results further extend our understanding of the function of MagR and provide new insights into the origin of magnetoreception from an evolutionary perspective.
基金supported by the China Postdoctoral Science Foundation(No.2019M651381)the Shanghai Talent Development Funding(No.2019115)to Chen Zhong。
文摘Objective:Plant-derived cytotoxic transgene expression,such as trichosanthin(tcs),regulated by recombinant adeno-associated virus(r AAV)vector is a promising cancer gene therapy.However,the cytotoxic transgene can hamper the vector production in the r AAV producer cell line,human embryonic kidney(HEK293)cells.Here,we explored micro RNA-122(miR122)and its target sequence to limit the expression of the cytotoxic gene in the r AAV producer cells.Methods:A miR122 target(122 T)sequence was incorporated into the 30 untranslated region of the tcs c DNA sequence.The firefly luciferase(fluc)transgene was used as an appropriate control.Cell line HEK293-mir122 was generated by the lentiviral vector-mediated genome integration of the mir122 gene in parental HEK293 cells.The effects of miR122 overexpression on cell growth,transgene expression,and r AAV production were determined.Results:The presence of 122 T sequence significantly reduced transgene expression in the miR122-enriched Huh7 cell line(in vitro),fresh human hepatocytes(ex vivo),and mouse liver(in vivo).Also,the normal liver physiology was unaffected by delivery of 122 T sequence by r AAV vectors.Compared with the parental cells,the miR122-overexpressing HEK293-mir122 cell line showed similar cell growth rate and expression of transgene without 122 T,as well as the ability to produce liver-targeting r AAV vectors.Fascinatingly,the yield of r AAV vectors carrying the tcs-122 T gene was increased by 77.7-fold in HEK293-mir122 cells.Moreover,the tcs-122 T-containing r AAV vectors significantly reduced the proliferation of hepatocellular carcinoma cells without affecting the normal liver cells.Conclusion:HEK293-mir122 cells along with the 122 T sequence provide a potential tool to attenuate the cytotoxic transgene expression,such as tcs,during r AAV vector production.
基金supported by the National Key Research Development Program of China(2021YFA1301203)the National Natural Science Foundation of China(82103031,82103918,81973408)+6 种基金the Clinical Research Incubation Project,West China Hospital,Sichuan University(22HXFH019)the China Postdoctoral Science Foundation(2019 M653416)the International Cooperation Project of Chengdu Municipal Science and Technology Bureau(2020-GH02-00017-HZ)the“1.3.5 Project for Disciplines of Excellence,West China Hospital,Sichuan University”(ZYJC18035,ZYJC18025,ZYYC20003,ZYJC18003)the GIST Research Institute(GRI)IIBR grants funded by the GISTthe National Research Foundation of Korea funded by the Korean government(MSIP)(2019R1C1C1005403,2019R1A4A1028802 and2021M3H9A2097520)the Post-Doctor Research Project,West China Hospital,Sichuan University(2021HXBH054)。
文摘The advent of single-cell RNA sequencing(scRNA-seq)has provided insight into the tumour immune microenvironment(TIME).This review focuses on the application of scRNA-seq in investigation of the TIME.Over time,scRNA-seq methods have evolved,and components of the TIME have been deciphered with high resolution.In this review,we first introduced the principle of scRNA-seq and compared different sequencing approaches.Novel cell types in the TIME,a continuous transitional state,and mutual intercommunication among TIME components present potential targets for prognosis prediction and treatment in cancer.Thus,we concluded novel cell clusters of cancerassociated fibroblasts(CAFs),T cells,tumour-associated macrophages(TAMs)and dendritic cells(DCs)discovered after the application of scRNA-seq in TIME.We also proposed the development of TAMs and exhausted T cells,as well as the possible targets to interrupt the process.In addition,the therapeutic interventions based on cellular interactions in TIME were also summarized.For decades,quantification of the TIME components has been adopted in clinical practice to predict patient survival and response to therapy and is expected to play an important role in the precise treatment of cancer.Summarizing the current findings,we believe that advances in technology and wide application of single-cell analysis can lead to the discovery of novel perspectives on cancer therapy,which can subsequently be implemented in the clinic.Finally,we propose some future directions in the field of TIME studies that can be aided by scRNA-seq technology.
基金supported by the National Natural Science Foundation of China(No.82003561).
文摘Objective Thrombotic thrombocytopenic purpura(TTP)is a rare and fatal disease caused by a severe deficiency in the metalloprotease ADAMTS13 and is characterized by thrombotic microangiopathy.The present study aimed to investigate the genes and variants associated with TTP in a Chinese population.Methods Target sequencing was performed on 220 genes related to complements,coagulation factors,platelets,fibrinolytic,endothelial,inflammatory,and anticoagulation systems in 207 TTP patients and 574 controls.Subsequently,logistic regression analysis was carried out to identify the TTP-associated genes based on the counts of rare deleterious variants in the region of a certain gene.Moreover,the associations between common variants and TTP were also investigated.Results ADAMTS13 was the only TTP-associated gene(OR=3.77;95%CI:1.82–7.81;P=3.6×10^(-4))containing rare deleterious variants in TTP patients.Among these 8 variants,5 novel rare variants that might contribute to TTP were identified,including rs200594025,rs782492477,c.T1928G(p.I643S),c.3336_3361del(p.Q1114Afs*20),and c.3469_3470del(p.A1158Sfs*17).No common variants associated with TTP were identified under the stringent criteria of correction for multiple testing.Conclusion ADAMTS13 is the primary gene related to TTP.The genetic variants associated with the occurrence of TTP were slightly different between the Chinese and European populations.
文摘The high-tech listed firms are not only representative for mainstream of nongovernmental businesses, but also afflux new livingness into the capital market development in China. By considering our country＇s high-tech listed firms as study object, we have disclosed different degree between diverse financial targets and business value, built a financial targets sequence model of impact on business value. The study supplies a certain extent reference to valid supervising and exalting of firms＇ financial core.
基金supported by a Methusalem grant(BOF08/01M01108)from Ghent Universityfunded by Ghent University+1 种基金FWOthe Flemish Government-department EWI。
文摘Pseudogenes are frequently encountered noncoding sequences with a high sequence similarity to their protein-coding paralogue.For this reason,their presence is often considered troublesome in molecular diagnostics.In pseudoxanthoma elasticum(PXE),a disease predominantly caused by mutations in ATPbinding cassette family C member 6(ABCC6),the presence of two pseudogenes complicates the analysis of sequence data.With whole-exome sequencing(WES)becoming the standard of care in molecular diagnostics,we wanted to evaluate whether this technique is as reliable as gene-specific targeted enrichment analysis for the analysis of ABCC6.We established a PCR-based targeted enrichment and next-generation sequencing testing approach and demonstrated that the ABCC6-specific enrichment combined with the applied mapping algorithm overcomes the complication of ABCC6 pseudogene aspecificities,contrary to WES.We propose a time-and cost-efficient diagnostic strategy for comprehensive and accurate molecular genetic testing of PXE,which is highly automatable.
