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
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
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
BACKGROUND Hereditary spherocytosis(HS)is characterized by anemia,jaundice,splenomegaly,and cholelithiasis,and is caused by abnormal genes encoding red blood cell membrane components.The most common mutations found in...BACKGROUND Hereditary spherocytosis(HS)is characterized by anemia,jaundice,splenomegaly,and cholelithiasis,and is caused by abnormal genes encoding red blood cell membrane components.The most common mutations found in HS are in the ANK1 gene.CASE SUMMARY A 4-mo-old girl was admitted to our hospital with pallor that had lasted for more than 2 mo.She presented with jaundice,anemia and splenomegaly.A heterozygous mutation of ANK1(exon23:c.G2467T:p.E823X)was identified,and the mutation was determined to be autosomal dominant.This mutation is linked to the relatively serious anemia she had after birth;this anemia improved with age.CONCLUSION The utilization of next-generation sequencing may assist with the accurate diagnosis of HS,especially in atypical cases.展开更多
The expanding application of targeted next-generation sequencing(tNGS)in clinical diagnostics has led to an increasing frequency of Nocardia spp.detection in respiratory specimens;however,the clinical relevance and in...The expanding application of targeted next-generation sequencing(tNGS)in clinical diagnostics has led to an increasing frequency of Nocardia spp.detection in respiratory specimens;however,the clinical relevance and interpretation of these findings remain uncertain.This study describes the epidemiology of Nocardia pneumonia and colonization in patients at The University of Hong Kong–Shenzhen Hospital,China,and evaluates the diagnostic role of tNGS.The analysis was conducted from January 2023 through December 2024 and included 22 patients in whom Nocardia spp.were detected in respiratory specimens via tNGS.Among these patients,81.8%(18/22)had comorbidities,most commonly chronic lung disease(40.9%,9/22).Common clinical symptoms included cough,sputum and fever.Nocardia farcinica and Nocardia abscessus were the predominant species identified.Although the median number of se-quence reads of Nocardia were higher in infected patients than in colonized individuals(289,IQR:132.5–3747 vs.133.5,IQR:51.75–261.3),the difference was not statistically significant(P>0.05).In a comparison of diagnostic performance,tNGS identified Nocardia sequences in all 25 respiratory samples collected from the enrolled patients,whereas conventional aerobic culture success-fully isolated the pathogen in only two patients.Additionally,tNGS exhibited a significantly shorter median turnaround time than culture(38 h vs.376.5 h).These findings highlight tNGS as a rapid tool for Nocardia detection.However,the final diagnosis of nocardiosis can-not rely solely on sequence read thresholds.Clinical,radiological and laboratory integration remains critical in distinguishing true infection from colonization,ensuring accurate diagnosis and management of nocardiosis.展开更多
Introduction:Vaccination stands as the most effective preventive measure against yellow fever(YF).However,the YF vaccination associated Adverse Events following Immunization(AEFI)cases occur occasionally.Methods:The G...Introduction:Vaccination stands as the most effective preventive measure against yellow fever(YF).However,the YF vaccination associated Adverse Events following Immunization(AEFI)cases occur occasionally.Methods:The Guangdong Provincial Center for Disease Control and Prevention utilized Targeted Next-Generation Sequencing(tNGS)to determine whether the imported suspected YF case was infected by the wild-type YF virus strain or experienced an AEFI.Results:tNGS analysis successfully yielded a 10.2 kb viral genomic sequence.Subsequent in-depth analysis revealed high similarity to the YF vaccine strain 17D-213 and classified the sequence within the West Africa II genotype,clustering with the 17D vaccine strain.Conclusions and Implications for Public Health Practice:This case represented YF AEFI.The implementation of tNGS technology enables more precise and expeditious pathogen sequencing,providing critical evidence for accurate disease diagnosis and informed public health interventions.展开更多
Objective To screen gene mutation in adult-onset hypoparathyroidism in Chinese through the targeted nextgeneration sequencing(NGS).Methods We recruited 17patients with adult-onset hypoparathyroidism who were regularly...Objective To screen gene mutation in adult-onset hypoparathyroidism in Chinese through the targeted nextgeneration sequencing(NGS).Methods We recruited 17patients with adult-onset hypoparathyroidism who were regularly followed or newly diagnosed at our centre during the past one year.Nine of them developed hypercalciuria during the treatment with calcium and vitamin D.展开更多
Next-generation sequencing （NGS） is getting routinely used in the diagnosis of hereditary diseases, such as human cardiomyopathies. Hence. it is of utter importance to secure high quality sequencing data, enabling t...Next-generation sequencing （NGS） is getting routinely used in the diagnosis of hereditary diseases, such as human cardiomyopathies. Hence. it is of utter importance to secure high quality sequencing data, enabling the identification of disease-relevant mutations or the conclusion of negative test results. During the process of sample preparation, each protocol for target enrichment library preparation has its own requirements for quality control （QC）; however, there is little evi- dence on the actual impact of these guidelines on resulting data quality. In this study, we analyzed the impact of QC during the diverse library preparation steps of Agilent SureSelect XT target enrichment and lllumina sequencing. We quantified the parameters for a cohort of around 600 samples, which include starting amount of DNA, amount of sheared DNA, smallest and largest fragment size of the starting DNA; amount of DNA after the pre-PCR, and smallest and largest fragment size of the resulting DNA; as well as the amount of the final library, the corresponding smallest and largest fragment size, and the number of detected variants. Intriguingly, there is a high tolerance for variations in all QC steps, meaning that within the boundaries proposed in the current study, a considerable variance at each step of QC can be well tolerated without compromising NGS quality.展开更多
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
In rice fields,rice plants usually grow alongside wild weeds and are attacked by various invertebrate species.Viruses are abundant in plants and invertebrates,playing crucial ecological roles in controlling microbial ...In rice fields,rice plants usually grow alongside wild weeds and are attacked by various invertebrate species.Viruses are abundant in plants and invertebrates,playing crucial ecological roles in controlling microbial abundance and maintaining community structures.To date,only 16 rice viruses have been documented in rice-growing regions.These viruses pose serious threats to rice production and have traditionally been identified only from rice plants and insect vectors by isolation techniques.Advances in next-generation sequencing(NGS)have made it feasible to discover viruses on a global scale.Recently,numerous viruses have been identified in plants and invertebrates using NGS technologies.In this review,we discuss viral studies in rice plants,invertebrate species,and weeds in rice fields.Many novel viruses have been discovered in rice ecosystems through NGS technologies,with some also detected using metatranscriptomic and small RNA sequencing.These analyses greatly expand our understanding of viruses in rice fields and provide valuable insights for developing efficient strategies to manage insect pests and virus-mediated rice diseases.展开更多
This study investigates the diversity of gut microbiota in Metaphire peguana,an earthworm species commonly found in agricultural areas of Thailand.Earthworms play a critical role in soil ecosystems by supporting nutri...This study investigates the diversity of gut microbiota in Metaphire peguana,an earthworm species commonly found in agricultural areas of Thailand.Earthworms play a critical role in soil ecosystems by supporting nutrient cycling and breaking down organic matter.Understanding the microbial diversity in their gut is essential for exploring their ecological contributions.Using Next Generation Sequencing(NGS),we analyzed the mycobiome in the gut of M.peguana.Our findings revealed a high diversity of fungal species,primarily belonging to two major phyla:Ascomycota and Basidiomycota.Ascomycota was the most abundant phylum,comprising 40.1% of the total fungal species identified.A total of 33 distinct fungal species were identified,which underscores the richness of microbial life within the earthworm gut.This study successfully created the first genetic database of the microbial community in M.peguana,providing a foundation for future research in agricultural applications.The microbial species identified,particularly siderophoreproducing fungi,could have significant implications for improving soil fertility and promoting sustainable agricultural practices.The use of NGS technology has enabled comprehensive profiling of microbial communities,allowing for precise identification of fungi that may play essential roles in soil health.Furthermore,the study paves the way for future studies on the potential applications of earthworm gut microbiomes in biotechnology,especially in enhancing soil nutrient availability and plant growth.The findings of this research contribute to the broader understanding of the ecological roles of earthworms and their microbiomes in soil ecosystems.展开更多
Objective and Background Early and accurate diagnosis of spinal infections,including spinal tuberculosis,is pivotal for effective treatment but remains challenging.This study aims to assess the diagnostic yield of met...Objective and Background Early and accurate diagnosis of spinal infections,including spinal tuberculosis,is pivotal for effective treatment but remains challenging.This study aims to assess the diagnostic yield of metagenomic next-generation sequencing(mNGS)compared with that of conventional microbiological tests(CMTs)in identifying pathogens associated with spinal pathologies,with a special focus on infections leading to surgical interventions.Methods We enrolled 85 patients who underwent spinal surgery,comprising 63 patients with clinically diagnosed spinal infections,including patients with spinal tuberculosis,and 22 patients with noninfectious spinal conditions.The procedures involved irrigation and debridement for persistent wound drainage,with subsequent DNA extraction from plasma and joint fluid for mNGS and CMT analysis.Results Significantly increased C-reactive protein(CRP)levels were observed in patients with infections.The mNGS approach showed greater diagnostic sensitivity(92.06%)for detecting pathogens,including Mycobacterium tuberculosis,than did CMTs(36.51%).Despite its low specificity,mNGS had considerable negative predictive value(70.59%),underscoring its utility in ruling out infections.Conclusions The mNGS offers superior sensitivity over CMTs in the diagnosis of a variety of spinal infections,notably spinal tuberculosis.This study highlights the potential of mNGS in enhancing the diagnosis of complex spinal infections,thereby informing targeted treatment strategies.展开更多
BACKGROUND Leuconostoc garlicum is commonly found in fermented foods and very few infected patients have been reported,who typically present symptoms such as fever and fatigue.Conventional clinical examinations often ...BACKGROUND Leuconostoc garlicum is commonly found in fermented foods and very few infected patients have been reported,who typically present symptoms such as fever and fatigue.Conventional clinical examinations often struggle to identify this bacterium,and routine anti-infective treatments are generally ineffective.Both diagnostic challenges and therapeutic limitations pose significant difficulties for clinicians.CASE SUMMARY We report a patient ultimately diagnosed with Leuconostoc garlicum infection.The primary manifestations included persistent fever,cough and fatigue.These symptoms lasted for 2 months.He received anti-infective treatment at a community hospital,but this was ineffective.After inquiring about the patient's medical history and conducting a physical examination,the patient underwent laboratory tests.Complete blood count tests revealed that the patient had a high proportion of neutrophils,C-reactive protein level was 235.9 mg/L,erythrocyte sedimentation rate was 67 mm/h,respiratory pathogen testing was negative,and he was then thought to have an infectious disease.However,conventional anti-infective treatments were ineffective.After excluding infectious neurological diseases,urologic diseases and digestive problems,we ultimately focused our attention on the lungs.A lung computed tomography scan indicated pulmonary inflammation.Bronchoalveolar lavage fluid for next-generation sequencing suggested lung infection with Leuconostoc garlicum.The patient's symptoms gradually improved following treatment with piperacillin tazobactam and linezolid.During the follow-up period,the patient's temperature remained normal.CONCLUSION For patients with suspected bacterial infection and experiencing fever,conventional anti-infective treatment can be ineffective in controlling their symptoms,and an infection due to rare bacteria or drug-resistant bacteria should be considered.Next-generation sequencing enables rapid and precise identification of infection-related pathogens in febrile patients.展开更多
In this study,an amine-reactive poly(pentafluorophenyl acrylate)(PPFPA)platform was developed for advanced surface engineering of next-generation sequencing(NGS)chips.Through post-polymerization modification,PPFPA was...In this study,an amine-reactive poly(pentafluorophenyl acrylate)(PPFPA)platform was developed for advanced surface engineering of next-generation sequencing(NGS)chips.Through post-polymerization modification,PPFPA was functionalized with dual moieties:azide groups for covalent immobilization of DBCO-modified DNA primers via click chemistry and tunable hydrophilic side chains to optimize biocompatibility and surface properties.Systematic screening revealed that hydrophobic azide carriers combined with neutral hydroxyl groups maximized the DNA immobilization efficacy,approaching the performance of commercial polyacrylamide-based polymers.The negatively charged carboxyl groups severely impede DNA primer attachment.Higher molecular weight derivatives further enhance the efficacy of DNA immobilization.In NGS validation,optimized surface modification polymers achieved robust surface density of clustered DNA and high sequencing accuracy,surpassing quality benchmarks and comparable to those of conventional analogs.This platform demonstrates significant potential for tailoring high-sensitivity surfaces for genomic applications,advancing clinical diagnostics,and personalized medicine.展开更多
The improvement of soybean seed carotenoid contents is very important due to the beneficial role of carotenoids in human health and nutrition. However, the genetic architecture underlying soybean carotenoid biosynthes...The improvement of soybean seed carotenoid contents is very important due to the beneficial role of carotenoids in human health and nutrition. However, the genetic architecture underlying soybean carotenoid biosynthesis remains largely unknown. In the present study, we employed next generation sequencing-based bulked-segregant analysis to identify new genomic regions governing seed carotenoids in 1,551 natural soybean accessions. The genomic DNA samples of individual plants with extreme phenotypes were pooled to form two bulks with high(50 accessions) and low(50 accessions) carotenoid contents for Illumina sequencing. A total of 125.09 Gb of clean bases and 89.82% of Q30 were obtained, and the average alignment efficiency was 99.45% with an average coverage depth of 62.20× and 99.75% genome coverage. Based on the G prime statistic algorithm(G') method analysis, 16 candidate genomic loci with a total length 20.41 Mb were found to be related to the trait. Of these loci, the most significant regions displaying the highest elevated G' values were found in chromosome 06 at a position of 18.53–22.67 Mb, and chromosome 19 at genomic region intervals of 8.36–10.94, 12.06–13.79 and 18.45–20.26 Mb. These regions were then used to identify the key candidate genes. In these regions, 250 predicted genes were found and analyzed to obtain 90 significantly enriched(P<0.05) Gene Ontology(GO) terms. Based on ANNOVAR analysis, 50 genes with non-synonymous and stopgained mutations were preferentially selected as potential candidate genes. Of those 50 genes, following their gene annotation functions and high significant haplotype variations in various environments,five genes were identified as the most promising candidate genes regulating soybean seed carotenoid accumulation, and they should be investigated in further functional validation studies. Collectively, understanding the genetic basis of carotenoid pigments and identifying genes underpinning carotenoid accumulation via a bulked-segregant analysis-based sequencing(BSA-seq) approach provide new insights for exploring future molecular breeding efforts to produce soybean cultivars with high carotenoid content.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
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
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
Next-generation sequencing(NGS) technology is capable of sequencing millions or billions of DNA molecules simultaneously.Therefore, it represents a promising tool for the analysis of molecular targets for the initial ...Next-generation sequencing(NGS) technology is capable of sequencing millions or billions of DNA molecules simultaneously.Therefore, it represents a promising tool for the analysis of molecular targets for the initial diagnosis of disease, monitoring of disease progression, and identifying the mechanism of drug resistance. On behalf of the Tumor Biomarker Committee of the Chinese Society of Clinical Oncology(CSCO) and the China Actionable Genome Consortium(CAGC), the present expert group hereby proposes advisory guidelines on clinical applications of NGS technology for the analysis of cancer driver genes for precision cancer therapy. This group comprises an assembly of laboratory cancer geneticists, clinical oncologists, bioinformaticians,pathologists, and other professionals. After multiple rounds of discussions and revisions, the expert group has reached a preliminary consensus on the need of NGS in clinical diagnosis, its regulation, and compliance standards in clinical sample collection. Moreover, it has prepared NGS criteria, the sequencing standard operation procedure(SOP), data analysis, report, and NGS platform certification and validation.展开更多
Cystic echinococcosis (CE) is a prevalent zoonotic disease caused by Echinococcus granulosus, with a cosmopolitan distribution. The parasite is transmitted cyclically between canines and numerous intermediate herbivor...Cystic echinococcosis (CE) is a prevalent zoonotic disease caused by Echinococcus granulosus, with a cosmopolitan distribution. The parasite is transmitted cyclically between canines and numerous intermediate herbivorous livestock animals. Also, other Taeniid tapeworms could infect domestic dogs and they pose significant veterinary and public health concerns worldwide. This study aimed to develop a sensitive molecular method for detecting Echinococcus spp. DNA in dog fecal samples using next-generation sequencing (NGS). A set of PCR primers targeting conserved regions of Taeniid tapeworms’ 18s rRNA genes was designed and tested for amplifying genomic DNA from various tapeworm species. The PCR system demonstrated high sensitivity, amplifying DNA from all tested tapeworm species, with differences observed in amplified band sizes. The primers were adapted for NGS analysis by adding forward and reverse adapters, enabling the sequencing of amplified DNA fragments. Application of the developed PCR system to dog fecal samples collected from Yatta town, Palestine, revealed the presence of E. granulosus DNA in five out of 50 samples. NGS analysis confirmed the specificity of the amplified DNA fragments, showing 98% - 99% similarity with the 18s rDNA gene of E. granulosus. This study demonstrates the utility of NGS-based molecular methods for accurate and sensitive detection of Echinococcus spp. in dog fecal samples, providing valuable insights for epidemiological surveillance and control programs of echinococcosis in endemic regions.展开更多
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.
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Supported by the Natural Science Foundation of Shanghai Science Committee,No.18ZR1431200Research Foundation of Shanghai Municipal Health Commission,No.20194Y0112Clinical Research Plan of SHDC,No.SHDC2020CR4089.
文摘BACKGROUND Hereditary spherocytosis(HS)is characterized by anemia,jaundice,splenomegaly,and cholelithiasis,and is caused by abnormal genes encoding red blood cell membrane components.The most common mutations found in HS are in the ANK1 gene.CASE SUMMARY A 4-mo-old girl was admitted to our hospital with pallor that had lasted for more than 2 mo.She presented with jaundice,anemia and splenomegaly.A heterozygous mutation of ANK1(exon23:c.G2467T:p.E823X)was identified,and the mutation was determined to be autosomal dominant.This mutation is linked to the relatively serious anemia she had after birth;this anemia improved with age.CONCLUSION The utilization of next-generation sequencing may assist with the accurate diagnosis of HS,especially in atypical cases.
基金supported by the Health Commission of Shenzhen Municipality under the Sanming Project of Medicine(SZSM201911014)in Shenzhenthe High Level-Hospital Program,Health Commission of Guangdong Province,China.
文摘The expanding application of targeted next-generation sequencing(tNGS)in clinical diagnostics has led to an increasing frequency of Nocardia spp.detection in respiratory specimens;however,the clinical relevance and interpretation of these findings remain uncertain.This study describes the epidemiology of Nocardia pneumonia and colonization in patients at The University of Hong Kong–Shenzhen Hospital,China,and evaluates the diagnostic role of tNGS.The analysis was conducted from January 2023 through December 2024 and included 22 patients in whom Nocardia spp.were detected in respiratory specimens via tNGS.Among these patients,81.8%(18/22)had comorbidities,most commonly chronic lung disease(40.9%,9/22).Common clinical symptoms included cough,sputum and fever.Nocardia farcinica and Nocardia abscessus were the predominant species identified.Although the median number of se-quence reads of Nocardia were higher in infected patients than in colonized individuals(289,IQR:132.5–3747 vs.133.5,IQR:51.75–261.3),the difference was not statistically significant(P>0.05).In a comparison of diagnostic performance,tNGS identified Nocardia sequences in all 25 respiratory samples collected from the enrolled patients,whereas conventional aerobic culture success-fully isolated the pathogen in only two patients.Additionally,tNGS exhibited a significantly shorter median turnaround time than culture(38 h vs.376.5 h).These findings highlight tNGS as a rapid tool for Nocardia detection.However,the final diagnosis of nocardiosis can-not rely solely on sequence read thresholds.Clinical,radiological and laboratory integration remains critical in distinguishing true infection from colonization,ensuring accurate diagnosis and management of nocardiosis.
基金Supported by the Guangdong Provincial Key Laboratory of Pathogen Detection for Emerging Infectious Disease Response(2023B1212010010).
文摘Introduction:Vaccination stands as the most effective preventive measure against yellow fever(YF).However,the YF vaccination associated Adverse Events following Immunization(AEFI)cases occur occasionally.Methods:The Guangdong Provincial Center for Disease Control and Prevention utilized Targeted Next-Generation Sequencing(tNGS)to determine whether the imported suspected YF case was infected by the wild-type YF virus strain or experienced an AEFI.Results:tNGS analysis successfully yielded a 10.2 kb viral genomic sequence.Subsequent in-depth analysis revealed high similarity to the YF vaccine strain 17D-213 and classified the sequence within the West Africa II genotype,clustering with the 17D vaccine strain.Conclusions and Implications for Public Health Practice:This case represented YF AEFI.The implementation of tNGS technology enables more precise and expeditious pathogen sequencing,providing critical evidence for accurate disease diagnosis and informed public health interventions.
文摘Objective To screen gene mutation in adult-onset hypoparathyroidism in Chinese through the targeted nextgeneration sequencing(NGS).Methods We recruited 17patients with adult-onset hypoparathyroidism who were regularly followed or newly diagnosed at our centre during the past one year.Nine of them developed hypercalciuria during the treatment with calcium and vitamin D.
基金supported by the German Centre for Cardiovascular Research (DZHK), the European Union (FP7 Inheritance and FP7 Best Ageing), and the German Ministry of Education and Research (BMBF)
文摘Next-generation sequencing （NGS） is getting routinely used in the diagnosis of hereditary diseases, such as human cardiomyopathies. Hence. it is of utter importance to secure high quality sequencing data, enabling the identification of disease-relevant mutations or the conclusion of negative test results. During the process of sample preparation, each protocol for target enrichment library preparation has its own requirements for quality control （QC）; however, there is little evi- dence on the actual impact of these guidelines on resulting data quality. In this study, we analyzed the impact of QC during the diverse library preparation steps of Agilent SureSelect XT target enrichment and lllumina sequencing. We quantified the parameters for a cohort of around 600 samples, which include starting amount of DNA, amount of sheared DNA, smallest and largest fragment size of the starting DNA; amount of DNA after the pre-PCR, and smallest and largest fragment size of the resulting DNA; as well as the amount of the final library, the corresponding smallest and largest fragment size, and the number of detected variants. Intriguingly, there is a high tolerance for variations in all QC steps, meaning that within the boundaries proposed in the current study, a considerable variance at each step of QC can be well tolerated without compromising NGS quality.
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.31972983 and 32072487)the Key Technology R&D Program of Zhejiang Province,China(Grant No.2021C02006)the Zhejiang Provincial Natural Science Foundation of China(Grant No.LY23C140001).
文摘In rice fields,rice plants usually grow alongside wild weeds and are attacked by various invertebrate species.Viruses are abundant in plants and invertebrates,playing crucial ecological roles in controlling microbial abundance and maintaining community structures.To date,only 16 rice viruses have been documented in rice-growing regions.These viruses pose serious threats to rice production and have traditionally been identified only from rice plants and insect vectors by isolation techniques.Advances in next-generation sequencing(NGS)have made it feasible to discover viruses on a global scale.Recently,numerous viruses have been identified in plants and invertebrates using NGS technologies.In this review,we discuss viral studies in rice plants,invertebrate species,and weeds in rice fields.Many novel viruses have been discovered in rice ecosystems through NGS technologies,with some also detected using metatranscriptomic and small RNA sequencing.These analyses greatly expand our understanding of viruses in rice fields and provide valuable insights for developing efficient strategies to manage insect pests and virus-mediated rice diseases.
文摘This study investigates the diversity of gut microbiota in Metaphire peguana,an earthworm species commonly found in agricultural areas of Thailand.Earthworms play a critical role in soil ecosystems by supporting nutrient cycling and breaking down organic matter.Understanding the microbial diversity in their gut is essential for exploring their ecological contributions.Using Next Generation Sequencing(NGS),we analyzed the mycobiome in the gut of M.peguana.Our findings revealed a high diversity of fungal species,primarily belonging to two major phyla:Ascomycota and Basidiomycota.Ascomycota was the most abundant phylum,comprising 40.1% of the total fungal species identified.A total of 33 distinct fungal species were identified,which underscores the richness of microbial life within the earthworm gut.This study successfully created the first genetic database of the microbial community in M.peguana,providing a foundation for future research in agricultural applications.The microbial species identified,particularly siderophoreproducing fungi,could have significant implications for improving soil fertility and promoting sustainable agricultural practices.The use of NGS technology has enabled comprehensive profiling of microbial communities,allowing for precise identification of fungi that may play essential roles in soil health.Furthermore,the study paves the way for future studies on the potential applications of earthworm gut microbiomes in biotechnology,especially in enhancing soil nutrient availability and plant growth.The findings of this research contribute to the broader understanding of the ecological roles of earthworms and their microbiomes in soil ecosystems.
基金supported by the Hubei Provincial Natural Science Foundation of China(No.2023AFB646)Knowledge Innovation Program of Wuhan(No.2023020201010155)Educational Research Program of Huazhong University of Science and Technology(No.2022135).
文摘Objective and Background Early and accurate diagnosis of spinal infections,including spinal tuberculosis,is pivotal for effective treatment but remains challenging.This study aims to assess the diagnostic yield of metagenomic next-generation sequencing(mNGS)compared with that of conventional microbiological tests(CMTs)in identifying pathogens associated with spinal pathologies,with a special focus on infections leading to surgical interventions.Methods We enrolled 85 patients who underwent spinal surgery,comprising 63 patients with clinically diagnosed spinal infections,including patients with spinal tuberculosis,and 22 patients with noninfectious spinal conditions.The procedures involved irrigation and debridement for persistent wound drainage,with subsequent DNA extraction from plasma and joint fluid for mNGS and CMT analysis.Results Significantly increased C-reactive protein(CRP)levels were observed in patients with infections.The mNGS approach showed greater diagnostic sensitivity(92.06%)for detecting pathogens,including Mycobacterium tuberculosis,than did CMTs(36.51%).Despite its low specificity,mNGS had considerable negative predictive value(70.59%),underscoring its utility in ruling out infections.Conclusions The mNGS offers superior sensitivity over CMTs in the diagnosis of a variety of spinal infections,notably spinal tuberculosis.This study highlights the potential of mNGS in enhancing the diagnosis of complex spinal infections,thereby informing targeted treatment strategies.
文摘BACKGROUND Leuconostoc garlicum is commonly found in fermented foods and very few infected patients have been reported,who typically present symptoms such as fever and fatigue.Conventional clinical examinations often struggle to identify this bacterium,and routine anti-infective treatments are generally ineffective.Both diagnostic challenges and therapeutic limitations pose significant difficulties for clinicians.CASE SUMMARY We report a patient ultimately diagnosed with Leuconostoc garlicum infection.The primary manifestations included persistent fever,cough and fatigue.These symptoms lasted for 2 months.He received anti-infective treatment at a community hospital,but this was ineffective.After inquiring about the patient's medical history and conducting a physical examination,the patient underwent laboratory tests.Complete blood count tests revealed that the patient had a high proportion of neutrophils,C-reactive protein level was 235.9 mg/L,erythrocyte sedimentation rate was 67 mm/h,respiratory pathogen testing was negative,and he was then thought to have an infectious disease.However,conventional anti-infective treatments were ineffective.After excluding infectious neurological diseases,urologic diseases and digestive problems,we ultimately focused our attention on the lungs.A lung computed tomography scan indicated pulmonary inflammation.Bronchoalveolar lavage fluid for next-generation sequencing suggested lung infection with Leuconostoc garlicum.The patient's symptoms gradually improved following treatment with piperacillin tazobactam and linezolid.During the follow-up period,the patient's temperature remained normal.CONCLUSION For patients with suspected bacterial infection and experiencing fever,conventional anti-infective treatment can be ineffective in controlling their symptoms,and an infection due to rare bacteria or drug-resistant bacteria should be considered.Next-generation sequencing enables rapid and precise identification of infection-related pathogens in febrile patients.
基金financially supported by the Science and Technology Commission of Shanghai Municipality(No.24ZR1401400)Shenzhen Salus Bio Med Company for their strong support in this study。
文摘In this study,an amine-reactive poly(pentafluorophenyl acrylate)(PPFPA)platform was developed for advanced surface engineering of next-generation sequencing(NGS)chips.Through post-polymerization modification,PPFPA was functionalized with dual moieties:azide groups for covalent immobilization of DBCO-modified DNA primers via click chemistry and tunable hydrophilic side chains to optimize biocompatibility and surface properties.Systematic screening revealed that hydrophobic azide carriers combined with neutral hydroxyl groups maximized the DNA immobilization efficacy,approaching the performance of commercial polyacrylamide-based polymers.The negatively charged carboxyl groups severely impede DNA primer attachment.Higher molecular weight derivatives further enhance the efficacy of DNA immobilization.In NGS validation,optimized surface modification polymers achieved robust surface density of clustered DNA and high sequencing accuracy,surpassing quality benchmarks and comparable to those of conventional analogs.This platform demonstrates significant potential for tailoring high-sensitivity surfaces for genomic applications,advancing clinical diagnostics,and personalized medicine.
基金financially supported by the National Natural Science Foundation of China (32161143033, 32272178, and 32001574)National Key Research and Development Program of China (2021YFD1201605)the Agricultural Science and Technology Innovation Project of CAAS。
文摘The improvement of soybean seed carotenoid contents is very important due to the beneficial role of carotenoids in human health and nutrition. However, the genetic architecture underlying soybean carotenoid biosynthesis remains largely unknown. In the present study, we employed next generation sequencing-based bulked-segregant analysis to identify new genomic regions governing seed carotenoids in 1,551 natural soybean accessions. The genomic DNA samples of individual plants with extreme phenotypes were pooled to form two bulks with high(50 accessions) and low(50 accessions) carotenoid contents for Illumina sequencing. A total of 125.09 Gb of clean bases and 89.82% of Q30 were obtained, and the average alignment efficiency was 99.45% with an average coverage depth of 62.20× and 99.75% genome coverage. Based on the G prime statistic algorithm(G') method analysis, 16 candidate genomic loci with a total length 20.41 Mb were found to be related to the trait. Of these loci, the most significant regions displaying the highest elevated G' values were found in chromosome 06 at a position of 18.53–22.67 Mb, and chromosome 19 at genomic region intervals of 8.36–10.94, 12.06–13.79 and 18.45–20.26 Mb. These regions were then used to identify the key candidate genes. In these regions, 250 predicted genes were found and analyzed to obtain 90 significantly enriched(P<0.05) Gene Ontology(GO) terms. Based on ANNOVAR analysis, 50 genes with non-synonymous and stopgained mutations were preferentially selected as potential candidate genes. Of those 50 genes, following their gene annotation functions and high significant haplotype variations in various environments,five genes were identified as the most promising candidate genes regulating soybean seed carotenoid accumulation, and they should be investigated in further functional validation studies. Collectively, understanding the genetic basis of carotenoid pigments and identifying genes underpinning carotenoid accumulation via a bulked-segregant analysis-based sequencing(BSA-seq) approach provide new insights for exploring future molecular breeding efforts to produce soybean cultivars with high carotenoid content.
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.
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.
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 grants from Guangdong Provincial Key Lab of Translational Medicine in Lung Cancer (Grant No. 2017B030314120)General Research Project of Guangzhou Science and Technology Bureau (Grant No. 201607010391)+1 种基金National Key Research and Development Program of China (Grant No. 2016YFC1303800)Guangdong Provincial Applied S&T R&D Program (Grant No. 2016B020237006)
文摘Next-generation sequencing(NGS) technology is capable of sequencing millions or billions of DNA molecules simultaneously.Therefore, it represents a promising tool for the analysis of molecular targets for the initial diagnosis of disease, monitoring of disease progression, and identifying the mechanism of drug resistance. On behalf of the Tumor Biomarker Committee of the Chinese Society of Clinical Oncology(CSCO) and the China Actionable Genome Consortium(CAGC), the present expert group hereby proposes advisory guidelines on clinical applications of NGS technology for the analysis of cancer driver genes for precision cancer therapy. This group comprises an assembly of laboratory cancer geneticists, clinical oncologists, bioinformaticians,pathologists, and other professionals. After multiple rounds of discussions and revisions, the expert group has reached a preliminary consensus on the need of NGS in clinical diagnosis, its regulation, and compliance standards in clinical sample collection. Moreover, it has prepared NGS criteria, the sequencing standard operation procedure(SOP), data analysis, report, and NGS platform certification and validation.
文摘Cystic echinococcosis (CE) is a prevalent zoonotic disease caused by Echinococcus granulosus, with a cosmopolitan distribution. The parasite is transmitted cyclically between canines and numerous intermediate herbivorous livestock animals. Also, other Taeniid tapeworms could infect domestic dogs and they pose significant veterinary and public health concerns worldwide. This study aimed to develop a sensitive molecular method for detecting Echinococcus spp. DNA in dog fecal samples using next-generation sequencing (NGS). A set of PCR primers targeting conserved regions of Taeniid tapeworms’ 18s rRNA genes was designed and tested for amplifying genomic DNA from various tapeworm species. The PCR system demonstrated high sensitivity, amplifying DNA from all tested tapeworm species, with differences observed in amplified band sizes. The primers were adapted for NGS analysis by adding forward and reverse adapters, enabling the sequencing of amplified DNA fragments. Application of the developed PCR system to dog fecal samples collected from Yatta town, Palestine, revealed the presence of E. granulosus DNA in five out of 50 samples. NGS analysis confirmed the specificity of the amplified DNA fragments, showing 98% - 99% similarity with the 18s rDNA gene of E. granulosus. This study demonstrates the utility of NGS-based molecular methods for accurate and sensitive detection of Echinococcus spp. in dog fecal samples, providing valuable insights for epidemiological surveillance and control programs of echinococcosis in endemic regions.
