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1.
Nat Commun ; 12(1): 4702, 2021 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-34349104

RESUMO

Mycobacterium tuberculosis can adapt to changing environments by non-heritable mechanisms. Frame-shifting insertions and deletions (indels) may also participate in adaptation through gene disruption, which could be reversed by secondary introduction of a frame-restoring indel. We present ScarTrek, a program that scans genomic data for indels, including those that together disrupt and restore a gene's reading frame, producing "frame-shift scars" suggestive of reversible gene inactivation. We use ScarTrek to analyze 5977 clinical M. tuberculosis isolates. We show that indel frequency inversely correlates with genomic linguistic complexity and varies with gene-position and gene-essentiality. Using ScarTrek, we detect 74 unique frame-shift scars in 48 genes, with a 3.74% population-level incidence of unique scar events. We find multiple scars in the ESX-1 gene cluster. Six scars show evidence of convergent evolution while the rest shared a common ancestor. Our results suggest that sequential indels are a mechanism for reversible gene silencing and adaptation in M. tuberculosis.


Assuntos
Adaptação Fisiológica/genética , Inativação Gênica , Mycobacterium tuberculosis/genética , Fases de Leitura/genética , Evolução Molecular , Genes Bacterianos/genética , Genoma Bacteriano/genética , Humanos , Mutação INDEL , Mycobacterium tuberculosis/isolamento & purificação , Tuberculose/microbiologia
2.
Gene ; 802: 145867, 2021 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-34352299

RESUMO

Genome-wide association studies (GWAS) have identified DENND1A as a potential candidate gene linked to the fertility-related phenotypes in dairy cows. However, to date, no studies have examined the association of the DENND1A insertion/deletions (indels) to bovine fertility on a large scale. Herein, two indel sites, including P4-del-26-bp and P8-ins-15-bp were identified in 1064 Holstein cows. The values of the minor allelic frequency (MAF) ranged between 0.471 (deletion) and 0.230 (deletion), respectively, and combined four different haplotypes by analyzing the haplotype combination. It is noteworthy that P4-del-26-bp is associated with the ovarian width (P = 0.0004) and corpus luteum diameter (P = 0.004). Meanwhile, P8-ins-15-bp was found to have a significant association with the ovarian width (P = 0.020), ovarian weight (P = 0.004), the number of mature follicles (P = 0.020), and diameter of the mature follicles (P = 0.016). Furthermore, the combinatorial analysis showed that the two indel combined-genotypes were significantly related to several reproductive traits (ovarian width, ovarian weight, etc.). Collectively, our findings indicated that these two novel indels and their combinations are correlated with the reproductive traits, and hence, they can serve in the marker-assisted selection (MAS) in cattle breeding. Nevertheless, further functional experiments are needed for understanding the mechanisms of these indels in cattle reproduction in a better way.


Assuntos
Doenças dos Bovinos/genética , Mutação INDEL , Proteínas de Membrana Transportadoras/genética , Síndrome do Ovário Policístico/genética , Animais , Bovinos , Feminino , Fertilidade/genética , Frequência do Gene , Estudo de Associação Genômica Ampla/veterinária , Haplótipos , Reprodução/genética
3.
BMC Genomics ; 22(1): 595, 2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34353285

RESUMO

BACKGROUND: Cannabis sativa L., a dioecious plant derived from China, demonstrates important medicinal properties and economic value worldwide. Cannabis properties have been usually harnessed depending on the sex of the plant. To analyse the genetic structure of Chinese Cannabis and identify sex-linked makers, genome-wide insertion-deletion (InDel) markers were designed and used. RESULTS: In this study, a genome-wide analysis of insertion-deletion (InDel) polymorphisms was performed based on the recent genome sequences. In total, 47,558 InDels were detected between the two varieties, and the length of InDels ranged from 4 bp to 87 bp. The most common InDels were tetranucleotides, followed by pentanucleotides. Chromosome 5 exhibited the highest number of InDels among the Cannabis chromosomes, while chromosome 10 exhibited the lowest number. Additionally, 31,802 non-redundant InDel markers were designed, and 84 primers evenly distributed in the Cannabis genome were chosen for polymorphism analysis. A total of 38 primers exhibited polymorphisms among three accessions, and of the polymorphism primers, 14 biallelic primers were further used to analyse the genetic structure. A total of 39 fragments were detected, and the PIC value ranged from 0.1209 to 0.6351. According to the InDel markers and the flowering time, the 115 Chinese germplasms were divided into two subgroups, mainly composed of cultivars obtained from the northernmost and southernmost regions, respectively. Additional two markers, "Cs-I1-10" and "Cs-I1-15", were found to amplify two bands (398 bp and 251 bp; 293 bp and 141 bp) in the male plants, while 389-bp or 293-bp bands were amplified in female plants. Using the two markers, the feminized and dioecious varieties could also be distinguished. CONCLUSION: Based on the findings obtained herein, we believe that this study will facilitate the genetic improvement and germplasm conservation of Cannabis in China, and the sex-linked InDel markers will provide accurate sex identification strategies for Cannabis breeding and production.


Assuntos
Cannabis , Cannabis/genética , China , Marcadores Genéticos , Genoma , Mutação INDEL , Melhoramento Vegetal
4.
BMC Plant Biol ; 21(1): 363, 2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34376154

RESUMO

BACKGROUND: To meet increasing demand for forest-based products and protect natural forests from further deforestation requires increased productivity from planted forests. Genetic improvement of conifers by traditional breeding is time consuming due to the long juvenile phase and genome complexity. Genetic modification (GM) offers the opportunity to make transformational changes in shorter time frames but is challenged by current genetically modified organism (GMO) regulations. Genome editing, which can be used to generate site-specific mutations, offers the opportunity to rapidly implement targeted improvements and is globally regulated in a less restrictive way than GM technologies. RESULTS: We have demonstrated CRISPR/Cas9 genome editing in P. radiata targeting a single-copy cell wall gene GUX1 in somatic embryogenic tissue and produced plantlets from the edited tissue. We generated biallelic INDELs with an efficiency of 15 % using a single gRNA. 12 % of the transgenic embryogenic tissue was edited when two gRNAs were used and deletions of up to 1.3 kb were identified. However, the regenerated plants did not contain large deletions but had single nucleotide insertions at one of the target sites. We assessed the use of CRISPR/Cas9 ribonucleoproteins (RNPs) for their ability to accomplish DNA-free genome editing in P. radiata. We chose a hybrid approach, with RNPs co-delivered with a plasmid-based selectable marker. A two-gRNA strategy was used which produced an editing efficiency of 33 %, and generated INDELs, including large deletions. Using the RNP approach, deletions found in embryogenic tissue were also present in the plantlets. But, all plants produced using the RNP strategy were monoallelic. CONCLUSIONS: We have demonstrated the generation of biallelic and monoallelic INDELs in the coniferous tree P. radiata with the CRISPR/Cas9 system using plasmid expressed Cas9 gRNA and RNPs respectively. This opens the opportunity to apply genome editing in conifers to rapidly modify key traits of interest.


Assuntos
Sistemas CRISPR-Cas , Edição de Genes/métodos , Genoma de Planta , Pinus/genética , DNA de Plantas , Mutação INDEL , RNA Guia , Ribonucleoproteínas/genética
5.
BMC Bioinformatics ; 22(1): 404, 2021 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-34391391

RESUMO

BACKGROUND: Modern Next Generation- and Third Generation- Sequencing methods such as Illumina and PacBio Circular Consensus Sequencing platforms provide accurate sequencing data. Parallel developments in Deep Learning have enabled the application of Deep Neural Networks to variant calling, surpassing the accuracy of classical approaches in many settings. DeepVariant, arguably the most popular among such methods, transforms the problem of variant calling into one of image recognition where a Deep Neural Network analyzes sequencing data that is formatted as images, achieving high accuracy. In this paper, we explore an alternative approach to designing Deep Neural Networks for variant calling, where we use meticulously designed Deep Neural Network architectures and customized variant inference functions that account for the underlying nature of sequencing data instead of converting the problem to one of image recognition. RESULTS: Results from 27 whole-genome variant calling experiments spanning Illumina, PacBio and hybrid Illumina-PacBio settings suggest that our method allows vastly smaller Deep Neural Networks to outperform the Inception-v3 architecture used in DeepVariant for indel and substitution-type variant calls. For example, our method reduces the number of indel call errors by up to 18%, 55% and 65% for Illumina, PacBio and hybrid Illumina-PacBio variant calling respectively, compared to a similarly trained DeepVariant pipeline. In these cases, our models are between 7 and 14 times smaller. CONCLUSIONS: We believe that the improved accuracy and problem-specific customization of our models will enable more accurate pipelines and further method development in the field. HELLO is available at https://github.com/anands-repo/hello.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Redes Neurais de Computação , Humanos , Mutação INDEL
6.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34210100

RESUMO

Cas endonuclease-mediated genome editing provides a long-awaited molecular biological approach to the modification of predefined genomic target sequences in living organisms. Although cas9/guide (g)RNA constructs are straightforward to assemble and can be customized to target virtually any site in the plant genome, the implementation of this technology can be cumbersome, especially in species like triticale that are difficult to transform, for which only limited genome information is available and/or which carry comparatively large genomes. To cope with these challenges, we have pre-validated cas9/gRNA constructs (1) by frameshift restitution of a reporter gene co-introduced by ballistic DNA transfer to barley epidermis cells, and (2) via transfection in triticale protoplasts followed by either a T7E1-based cleavage assay or by deep-sequencing of target-specific PCR amplicons. For exemplification, we addressed the triticale ABA 8'-hydroxylase 1 gene, one of the putative determinants of pre-harvest sprouting of grains. We further show that in-del induction frequency in triticalecan beincreased by TREX2 nuclease activity, which holds true for both well- and poorly performing gRNAs. The presented results constitute a sound basis for the targeted induction of heritable modifications in triticale genes.


Assuntos
Sistema Enzimático do Citocromo P-450/metabolismo , Edição de Genes/métodos , Proteínas de Plantas/metabolismo , Triticale/metabolismo , Sistemas CRISPR-Cas , Sistema Enzimático do Citocromo P-450/genética , Genes Reporter , Mutação INDEL , Mutagênese Sítio-Dirigida , Proteínas de Plantas/genética , Transfecção , Triticale/genética
7.
Methods Mol Biol ; 2312: 225-233, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34228293

RESUMO

The CRISPR-Cas9 system offers targeted genome manipulation with simplicity. Combining the CRISPR-Cas9 with optogenetics technology, we have engineered photoactivatable Cas9 to precisely control the genome sequence in a spatiotemporal manner. Here we provide a detailed protocol for optogenetic genome editing experiments using photoactivatable Cas9, including that for the generation of guide RNA vectors, light-mediated Cas9 activation, and quantification of genome editing efficiency in mammalian cells.


Assuntos
Proteína 9 Associada à CRISPR/efeitos da radiação , Sistemas CRISPR-Cas/efeitos da radiação , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Edição de Genes , Regulação da Expressão Gênica/efeitos da radiação , Luz , Optogenética , Proteína 9 Associada à CRISPR/genética , Proteína 9 Associada à CRISPR/metabolismo , Técnicas de Cultura de Células , Reparo do DNA por Junção de Extremidades , Células HEK293 , Humanos , Mutação INDEL , Mutação Puntual , RNA Guia/genética , RNA Guia/metabolismo
8.
Theranostics ; 11(14): 7092-7109, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34093873

RESUMO

Background: Recent studies in non-small cell lung cancer (NSCLC) patients have demonstrated that first-line immunotherapy is associated with better therapeutic response than second-line treatment. So far, the mechanisms need to be explored. It prompted us to evaluate the association between first-line chemotherapy and subsequent immunotherapy in NSCLC as well as its underlying mechanisms at the genomic and transcriptomic level. Methods: We launched a prospective, observational clinical study, paired tumor biopsies before and after chemotherapy were collected from NSCLC patients without tyrosine kinase inhibitor (TKI)-related driver gene mutations. The analyses included genomic and transcriptional changes performed by next-generation sequencing (NGS)-based whole-exome sequencing (WES) and messager ribonucleic acid (mRNA) sequencing. Characteristic mutational alterations in 1574 genes were investigated based on mutational status, clinicopathological factors, and chemotherapy responses. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, neoantigen prediction and intratumoral heterogeneity evaluation were also performed. Results: Samples and information from 32 NSCLC patients without TKI-related driver gene mutations were obtained. We found that the total number of single nucleotide variants (SNV)/insertion-deletion (INDEL) mutations did not change significantly after chemotherapy. The tumor mutation burden (TMB) decreased significantly after chemotherapy in smoking patients and the decreased TMB correlated with a better survival of smoking patients. The change in copy number variations (CNVs) exhibited a decreasing trend during chemotherapy. Subsequent analysis at mRNA level revealed a significant decrease in the expression levels of genes related to antigen processing and presentation as well as other factors relevant for response to immunotherapy. Pathway enrichment analysis confirmed that the immune-related signaling pathways or biological processes were decreased after first-line chemotherapy. Conclusions: Our study presents an explanation for the unsatisfactory results of immunotherapy when given after chemotherapy, and suggests that first-line chemotherapy is able to influence the tumor microenvironment and decrease the efficacy of subsequent immunotherapy. The study was registered at ClinicalTrials.gov, number NCT03764917, and has completed enrolment; patients are still in follow-up.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Imunoterapia/métodos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Biópsia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/terapia , Inibidores Enzimáticos/metabolismo , Perfilação da Expressão Gênica , Ontologia Genética , Genômica , Humanos , Mutação INDEL , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/terapia , Estudos Observacionais como Assunto , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Estudos Prospectivos , RNA-Seq , Fumantes , Microambiente Tumoral , Sequenciamento Completo do Exoma
9.
J Forensic Sci ; 66(5): 1973-1979, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34106477

RESUMO

Sampling of healthy multi-rooted teeth is recommended for the genetic identification of human skeletal remains. However, this may not always be possible, as in the reported case consisting of an isolated human cranium found in an aggregate crushing and processing plant in Piedmont, Northwest Italy. The cranium displayed significant weathering, suggesting a post-mortem interval of several years, and was edentulous with the exception of the apical root fragment of the upper left canine, consequence of an antemortem horizontal fracture. Prolonged decalcification of the root fragment followed by powder-free DNA extraction from ~10 mg of root tip tissue led to the recovery of >10 ng of high molecular weight human DNA, in comparison with ~0.01 ng of DNA per mg of bone powder obtained from the petrous portion of the temporal bone. Quantity and quality of DNA isolated from apical tooth tissue enabled multiple genotyping, including a reportable female STR profile, mitochondrial DNA analysis, and ancestry-informative insertion/deletion polymorphisms. Although the cranium remained unidentified after DNA comparisons, our findings confirm that apical tooth tissue is a promising source of DNA, easily obtained through a powder-free extraction protocol. Results also indicate that root tips should not be overlooked in challenging identification cases, even in the presence of compromised tooth specimens.


Assuntos
Impressões Digitais de DNA/métodos , DNA/isolamento & purificação , Raiz Dentária/química , DNA Mitocondrial/genética , Feminino , Humanos , Mutação INDEL , Repetições de Microssatélites , Crânio , Ápice Dentário
10.
Medicine (Baltimore) ; 100(19): e25910, 2021 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-34106654

RESUMO

ABSTRACT: It is well known that many genetic factors are involved in the occurrence and progression of atrioventricular block (AV block) and atrial fibrillation (AF). However, the genetic variants discovered so far have only explained parts of these processes. More genes and variants remain to be identified. In the present study, a three-generation family with an autosomal dominant form of AV block and AF was enrolled. Whole exome sequencing was conducted in three affected and one unaffected family member. A total of 64 nonsynonymous variants was shared by three affected individuals and not present in the unaffected individual. By selection of variants absent in the known databases and were predicted to be deleterious, 4 novel variants were identified. Only one novel frameshift insertion in the LMNA gene (c.825_826insCAGG) was identified in another affected family member and not detected in other non-affected family members and the 100 controls. Our finding expanded the spectrum of variants associated with AV block and AF, and was valuable in the genetic diagnosis of AV block and AF.


Assuntos
Fibrilação Atrial/genética , Bloqueio Atrioventricular/genética , Lamina Tipo A/genética , Adulto , Estudos de Casos e Controles , Feminino , Humanos , Mutação INDEL , Masculino , Pessoa de Meia-Idade , Sequenciamento Completo do Exoma , Adulto Jovem
11.
BMC Genomics ; 22(1): 407, 2021 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-34082700

RESUMO

BACKGROUND: Next-generation sequencing (NGS) is an efficient tool used for identifying pathogenic variants that cause Mendelian disorders. However, the lack of bioinformatics training of researchers makes the interpretation of identified variants a challenge in terms of precision and efficiency. In addition, the non-standardized phenotypic description of human diseases also makes it difficult to establish an integrated analysis pathway for variant annotation and interpretation. Solutions to these bottlenecks are urgently needed. RESULTS: We develop a tool named "Cruxome" to automatically annotate and interpret single nucleotide variants (SNVs) and small insertions and deletions (InDels). Our approach greatly simplifies the current burdensome task of clinical geneticists and scientists to identify the causative pathogenic variants and build personal knowledge reference bases. The integrated architecture of Cruxome offers key advantages such as an interactive and user-friendly interface and the assimilation of electronic health records of the patient. By combining a natural language processing algorithm, Cruxome can efficiently process the clinical description of diseases to HPO standardized vocabularies. By using machine learning, in silico predictive algorithms, integrated multiple databases and supplementary tools, Cruxome can automatically process SNVs and InDels variants (trio-family or proband-only cases) and clinical diagnosis records, then annotate, score, identify and interpret pathogenic variants to finally generate a standardized clinical report following American College of Medical Genetics and Genomics/ Association for Molecular Pathology (ACMG/AMP) guidelines. Cruxome also provides supplementary tools to examine and visualize the genes or variations in historical cases, which can help to better understand the genetic basis of the disease. CONCLUSIONS: Cruxome is an efficient tool for annotation and interpretation of variations and dramatically reduces the workload for clinical geneticists and researchers to interpret NGS results, simplifying their decision-making processes. We present an online version of Cruxome, which is freely available to academics and clinical researchers. The site is accessible at http://114.251.61.49:10024/cruxome/ .


Assuntos
Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Algoritmos , Biologia Computacional , Bases de Dados Genéticas , Variação Genética , Humanos , Mutação INDEL , Software
12.
Int J Mol Sci ; 22(11)2021 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-34073702

RESUMO

The combination of phage display technology with high-throughput sequencing enables in-depth analysis of library diversity and selection-driven dynamics. We applied short-read sequencing of the mutagenized region on focused display libraries of two homologous nucleic acid modification eraser proteins-AlkB and FTO-biopanned against methylated DNA. This revealed enriched genotypes with small indels and concomitant doubtful amino acid motifs within the FTO library. Nanopore sequencing of the entire display vector showed additional enrichment of large deletions overlooked by region-specific sequencing, and further impacted the interpretation of the obtained amino acid motifs. We could attribute enrichment of these corrupted clones to amplification bias due to arduous FTO display slowing down host cell growth as well as phage production. This amplification bias appeared to be stronger than affinity-based target selection. Recommendations are provided for proper sequence analysis of phage display data, which can improve motive discovery in libraries of proteins that are difficult to display.


Assuntos
Bacteriófagos/genética , Variação Genética , Sequenciamento de Nucleotídeos em Larga Escala , Biblioteca de Peptídeos , Mutação INDEL , Análise de Sequência de DNA
13.
Nat Commun ; 12(1): 3281, 2021 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-34078888

RESUMO

Engineered reproductive species barriers are useful for impeding gene flow and driving desirable genes into wild populations in a reversible threshold-dependent manner. However, methods to generate synthetic barriers are lacking in advanced eukaryotes. Here, to overcome this challenge, we engineer SPECIES (Synthetic Postzygotic barriers Exploiting CRISPR-based Incompatibilities for Engineering Species), an engineered genetic incompatibility approach, to generate postzygotic reproductive barriers. Using this approach, we create multiple reproductively isolated SPECIES and demonstrate their reproductive isolation and threshold-dependent gene drive capabilities in D. melanogaster. Given the near-universal functionality of CRISPR tools, this approach should be portable to many species, including insect disease vectors in which confinable gene drives could be of great practical utility.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Drosophila melanogaster/genética , Tecnologia de Impulso Genético/métodos , Genes Letais , Especiação Genética , Dinâmica Populacional , Animais , Proteína 9 Associada à CRISPR/deficiência , Proteína 9 Associada à CRISPR/genética , Drosophila melanogaster/metabolismo , Feminino , Fluxo Gênico , Mutação INDEL , Masculino , RNA Guia/genética , RNA Guia/metabolismo , Isolamento Reprodutivo
14.
Nat Commun ; 12(1): 3551, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34112792

RESUMO

Species harbor extensive structural variation underpinning recent adaptive evolution. However, the causality between genomic features and the induction of new rearrangements is poorly established. Here, we analyze a global set of telomere-to-telomere genome assemblies of a fungal pathogen of wheat to establish a nucleotide-level map of structural variation. We show that the recent emergence of pesticide resistance has been disproportionally driven by rearrangements. We use machine learning to train a model on structural variation events based on 30 chromosomal sequence features. We show that base composition and gene density are the major determinants of structural variation. Retrotransposons explain most inversion, indel and duplication events. We apply our model to Arabidopsis thaliana and show that our approach extends to more complex genomes. Finally, we analyze complete genomes of haploid offspring in a four-generation pedigree. Meiotic crossover locations are enriched for new rearrangements consistent with crossovers being mutational hotspots. The model trained on species-wide structural variation accurately predicts the position of >74% of newly generated variants along the pedigree. The predictive power highlights causality between specific sequence features and the induction of chromosomal rearrangements. Our work demonstrates that training sequence-derived models can accurately identify regions of intrinsic DNA instability in eukaryotic genomes.


Assuntos
Ascomicetos/genética , Ascomicetos/patogenicidade , Cromossomos/genética , Variação Genética , Genoma , Genômica/métodos , Aprendizado de Máquina , Meiose/genética , Arabidopsis/genética , Cromossomos/metabolismo , Simulação por Computador , Troca Genética , Eucariotos/genética , Evolução Molecular , Genes Duplicados , Estudo de Associação Genômica Ampla , Mutação INDEL , Modelos Genéticos , Linhagem , Filogenia , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único , Retroelementos/genética , Inversão de Sequência
15.
Int J Mol Sci ; 22(11)2021 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-34072027

RESUMO

A high-density genetic linkage map is essential for genetic and genomic studies including QTL mapping, genome assembly, and comparative genomic analysis. Here, we constructed a citrus high-density linkage map using SSR and SNP markers, which are evenly distributed across the citrus genome. The integrated linkage map contains 4163 markers with an average distance of 1.12 cM. The female and male linkage maps contain 1478 and 2976 markers with genetic lengths of 1093.90 cM and 1227.03 cM, respectively. Meanwhile, a genetic map comparison demonstrates that the linear order of common markers is highly conserved between the clementine mandarin and Poncirus trifoliata. Based on this high-density integrated citrus genetic map and two years of deciduous phenotypic data, two loci conferring leaf abscission phenotypic variation were detected on scaffold 1 (including 36 genes) and scaffold 8 (including 107 genes) using association analysis. Moreover, the expression patterns of 30 candidate genes were investigated under cold stress conditions because cold temperature is closely linked with the deciduous trait. The developed high-density genetic map will facilitate QTL mapping and genomic studies, and the localization of the leaf abscission deciduous trait will be valuable for understanding the mechanism of this deciduous trait and citrus breeding.


Assuntos
Mapeamento Cromossômico , Poncirus/genética , Locos de Características Quantitativas , Característica Quantitativa Herdável , Resposta ao Choque Frio , Biologia Computacional/métodos , Ligação Genética , Marcadores Genéticos , Humanos , Mutação INDEL , Repetições de Microssatélites , Fenótipo , Polimorfismo de Nucleotídeo Único
16.
Theranostics ; 11(13): 6427-6444, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33995666

RESUMO

Background: Reportedly, nasopharyngeal carcinoma (NPC) patients with MHC I Class aberration are prone to poor survival outcomes, which indicates that the deficiency of tumor neoantigens might represent a mechanism of immune surveillance escape in NPC. Methods: To clearly delineate the landscape of neoantigens in NPC, we performed DNA and RNA sequencing on paired primary tumor, regional lymph node metastasis and distant metastasis samples from 26 patients. Neoantigens were predicted using pVACseq pipeline. Subtype prediction model was built using random forest algorithm. Results: Portraying the landscape of neoantigens in NPC for the first time, we found that the neoantigen load of NPC was above average compared to that of other cancers in The Cancer Genome Atlas program. While the quantity and quality of neoantigens were similar among primary tumor, regional lymph node metastasis and distant metastasis samples, neoantigen depletion was more severe in metastatic sites than in primary tumors. Upon tracking the clonality change of neoantigens, we found that neoantigen reduction occurred during metastasis. Building a subtype prediction model based on reported data, we observed that subtype I lacked T cells and suffered from severe neoantigen depletion, subtype II highly expressed immune checkpoint molecules and suffered from the least neoantigen depletion, and subtype III was heterogenous. Conclusions: These results indicate that neoantigens are conducive to the guidance of clinical treatment, and personalized therapeutic vaccines for NPC deserve deeper basic and clinical investigations to make them feasible in the future.


Assuntos
Antígenos de Neoplasias/imunologia , Carcinoma Nasofaríngeo/secundário , Neoplasias Nasofaríngeas/imunologia , Adulto , Antígenos de Neoplasias/genética , DNA de Neoplasias/genética , DNA Viral/análise , DNA Viral/genética , Feminino , Antígenos HLA/genética , Antígenos HLA/imunologia , Herpesvirus Humano 4/genética , Humanos , Mutação INDEL , Inibidores de Checkpoint Imunológico/uso terapêutico , Estimativa de Kaplan-Meier , Metástase Linfática/imunologia , Masculino , Pessoa de Meia-Idade , Carcinoma Nasofaríngeo/imunologia , Carcinoma Nasofaríngeo/terapia , Carcinoma Nasofaríngeo/virologia , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/terapia , Neoplasias Nasofaríngeas/virologia , Polimorfismo de Nucleotídeo Único , Intervalo Livre de Progressão , RNA Neoplásico/genética , Receptores de Antígenos de Linfócitos T/imunologia , Evasão Tumoral , Microambiente Tumoral/imunologia , Infecções Tumorais por Vírus/virologia
17.
Gene ; 790: 145696, 2021 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-33964377

RESUMO

BACKGROUND AND AIMS: Hemorrhagic stroke (HS) results in significant mortality and disability worldwide. Angiotensin Converting Enzyme (ACE) is responsible for blood pressure regulation and vascular homeostasis. Our objective was to conduct a comprehensive meta-analysis for ascertaining the association of ACE I/D polymorphism with HS since a number of studies depicted inconclusive evidence. METHODS: Literature search was performed till July 10, 2020 in PubMed, EMBASE, Cochrane, Chinese National Knowledge Information and Google Scholar databases with keywords: ('Angiotensin Converting Enzyme' OR 'ACE') AND ('Single Nucleotide polymorphisms' OR 'SNP') AND ('Hemorrhagic stroke or 'HS'). Pooled Odds Ratio (OR) and 95% Confidence Interval (CI) were determined for gene-disease association using either fixed (when I2 < 50%) or random effect (when I2 > 50%) models. Risk of bias in studies was assessed using funnel plots and sensitivity analyses. Statistical analysis was performed using STATA version 13.0 software. RESULTS: A total of 53 studies having 5186 HS and 7347 healthy control subjects were included in our meta-analysis. Pooled analyses showed that ACE I/D gene polymorphism had significant association with risk of HS in overall study population [(dominant model: OR = 1.29, 95% CI = 1.12-1.50 & recessive model: OR = 1.79, 95% CI = 1.46-2.20)]. Population subgroup analyses further revealed significant relationship of ACE I/D polymorphism with ICH in Asians (recessive: OR 1.97, 95% CI = 1.57-2.47) but not in Caucasians (recessive: OR 1.02, 95% CI = 0.76-1.36). CONCLUSION: This meta-analysis suggests that ACE I/D polymorphism may lead to risk of HS and can be a potential biomarker for HS susceptibility especially in Asian population.


Assuntos
AVC Hemorrágico/etiologia , Mutação INDEL , Peptidil Dipeptidase A/genética , AVC Hemorrágico/patologia , Humanos
18.
Nat Commun ; 12(1): 2474, 2021 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-33931648

RESUMO

As more clinically-relevant genomic features of myeloid malignancies are revealed, it has become clear that targeted clinical genetic testing is inadequate for risk stratification. Here, we develop and validate a clinical transcriptome-based assay for stratification of acute myeloid leukemia (AML). Comparison of ribonucleic acid sequencing (RNA-Seq) to whole genome and exome sequencing reveals that a standalone RNA-Seq assay offers the greatest diagnostic return, enabling identification of expressed gene fusions, single nucleotide and short insertion/deletion variants, and whole-transcriptome expression information. Expression data from 154 AML patients are used to develop a novel AML prognostic score, which is strongly associated with patient outcomes across 620 patients from three independent cohorts, and 42 patients from a prospective cohort. When combined with molecular risk guidelines, the risk score allows for the re-stratification of 22.1 to 25.3% of AML patients from three independent cohorts into correct risk groups. Within the adverse-risk subgroup, we identify a subset of patients characterized by dysregulated integrin signaling and RUNX1 or TP53 mutation. We show that these patients may benefit from therapy with inhibitors of focal adhesion kinase, encoded by PTK2, demonstrating additional utility of transcriptome-based testing for therapy selection in myeloid malignancy.


Assuntos
Biomarcadores Tumorais/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/metabolismo , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Estudos de Coortes , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Feminino , Fusão Gênica , Humanos , Mutação INDEL , Integrinas/genética , Integrinas/metabolismo , Leucemia Mieloide Aguda/genética , Masculino , Polimorfismo de Nucleotídeo Único , Prognóstico , Estudos Prospectivos , RNA-Seq , Fatores de Risco , Transdução de Sinais/genética , Análise de Sobrevida , Transcriptoma , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Sequenciamento Completo do Exoma , Sequenciamento Completo do Genoma
19.
PLoS Genet ; 17(4): e1009329, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33857147

RESUMO

Nicks are the most frequent form of DNA damage and a potential source of mutagenesis in human cells. By deep sequencing, we have identified factors and pathways that promote and limit mutagenic repair at a targeted nick in human cells. Mutations were distributed asymmetrically around the nick site. BRCA2 inhibited all categories of mutational events, including indels, SNVs and HDR. DNA2 and RPA promoted resection. DNA2 inhibited 1 bp deletions but contributed to longer deletions, as did REV7. POLQ stimulated SNVs. Parallel analysis of DSBs targeted to the same site identified similar roles for DNA2 and POLQ (but not REV7) in promoting deletions and for POLQ in stimulating SNVs. Insertions were infrequent at nicks, and most were 1 bp in length, as at DSBs. The translesion polymerase REV1 stimulated +1 insertions at one nick site but not another, illustrating the potential importance of sequence context in determining the outcome of mutagenic repair. These results highlight the potential for nicks to promote mutagenesis, especially in BRCA-deficient cells, and identify mutagenic signatures of DNA2, REV1, REV3, REV7 and POLQ.


Assuntos
Proteína BRCA2/genética , Quebras de DNA de Cadeia Simples , Dano ao DNA/genética , Mutagênese/genética , Ciclo Celular/genética , Quebras de DNA de Cadeia Dupla , DNA Helicases/genética , Reparo do DNA/genética , Proteínas de Ligação a DNA/genética , DNA Polimerase Dirigida por DNA/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mutação INDEL/genética , Proteínas Mad2/genética , Nucleotidiltransferases/genética , RNA Guia/genética , Transdução de Sinais/genética
20.
Int J Legal Med ; 135(4): 1359-1367, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33907868

RESUMO

Most of insertion/deletion polymorphisms are diallelic molecular markers characterized as small amplicon sizes, high inter-population diversities, and low mutation rates, which make them the promising genetic markers in biogeographic ancestor inference field. The developmental validations of a 39 ancestry informative marker-insertion/deletion (AIM-InDel) panel and the genetic polymorphic investigations of this panel were performed in the Shaanxi Han population of China. The developmental validation included the optimizations of PCR-related indicators, repeatability, reproducibility, precision, accuracy, sensitivity, species specificity, stability of the panel, and the abilities in analyzing degraded, casework, and mixture samples, and the present results demonstrated that this 39 AIM-InDel panel was robust, sensitive, and accurate. For the population diversity analyses, the combined discrimination power value of 38 AIM-InDel loci except for rs36038238 locus was 0.999999999931257, indicating that this novel panel was highly polymorphic, biogeographic informative, and could be also used in individual identifications in the Shaanxi Han population.


Assuntos
Grupo com Ancestrais do Continente Asiático/genética , Genética Forense/instrumentação , Análise de Sequência de DNA/métodos , China/etnologia , Marcadores Genéticos , Humanos , Mutação INDEL , Linhagem , Polimorfismo Genético , Reprodutibilidade dos Testes , Especificidade da Espécie
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