scholarly journals Genome-wide characterization of copy number variations in the host genome in genetic resistance to Marek's disease using next generation sequencing

2020 ◽  
Author(s):  
Hao Bai ◽  
Yanghua He ◽  
Yi Ding ◽  
Huanmin Zhang ◽  
Jilan Chen ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Copy Number ◽  
Marek's Disease ◽  
Neoplastic Disease ◽  
Marek’S Disease ◽  
Next Generation ◽  
Qrt Pcr ◽  
Genome Wide ◽  
A Genome ◽  
Generation Sequencing

Abstract Background: Marek’s disease (MD) is a highly neoplastic disease primarily affecting chickens, and remains as a chronic infectious disease that threatens the poultry industry. Copy number variation (CNV) has been examined in many species and is recognized as a major source of genetic variation that directly contributes to phenotypic variation such as resistance to infectious diseases. Two highly inbred chicken lines 63 (MD-resistant) and 72 (MD-susceptible), as well as their F1 generation and six recombinant congenic strains (RCSs) with varied susceptibility to MD, are considered as ideal models to identify the complex mechanisms of genetic and molecular resistance to MD.Results: In the present study, to unravel the potential genetic mechanisms underlying resistance to MD, we performed a genome-wide CNV detection using next generation sequencing on the inbred chicken lines with the assistance of CNVnator. As a result, a total of 1,649 CNV regions (CNVRs) were successfully identified after merging all the nine datasets, of which 90 CNVRs were overlapped across all the chicken lines. Within these shared regions, 1,360 harbored genes were identified. In addition, 55 and 44 CNVRs with 62 and 57 harbored genes were specifically identified in line 63 and 72, respectively. Bioinformatics analysis showed that the nearby genes were significantly enriched in 36 GO terms and 6 KEGG pathways including JAK/STAT signaling pathway. Ten CNVRs (nine deletions and one duplication) involved in 10 disease-related genes were selected for validation by using qRT-PCR, all of which were successfully confirmed. Finally, qRT-PCR was also used to validate two deletion events in line 72 that were definitely normal in line 63. One high-confidence gene, IRF2 was identified as the most promising candidate gene underlying resistance and susceptibility to MD in view of its function and overlaps with data from previous study.Conclusions: Our findings provide valuable insights for understanding the genetic mechanism of resistance to MD and the identified gene and pathway could be considered as the subject of further functional characterization.

scholarly journals Genome-wide characterization of copy number variations in genetic resistance to Marek’s disease using next generation sequencing

2019 ◽  
Author(s):  
Hao Bai ◽  
Yanghua He ◽  
Yi Ding ◽  
Huanmin Zhang ◽  
Jilan Chen ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Copy Number ◽  
Marek's Disease ◽  
Neoplastic Disease ◽  
Marek’S Disease ◽  
Next Generation ◽  
Qrt Pcr ◽  
Genome Wide ◽  
A Genome ◽  
Generation Sequencing

Abstract Background: Marek’s disease (MD) is a highly neoplastic disease primarily affecting chickens, and remains as a chronic infectious disease that threatens the poultry industry. Copy number variation (CNV) has been examined in many species and is recognized as a major source of genetic variation that directly contributes to phenotypic variation such as resistance to infectious diseases. Two highly inbred chicken lines 63 (MD-resistant) and 72 (MD-susceptible), as well as their F1 generation and six recombinant congenic strains (RCSs) with varied susceptibility to MD, are considered as ideal models to identify the complex mechanisms of genetic and molecular resistance to MD. Results: In the present study, to unravel the potential genetic mechanisms underlying MD, we performed a genome-wide CNV detection using next generation sequencing on the inbred chicken lines with the assistance of CNVnator. As a result, a total of 1,649 CNV regions (CNVRs) were successfully identified after merging all the nine datasets, of which 90 CNVRs were overlapped across all the chicken lines. Within these shared regions, 1,360 harbored genes were identified. In addition, 55 and 44 CNVRs with 62 and 57 harbored genes were specifically identified in line 63 and 72, respectively. Bioinformatics analysis showed that the nearby genes were significantly enriched in 36 GO terms and 6 KEGG pathways including JAK/STAT signaling pathway. Ten CNVRs (nine deletions and one duplication) involved in 10 disease-related genes were selected for validation by using qRT-PCR, all of which were successfully confirmed. Finally, qRT-PCR was also used to validate two deletion events in line 72 that were definitely normal in line 63. One high-confidence gene, IRF2 was identified as the most promising candidate gene underlying MD in view of its function and overlaps with previous study. Conclusions: Our findings provide valuable insights for understanding the genetic mechanism of resistance to MD and the identified gene and pathway could be considered as the subject of further functional characterization.

scholarly journals Identification of a genome-wide serum microRNA expression profile as potential noninvasive biomarkers for chronic kidney disease using next-generation sequencing

2020 ◽  
Vol 48 (12) ◽  
pp. 030006052096948
Author(s):  
Xinying Liu ◽  
Weijie Wang ◽  
Yaling Bai ◽  
Huiran Zhang ◽  
Shenglei Zhang ◽  
...  
Keyword(s):  
Chronic Kidney Disease ◽  
Next Generation Sequencing ◽  
Kidney Disease ◽  
Expression Profile ◽  
Fold Change ◽  
Next Generation ◽  
Qrt Pcr ◽  
Genome Wide ◽  
A Genome ◽  
Generation Sequencing

Objective To identify serum microRNAs (miRNAs) as potential non-invasive biomarkers for patients with chronic kidney disease (CKD). Methods We collected serum samples from healthy controls, CKD stage 1 (CKD1), and stage 5 (CKD5) patients with primary glomerulonephritis (GN), screened differentially expressed miRNAs (DEMs) using next-generation sequencing (NGS), and confirmed the sequencing data using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Results We identified 20 and 42 DEMs in the CKD1 and CKD5 patients compared with the controls, respectively, and 70 DEMs in the CKD5 compared with the CKD1 patients. The qRT-PCR results showed that miR-483-5p was up-regulated in the CKD1 and CKD5 patients compared with controls (fold change = 2.56 and 18.77, respectively). miR-363-3p was down-regulated in the CKD5 patients compared with the controls and CKD1 patients (fold change = 0.27 and 0.48, respectively). Conclusion We identified a genome-wide serum miRNA expression profile in CKD patients, and serum miR-483-5p and miR-363-3p may act as potential diagnostic biomarkers for CKD.

Next generation sequencing allows deeper analysis and understanding of genomes and transcriptomes including aspects to fertility

2011 ◽  
Vol 23 (1) ◽  
pp. 75 ◽  
Author(s):  
Thomas Werner
Keyword(s):  
Next Generation Sequencing ◽  
Transcriptional Control ◽  
Target Genes ◽  
De Novo ◽  
Alternative Promoters ◽  
Next Generation ◽  
Sequencing Data ◽  
Genome Wide ◽  
A Genome ◽  
Generation Sequencing

Reproduction and fertility are controlled by specific events naturally linked to oocytes, testes and early embryonal tissues. A significant part of these events involves gene expression, especially transcriptional control and alternative transcription (alternative promoters and alternative splicing). While methods to analyse such events for carefully predetermined target genes are well established, until recently no methodology existed to extend such analyses into a genome-wide de novo discovery process. With the arrival of next generation sequencing (NGS) it becomes possible to attempt genome-wide discovery in genomic sequences as well as whole transcriptomes at a single nucleotide level. This does not only allow identification of the primary changes (e.g. alternative transcripts) but also helps to elucidate the regulatory context that leads to the induction of transcriptional changes. This review discusses the basics of the new technological and scientific concepts arising from NGS, prominent differences from microarray-based approaches and several aspects of its application to reproduction and fertility research. These concepts will then be illustrated in an application example of NGS sequencing data analysis involving postimplantation endometrium tissue from cows.

scholarly journals ITR-Seq, a next-generation sequencing assay, identifies genome-wide DNA editing sites in vivo following adeno-associated viral vector-mediated genome editing

BMC Genomics ◽  
2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Camilo Breton ◽  
Peter M. Clark ◽  
Lili Wang ◽  
Jenny A. Greig ◽  
James M. Wilson
Keyword(s):  
Next Generation Sequencing ◽  
Genome Editing ◽  
Next Generation ◽  
Aav Vector ◽  
Genome Wide ◽  
A Genome ◽  
Vector Sequences ◽  
Target Activity ◽  
Generation Sequencing

Abstract Background Identifying nuclease-induced double-stranded breaks in DNA on a genome-wide scale is critical for assessing the safety and efficacy of genome editing therapies. We previously demonstrated that after administering adeno-associated viral (AAV) vector-mediated genome-editing strategies in vivo, vector sequences integrated into the host organism’s genomic DNA at double-stranded breaks. Thus, identifying the genomic location of inserted AAV sequences would enable us to identify DSB events, mainly derived from the nuclease on- and off-target activity. Results Here, we developed a next-generation sequencing assay that detects insertions of specific AAV vector sequences called inverted terminal repeats (ITRs). This assay, ITR-Seq, enables us to identify off-target nuclease activity in vivo. Using ITR-Seq, we analyzed liver DNA samples of rhesus macaques treated with AAV vectors expressing a meganuclease. We found dose-dependent off-target activity and reductions in off-target events induced by further meganuclease development. In mice, we identified the genomic locations of ITR integration after treatment with Cas9 nucleases and their corresponding single-guide RNAs. Conclusions In sum, ITR-Seq is a powerful method for identifying off-target sequences induced by AAV vector-delivered genome-editing nucleases. ITR-Seq will help us understand the specificity and efficacy of different genome-editing nucleases in animal models and clinical studies. This information can help enhance the safety profile of gene-editing therapies.

scholarly journals Application of Next-Generation Sequencing for the Genomic Characterization of Patients with Smoldering Myeloma

Cancers ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 1332 ◽  
Author(s):  
Martina Manzoni ◽  
Valentina Marchica ◽  
Paola Storti ◽  
Bachisio Ziccheddu ◽  
Gabriella Sammarelli ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Gene Mutations ◽  
Genomic Analysis ◽  
Next Generation ◽  
Genome Wide ◽  
A Genome ◽  
Low Pass ◽  
Genomic Characterization ◽  
Smoldering Myeloma ◽  
Generation Sequencing

Genomic analysis could contribute to a better understanding of the biological determinants of the evolution of multiple myeloma (MM) precursor disease and an improved definition of high-risk patients. To assess the feasibility and value of next-generation sequencing approaches in an asymptomatic setting, we performed a targeted gene mutation analysis and a genome-wide assessment of copy number alterations (CNAs) by ultra-low-pass whole genome sequencing (ULP-WGS) in six patients with monoclonal gammopathy of undetermined significance and 25 patients with smoldering MM (SMM). Our comprehensive genomic characterization highlighted heterogeneous but substantial values of the tumor fraction, especially in SMM; a rather high degree of genomic complexity, in terms of both mutations and CNAs, and inter-patient variability; a higher incidence of gene mutations and CNAs in SMM, confirming ongoing evolution; intraclonal heterogeneity; and instances of convergent evolution. ULP-WGS of these patients proved effective in revealing the marked genome-wide level of their CNAs, most of which are not routinely investigated. Finally, the analysis of our small SMM cohort suggested that chr(8p) deletions, the DNA tumor fraction, and the number of alterations may have clinical relevance in the progression to overt MM. Although validation in larger series is mandatory, these findings highlight the promising impact of genomic approaches in the clinical management of SMM.

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