The post-genomic era: epigenetic sequencing applications and data integration
by dr. ir. Maté Ongenaert - Center for Medical Genetics, Ghent University
The past decade is known as the post-genomic era. Ever since the first published human genomes, the pace to determine new genomes ever increased. In addition, a number of new sequencing applications gave access to previously unexplored areas at a genome-wide scale such as whole epigenomes.
In this talk, the data generated from a number of sequencing techniques to determine whole DNA-methylomes and whole genome histone marks will be discussed.
Main goal: to convince scientists that the analysis tools have matured to a level that, using a good manual and insight in the mechanisms behind the analysis, they can do their own basic analyses.
Starting from a raw sequence file, over quality control to mapping to the reference genome, peak calling, visualization and identification of differentially methylated sites: within the time-frame of this talk, the entire process will be demonstrated.
As epigenetics regulates genomic processes and literally is a layer above genetics, able to fine-tune regulatory processes, several layers of information should be look at to understand the underlying mechanisms.
Important aspect in the analysis of epigenetic datasets thus is the integration of several data sources (expression results, re-expression results, DNA-methylation information and histone-modifications).
Overview of epigenetics and its role in diseaseGarry D. Lasaga
Epigenetics is the study of heritable changes in gene expression (active versus inactive genes) that do not involve changes to the underlying DNA sequence — a change in phenotype without a change in genotype — which in turn affects how cells read the genes.
Introduction
Genetics of somatic cell
Somatic cell genetics
Somatic cell nuclear transfer
Somatic cell hybridization
Mapping human genes by using human rodent hybrids
In medical application
Production of monoclonal antibodies by using hybridoma technology
Conclusion
References
Introduction
Transcriptome analysis
Goal of functional genomics
Why we need functional genomics
Technique
1. At DNA level
2.At RNA level
3. At protein level
4. loss of function
5. functional genomic and bioinformatics
Application
Latest research and reviews
Websites of functional genomics
Conclusions
Reference
What is Genome,Genome mapping,types of Genome mapping,linkage or genetic mapping,Physical mapping,Somatic cell hybridization
Radiation hybridization ,Fish( =fluorescence in - situ hybridization),Types of probes for FISH,applications,Molecular markers,Rflp(= Restriction fragment length polymorphism),RFLPs may have the following Applications;Advantages of rflp,disAdvantages of rflp, Rapd(=Random amplification of polymorphic DNA),Process of rapd, Difference between rflp &rapd
"Epigenetics refers to genetic factors that change an organism’s appearance or biological functions without changing the actual DNA sequence. In other words, gene expression changes but the genes themselves don’t. Epigenetics adds an additional level of complexity to the genetic code." - Public Health Cafe
The utility of 18F-fluorocholine PET/CT in the imaging of parathyroid adenomasNukleer Tıp Uzmanı
Introduction:
The aim of the study was to estimate the sensitivity of 18F-FCH PET/CT in preoperative localisation of hyperfunctioning parathyroid glands in patients with primary hyperparathyroidism (PHPT).
Material and methods:
Sixty-five consecutive patients with PHPT, who underwent neck ultrasound (US) and 99mTc/99mTc-MIBI dual-phase parathyroid scintigraphy, were prospectively enrolled. Twenty-two patients had unsuccessful parathyroid surgery prior to the study. PET/CT scans were performed 65.0 ± 13.3 min after injection of 218.5 ± 31.9 MBq of 18F-fluorocholine (FCH). Three experienced nuclear medicine physicians assessed the detection rate of hyperfunctioning parathyroid tissue. Response to parathyroidectomy and clinical follow-up served as a reference test. Per-patient sensitivity and positive predictive value (PPV) were calculated for patients who underwent surgery.
Results:
18F-FCH PET/CT was positive in 61 patients, and negative in 4. US and parathyroid scintigraphy showed positive and negative results in 20, 45 and 17, 48, respectively. US showed nodular goitre in 31 patients and chronic thyroiditis in 9 patients. Parathyroid surgery was performed in 43 (66%) patients. 18F-FCH PET/CT yielded a sensitivity of 100% (95% CI: 87.99–100) and PPV of 85.7% (95% CI: 70.77–94.06). Similar values were observed in patients with chronic thyroiditis, nodular goitre, and patients after an unsuccessful parathyroid surgery. PET/CT identified hyperparathyroidism complications (kidney stones, osteoporotic bone fractures, and brown tumours) in 11 patients.
Conclusions:
18F-FCH PET/CT effectively detected hyperfunctioning parathyroid tissue and its complications. The method showed excellent sensitivity and positive predictive value, including patients with nodular goitre, chronic thyroiditis, and prior unsuccessful parathyroidectomy. PET/CT performance was superior to neck ultrasound and parathyroid scintigraphy.
18F-FCH PET/CT; fluorocholine; hyperparathyroidism; parathyroid imaging
Overview of epigenetics and its role in diseaseGarry D. Lasaga
Epigenetics is the study of heritable changes in gene expression (active versus inactive genes) that do not involve changes to the underlying DNA sequence — a change in phenotype without a change in genotype — which in turn affects how cells read the genes.
Introduction
Genetics of somatic cell
Somatic cell genetics
Somatic cell nuclear transfer
Somatic cell hybridization
Mapping human genes by using human rodent hybrids
In medical application
Production of monoclonal antibodies by using hybridoma technology
Conclusion
References
Introduction
Transcriptome analysis
Goal of functional genomics
Why we need functional genomics
Technique
1. At DNA level
2.At RNA level
3. At protein level
4. loss of function
5. functional genomic and bioinformatics
Application
Latest research and reviews
Websites of functional genomics
Conclusions
Reference
What is Genome,Genome mapping,types of Genome mapping,linkage or genetic mapping,Physical mapping,Somatic cell hybridization
Radiation hybridization ,Fish( =fluorescence in - situ hybridization),Types of probes for FISH,applications,Molecular markers,Rflp(= Restriction fragment length polymorphism),RFLPs may have the following Applications;Advantages of rflp,disAdvantages of rflp, Rapd(=Random amplification of polymorphic DNA),Process of rapd, Difference between rflp &rapd
"Epigenetics refers to genetic factors that change an organism’s appearance or biological functions without changing the actual DNA sequence. In other words, gene expression changes but the genes themselves don’t. Epigenetics adds an additional level of complexity to the genetic code." - Public Health Cafe
The utility of 18F-fluorocholine PET/CT in the imaging of parathyroid adenomasNukleer Tıp Uzmanı
Introduction:
The aim of the study was to estimate the sensitivity of 18F-FCH PET/CT in preoperative localisation of hyperfunctioning parathyroid glands in patients with primary hyperparathyroidism (PHPT).
Material and methods:
Sixty-five consecutive patients with PHPT, who underwent neck ultrasound (US) and 99mTc/99mTc-MIBI dual-phase parathyroid scintigraphy, were prospectively enrolled. Twenty-two patients had unsuccessful parathyroid surgery prior to the study. PET/CT scans were performed 65.0 ± 13.3 min after injection of 218.5 ± 31.9 MBq of 18F-fluorocholine (FCH). Three experienced nuclear medicine physicians assessed the detection rate of hyperfunctioning parathyroid tissue. Response to parathyroidectomy and clinical follow-up served as a reference test. Per-patient sensitivity and positive predictive value (PPV) were calculated for patients who underwent surgery.
Results:
18F-FCH PET/CT was positive in 61 patients, and negative in 4. US and parathyroid scintigraphy showed positive and negative results in 20, 45 and 17, 48, respectively. US showed nodular goitre in 31 patients and chronic thyroiditis in 9 patients. Parathyroid surgery was performed in 43 (66%) patients. 18F-FCH PET/CT yielded a sensitivity of 100% (95% CI: 87.99–100) and PPV of 85.7% (95% CI: 70.77–94.06). Similar values were observed in patients with chronic thyroiditis, nodular goitre, and patients after an unsuccessful parathyroid surgery. PET/CT identified hyperparathyroidism complications (kidney stones, osteoporotic bone fractures, and brown tumours) in 11 patients.
Conclusions:
18F-FCH PET/CT effectively detected hyperfunctioning parathyroid tissue and its complications. The method showed excellent sensitivity and positive predictive value, including patients with nodular goitre, chronic thyroiditis, and prior unsuccessful parathyroidectomy. PET/CT performance was superior to neck ultrasound and parathyroid scintigraphy.
18F-FCH PET/CT; fluorocholine; hyperparathyroidism; parathyroid imaging
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The post-genomic era: epigenetic sequencing applications and data integration
1. The post-genomic era Epigenetic sequencing applications and data integration WOUD mini-symposium28/09/2011 Maté Ongenaert Center for Medical Genetics Ghent University Hospital, Belgium
2.
3. Epigenetics > Introduction -genetics Heritable changes to the DNA or histones without affecting the DNA sequence A whole range of changes are described DNA-methylation Histonetailmodifications Methylation Acetylation Phosphorylation …. Epigenetic changes are interconnected
13. Epigenetics > Detection / Prognosis / Prediction Prediction Chemotherapy respons (MGMT in brain cancer - temozolomide)
14.
15.
16. Sequencing the epigenome Shearing of DNA (Covaris) Sequencing Control of fragment sizeswith high sensitivity DNA chips Concentration determination of the fragmented DNA with Fluostar Optima plate reader MBD2 immunoprecipitation reaction (MethylCollector Kit)
22. Sequencing the epigenome Mapping macs14 -t IMR32.bowtie -f BOWTIE -g hs -n IMR32 -w --single-wig Input: mapped “treatment” reads and format of mapping (you can also provide a control sample) Parameters:-g hs: human reference genome (for size estimation)- n: name of output files - w: create wig-files for visualisation (counts)
29. Conclusions Sequencingepigenomesreveals a wealth of information There is no suchthing as the epigenome Methylome Hydroxymethylome Different histonemodifications Don’tforget the interplayand the dynamics… Start exploring the data byyourselfas youknow the application the best
![Overview ,[object Object],Introduction DNA-methylation Histonemodifications The interplaybetweenmethylationandhistonemodifications Applications of epigentics Epigeneticsequencing Sequencingthe epigenome Data analysis andintegration](data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7)