In this webinar, we will take a look at a large-scale SNP-based forensic identification panel for DNA analysis with massively parallel sequencing (MPS). The panel was specifically designed for the challenges of identifying missing persons; where DNA is frequently highly degraded, and relationship tests may involve reference samples from across several generations and in a deficient pedigree.
ICMP MPS SNP Panel for Missing Persons - Michelle Peck et al.QIAGEN
Optimization and Performance of a Very Large MGS SNP Panel for Missing Persons, by Michelle Peck et al., International Commission on Mission Persons. Presented May 3, 2018, at the QIAGEN Investigator Forum, San Antonio, TX.
An Approach to De-convolution of Mixtures in Touch DNA Samples. Download now!QIAGEN
7th QIAGEN Investigator Forum - Lisbon, March 8, 2018 . An Approach to De-convolution of Mixtures in Touch DNA Samples. Presenter: Lisa Dierig, Institute of Legal Medicine, Ulm
Validation of Identity and Ancestry
SNP Panels for the Ion PGM™
Christopher Phillips, Carla Santos, Maria de la Puente,
Manuel Fondevila, Ángel Carracedo, Maviky Lareu
Forensic Genetics Unit,
University of Santiago de Compostela
FORENSIC DNA PROFILING: Strengths and LimitationsHezekiah Fatoki
Forensic science is defined as the application of scientific knowledge and experimentation to legal contentions, be they civil or criminal matters. DNA profiling (also called DNA typing or DNA fingerprinting) is a forensic techniques used to identify individuals by characteristics of their DNA in crime cases. DNA profiling can be use to resolve paternal and ancestral issues. This process was built mainly on the knowledge of two scientific breakthroughs. First is the Polymerase Chain Reaction (PCR) which was conceived by Kary Mullis in 1983 at Cetus Corporation, USA. Second is the Restriction Fragment Length Polymorphism (RFLP) analysis of repeated DNA sequences which was discovered by Professor Sir Alec Jeffreys in 1985 at the University of Leicester, UK. The strengths and limitations of the current and emerging forensic DNA profiling are the focus of this seminar. It is my expectation that the newly proposed synthetic human genome project will aid the strength of this process in the future.
Molecular QC: Using Reference Standards in NGS PipelinesCandy Smellie
Since its inception, next-generation sequencing has found utility in a diverse set of industries, from biomarker discovery in pharma to ancestral identification in archeology. Across the board, NGS has the advantage of allowing us to answer questions that require a lot of data. Next-generation sequencing provides orders of magnitude more data than traditional Sanger sequencing as hundreds of “lanes” analyzed in parallel vs. hundreds of millions of “clusters” which allows for many samples to be multiplexed on a single-run.
By starting with different genetic material and following specific experimental workflows, NGS can be applied to many applications.
Here we focus on DNA resequencing applications, which implies the data generated will be compared to an existing reference sequence (such as the human genome). Specifically, we’ll focus on how we can analyze patient-derived material to identify onco-relevant mutations including single-nucleotide variants, insertions-deletions, copy number variants and translocations. We’ll also focus on how known reference standards have been shown to be vital in ensuring data generated from NGS assays is accurate and reproducible.
ICMP MPS SNP Panel for Missing Persons - Michelle Peck et al.QIAGEN
Optimization and Performance of a Very Large MGS SNP Panel for Missing Persons, by Michelle Peck et al., International Commission on Mission Persons. Presented May 3, 2018, at the QIAGEN Investigator Forum, San Antonio, TX.
An Approach to De-convolution of Mixtures in Touch DNA Samples. Download now!QIAGEN
7th QIAGEN Investigator Forum - Lisbon, March 8, 2018 . An Approach to De-convolution of Mixtures in Touch DNA Samples. Presenter: Lisa Dierig, Institute of Legal Medicine, Ulm
Validation of Identity and Ancestry
SNP Panels for the Ion PGM™
Christopher Phillips, Carla Santos, Maria de la Puente,
Manuel Fondevila, Ángel Carracedo, Maviky Lareu
Forensic Genetics Unit,
University of Santiago de Compostela
FORENSIC DNA PROFILING: Strengths and LimitationsHezekiah Fatoki
Forensic science is defined as the application of scientific knowledge and experimentation to legal contentions, be they civil or criminal matters. DNA profiling (also called DNA typing or DNA fingerprinting) is a forensic techniques used to identify individuals by characteristics of their DNA in crime cases. DNA profiling can be use to resolve paternal and ancestral issues. This process was built mainly on the knowledge of two scientific breakthroughs. First is the Polymerase Chain Reaction (PCR) which was conceived by Kary Mullis in 1983 at Cetus Corporation, USA. Second is the Restriction Fragment Length Polymorphism (RFLP) analysis of repeated DNA sequences which was discovered by Professor Sir Alec Jeffreys in 1985 at the University of Leicester, UK. The strengths and limitations of the current and emerging forensic DNA profiling are the focus of this seminar. It is my expectation that the newly proposed synthetic human genome project will aid the strength of this process in the future.
Molecular QC: Using Reference Standards in NGS PipelinesCandy Smellie
Since its inception, next-generation sequencing has found utility in a diverse set of industries, from biomarker discovery in pharma to ancestral identification in archeology. Across the board, NGS has the advantage of allowing us to answer questions that require a lot of data. Next-generation sequencing provides orders of magnitude more data than traditional Sanger sequencing as hundreds of “lanes” analyzed in parallel vs. hundreds of millions of “clusters” which allows for many samples to be multiplexed on a single-run.
By starting with different genetic material and following specific experimental workflows, NGS can be applied to many applications.
Here we focus on DNA resequencing applications, which implies the data generated will be compared to an existing reference sequence (such as the human genome). Specifically, we’ll focus on how we can analyze patient-derived material to identify onco-relevant mutations including single-nucleotide variants, insertions-deletions, copy number variants and translocations. We’ll also focus on how known reference standards have been shown to be vital in ensuring data generated from NGS assays is accurate and reproducible.
Molecular QC: Interpreting your Bioinformatics PipelineCandy Smellie
What is the impact of assay failure in your laboratory and how do you monitor for it?
The most heavily degraded samples are not suitable for standard exome coverage: sometimes it’s not even a matter of getting bad sequencing, you might get nothing at all!
FFPE artifacts increase with storage time
Artifacts go against the statistical power of your variant calling analysis
Molecular reference standards help filter out bad mappings and spurious variants
Bioinformatics pipelines allow adding Molecular Reference Standards in your joint variant calling pipeline
Genome In A Bottle Reference Standards are invaluable for validating variant calling analysis
NIST and its collaborators shared datasets created with most NGS technologies
Horizon Diagnostics shared annotated, merged variant calls from NIST for the Ashkenazim Trio
~35K variants are predicted having high or moderate impact within the Trio
GM24385 (Ashkenazim Son) includes 352 small variants with high/moderate impact which are absent in Father and Mother
Routinely monitor the performance of your workflows and assays with independent external controls
Understanding and controlling for sample and platform biases in NGS assaysCandy Smellie
What is the impact of assay failure in your laboratory and how do you monitor for it?
The advancement of next-generation sequencing has provided invaluable resources to researchers in multiple industries and disciplines, and will be a major driver during the personalized medicine revolution that is upon us. However, while the cost of generating sequencing data continues to decrease this does not take into account the significant costs associated with the infrastructure and expertise that are required to develop a robust, routine NGS pipeline.
Specifically, as predicted by Sboner, et al in 2011, the cost of the sequencing portion of the experiment continues to decrease and the costs associated with upfront experimental design and downstream analysis dominate the cost of each assay. This is true whether you are performing a pre-clinical R&D project, and perhaps even more so for clinical assays. In the paper, the authors note the unpredictable and considerable ‘human time’ spent on the upstream design and downstream analysis. Here at Horizon, we aim to develop tools that help researchers and clinicians optimize these workflows to make NGS more reliable and ultimately, more affordable by streamlining these resource intensive areas.
Phylogenomic methods for comparative evolutionary biology - University Colleg...Joe Parker
Invited research seminar given to MSc students at University College Dublin on 24th October 2013.
I introduce the discipline of phylogenomics - comparative phylogenetic analyses of DNA sequences across genomes - and some of the applications and recent breakthroughs in the field.
As an in-depth case study I explain the methods and significance of our 2013 Nature paper on adaptive genotypic molecular convergence in echolocating mammals.
I then highlight some of the avenues of study on the frontiers of current research.
Avances en genética. Utilidad de la NGS y la bioinformática.BBK Innova Sarea
27 Octubre 2014. Presentación de Pablo Lapunzina, Director del Instituto de Medicina Genética Médica y Molecular (INGEMM), de IDIPAZ y de CEBERER, en la "Jornada Avances en Genética y Tecnología Social. La experiencia de la Fundación Síndrome de Dravet ".
Forensic Genetics in the 21st century – meeting the challenges of biological ...Thermo Fisher Scientific
"Forensic Genetics in the 21st century – meeting the challenges of biological evidence
Presented by Peter M. Schneider, Institute of Legal Medicine
University of Cologne, Germany Human Identification Solutions Conference – Madrid, Spain
March 4, 2015 CO014206"
Supporting Genomics in the Practice of Medicine by Heidi RehmKnome_Inc
View the webinar at http://www.knome.com/webinar-supporting-genomics-practice-medicine. In this presentation, Dr. Heidi Rehm, Chief Laboratory Director of the Laboratory for Molecular Medicine at Partners Healthcare and one of the Principal Investigators on ClinGen, elucidates the challenges of genomics in medicine and outlined the path to integrating large scale sequencing into clinical practice.
Single-Cell Analysis - Powered by REPLI-g: Single Cell Analysis Series Part 1QIAGEN
What can you do from a single cell? Actually, quite a lot! Beginning with the genome, you can discover new biomarkers by identifying new genetic variances and their association with specific diseases, including cancers. Moving on to RNA, the recent advances in RNA sequencing technology have made single-cell transcriptomics a possibility. Along with these possibilities, come challenges that start from the moment you get the sample to the final step of gaining insights into the cell. This slidedeck will provide an overview on the multiple steps involved as you move from sample acquisition to analysis and data interpretation in different sample types.
Single cell analysis has exploded recently mainly due to the development of high-throughput technologies such as NGS. Single cell analysis is being pursued by researchers in many areas including developmental science, cancer, biomarker discovery and more. This presentation covers some of the recent applications from developed by QIAGEN customers.
Microhaplotype, A Powerful New Type of Genetic MarkerMojgan Talebian
Haplotyped SNPs allow more efficient inference of family relationships on a per locus basis because they constitute multiallelic loci, analogous to the STRs. Haplotypes are optimal type of forensically useful DNA marker, especially family or lineage inference.
A SNPs is a single nucleotide base difference in the DNA sequence. 10 million common SNPs are in the human genome, many of which are already annotated in SNP database. SNPS are Di-allelic and not highly polymorphism .
SNPs are so abundant throughout the genome that it is theoretically possible to type hundreds of them.
SNP’s sample processing and data analysis may be more fully automated because size-based separation is not required.
Molecular QC: Interpreting your Bioinformatics PipelineCandy Smellie
What is the impact of assay failure in your laboratory and how do you monitor for it?
The most heavily degraded samples are not suitable for standard exome coverage: sometimes it’s not even a matter of getting bad sequencing, you might get nothing at all!
FFPE artifacts increase with storage time
Artifacts go against the statistical power of your variant calling analysis
Molecular reference standards help filter out bad mappings and spurious variants
Bioinformatics pipelines allow adding Molecular Reference Standards in your joint variant calling pipeline
Genome In A Bottle Reference Standards are invaluable for validating variant calling analysis
NIST and its collaborators shared datasets created with most NGS technologies
Horizon Diagnostics shared annotated, merged variant calls from NIST for the Ashkenazim Trio
~35K variants are predicted having high or moderate impact within the Trio
GM24385 (Ashkenazim Son) includes 352 small variants with high/moderate impact which are absent in Father and Mother
Routinely monitor the performance of your workflows and assays with independent external controls
Understanding and controlling for sample and platform biases in NGS assaysCandy Smellie
What is the impact of assay failure in your laboratory and how do you monitor for it?
The advancement of next-generation sequencing has provided invaluable resources to researchers in multiple industries and disciplines, and will be a major driver during the personalized medicine revolution that is upon us. However, while the cost of generating sequencing data continues to decrease this does not take into account the significant costs associated with the infrastructure and expertise that are required to develop a robust, routine NGS pipeline.
Specifically, as predicted by Sboner, et al in 2011, the cost of the sequencing portion of the experiment continues to decrease and the costs associated with upfront experimental design and downstream analysis dominate the cost of each assay. This is true whether you are performing a pre-clinical R&D project, and perhaps even more so for clinical assays. In the paper, the authors note the unpredictable and considerable ‘human time’ spent on the upstream design and downstream analysis. Here at Horizon, we aim to develop tools that help researchers and clinicians optimize these workflows to make NGS more reliable and ultimately, more affordable by streamlining these resource intensive areas.
Phylogenomic methods for comparative evolutionary biology - University Colleg...Joe Parker
Invited research seminar given to MSc students at University College Dublin on 24th October 2013.
I introduce the discipline of phylogenomics - comparative phylogenetic analyses of DNA sequences across genomes - and some of the applications and recent breakthroughs in the field.
As an in-depth case study I explain the methods and significance of our 2013 Nature paper on adaptive genotypic molecular convergence in echolocating mammals.
I then highlight some of the avenues of study on the frontiers of current research.
Avances en genética. Utilidad de la NGS y la bioinformática.BBK Innova Sarea
27 Octubre 2014. Presentación de Pablo Lapunzina, Director del Instituto de Medicina Genética Médica y Molecular (INGEMM), de IDIPAZ y de CEBERER, en la "Jornada Avances en Genética y Tecnología Social. La experiencia de la Fundación Síndrome de Dravet ".
Forensic Genetics in the 21st century – meeting the challenges of biological ...Thermo Fisher Scientific
"Forensic Genetics in the 21st century – meeting the challenges of biological evidence
Presented by Peter M. Schneider, Institute of Legal Medicine
University of Cologne, Germany Human Identification Solutions Conference – Madrid, Spain
March 4, 2015 CO014206"
Supporting Genomics in the Practice of Medicine by Heidi RehmKnome_Inc
View the webinar at http://www.knome.com/webinar-supporting-genomics-practice-medicine. In this presentation, Dr. Heidi Rehm, Chief Laboratory Director of the Laboratory for Molecular Medicine at Partners Healthcare and one of the Principal Investigators on ClinGen, elucidates the challenges of genomics in medicine and outlined the path to integrating large scale sequencing into clinical practice.
Single-Cell Analysis - Powered by REPLI-g: Single Cell Analysis Series Part 1QIAGEN
What can you do from a single cell? Actually, quite a lot! Beginning with the genome, you can discover new biomarkers by identifying new genetic variances and their association with specific diseases, including cancers. Moving on to RNA, the recent advances in RNA sequencing technology have made single-cell transcriptomics a possibility. Along with these possibilities, come challenges that start from the moment you get the sample to the final step of gaining insights into the cell. This slidedeck will provide an overview on the multiple steps involved as you move from sample acquisition to analysis and data interpretation in different sample types.
Single cell analysis has exploded recently mainly due to the development of high-throughput technologies such as NGS. Single cell analysis is being pursued by researchers in many areas including developmental science, cancer, biomarker discovery and more. This presentation covers some of the recent applications from developed by QIAGEN customers.
Microhaplotype, A Powerful New Type of Genetic MarkerMojgan Talebian
Haplotyped SNPs allow more efficient inference of family relationships on a per locus basis because they constitute multiallelic loci, analogous to the STRs. Haplotypes are optimal type of forensically useful DNA marker, especially family or lineage inference.
A SNPs is a single nucleotide base difference in the DNA sequence. 10 million common SNPs are in the human genome, many of which are already annotated in SNP database. SNPS are Di-allelic and not highly polymorphism .
SNPs are so abundant throughout the genome that it is theoretically possible to type hundreds of them.
SNP’s sample processing and data analysis may be more fully automated because size-based separation is not required.
Using methylation patterns to determine origin of biological material and ageQIAGEN
In this QIAGEN sponsored webinar, our guest speakers from the San Francisco Police Department (SFPD) Crime Lab and Florida International University (FIU) discuss their research on the potential of epigenetic methylation as a procedure for body fluid identification and age estimation from DNA left at crime scenes. Several approaches have been studied, including an analysis of methyl array data and an initial validation of procedures such as pyrosequencing and real-time PCR. The presentation focuses on a number of tissue-specific epigenetic markers for body fluid and age determination with a promise of future integration of these markers into the forensic lab due to the simplicity of analysis and the ease of application.
Learn more about the Pyrosequencing technology and our solutions at
https://www.qiagen.com/resources/technologies/pyrosequencing-resource-center/
Take lung cancer research to a new molecular dimensionQIAGEN
Circulating Tumor Cells (CTCs) can provide researchers with important new discoveries on the mechanism of cancer. Find out more about the latest technology that provides researchers the necessary tools to conduct CTC research in lung cancer.
Circulating Tumor Cells (CTCs) can provide researchers with important new discoveries on the mechanism of cancer. Find out more about the latest technology that provides researchers the necessary tools to conduct CTC research in AR-V7 related prostate cancer.
Learn about the power of LNA (Locked Nucleic Acid) technology and QIAGEN's LNA enhanced product portfolio for RNA and DNA research. Download the slide deck!
Take your RNA research to the next level with QIAGEN LNA tools!QIAGEN
Download the flyer!
Experience truly exceptional RNA research with QIAGEN's next-generation, LNA®-enhanced tools. LNA (Locked Nucleic Acid) oligos bind with much higher affinity and specificity to RNA targets than standard DNA and RNA oligos – This enables specific and sensitive detection of small RNAs and discrimination between highly similar
sequences.
Assessment of Y chromosome degradation level using the Investigator® Quantipl...QIAGEN
Assessment of Y chromosome degradation level using the Investigator® Quantiplex® Pro RGQ Kit, presented by Dr. Tomasz Kupiec, Head of the Forensic Genetics Section, Institute of Forensic Research, Krakow, Poland on June 14, 2018.
Exploring the Temperate Leaf Microbiome: From Natural Forests to Controlled E...QIAGEN
The aerial surfaces of plants, the phyllosphere, harbors a diverse community of microorganisms. The increasing awareness of the potential roles of phyllosphere microbial communities calls for a greater understanding of their structure and dynamics in natural and urban ecosystems. To do so, we characterized the community structure and assembly dynamics of leaf bacterial communities in natural temperate forests of Quebec by comparing the relative influence of host species identity, site, and time on phyllosphere bacterial community structure. Second, we tested the value of characterizing a tree’s complete phyllosphere microbial community through a single sample by measuring the intra-individual, inter-individual and interspecific variation in leaf bacterial communities. Third, we quantified the relationships among phyllosphere bacterial diversity, plant species richness, plant functional diversity and identity, and plant community productivity in a biodiversity-ecosystem function experiment with trees. Finally, we compared tree leaf bacterial communities in natural and urban environments, as well as along a gradient of increasing anthropogenic pressures. The work presented here thus offers an original assessment of the dynamics at play in the tree phyllosphere.
Cancer Research & the Challenges of FFPE Samples – An IntroductionQIAGEN
A cascade of complex genetic and epigenetic changes regulate tumor formation and progression. Gene expression analyses can shed light on these changes at a molecular level and identify the key genes and associated pathways involved in cancer. Often the samples used in cancer research are FFPE samples, which pose a significant challenge in terms of nucleic acid quality. The quality of nucleic acids extracted from FFPE samples depends on a number of factors, including how the samples were handled before, during and after fixation and embedding.
Dr. Vishwadeepak Tripathi describes the variability of sample purification from FFPE samples – in particular, samples to be used in cancer research. What are the challenges and solutions, and what quality control approach can ensure credible results? This webinar will focus on sample purification and the quality control of FFPE samples and compare different automated purification procedures.
Introduction to real-Time Quantitative PCR (qPCR) - Download the slidesQIAGEN
This slidedeck introduces the concepts of real-time PCR and how to conduct a real-time PCR assay. The topics that are covered include an overview of real-time PCR chemistries, protocols, quantification methods, real-time PCR applications and factors for success.
The Microbiome of Research Animals : Implications for Reproducibility, Transl...QIAGEN
The human gut microbiota (GM) has emerged as a key factor in susceptibility to, as well as a potential biomarker of, several diseases and conditions. Similarly, researchers now appreciate that the GM of laboratory animals could affect the reproducibility and translatability of many disease models, including a complete loss of phenotype. While associations between characteristics of the GM and differential disease model phenotypes are of concern, they can also be viewed as sources of discovery related to disease pathogenesis. As such, there is considerable interest in factors that inadvertently influence the composition of the GM and methods of manipulating the GM prospectively to investigate such associations and standardize or optimize disease models. The webinar will present data on variables capable of influencing the GM of laboratory rodents citing several examples and animal models, considerations related to manipulation of the GM in mice and rats, and recent data supporting the use of “dirty” mice in biomedical research.
Rapid DNA isolation from diverse plant material for use in Next Generation Se...QIAGEN
Isolation of DNA from plant material is often a tedious process which involves significant hands on time and leads to varying results due to the diverse nature of the material. Different parts of the plants as well as the plants themselves differ in both consistency of material and presence of inhibitory substances, making dependable isolation of DNA difficult.
Here, we developed a method for the efficient extraction of DNA from different plant types, including strawberry leaf, pine needle, grape leaf, and cotton and coffee seeds (workflow at right). A novel bead beating method and lysis chemistry led to more efficient sample lysis with minimal hands-on time and significantly increased DNA yield compared to conventional methods. Through the use of multiple technologies to improve removal of secondary metabolites, such as polyphenols, complex polysaccharides, alkaloids and tannins that may inhibit downstream applications, the isolated DNA was of high quality and purity.
The resulting DNA is suitable for immediate use in downstream reactions, including PCR, qPCR and Next Generation Sequencing based applications. Using this method we were further able to design a workflow that included DNA isolation, library preparation and bioinformatics analyses for the efficient detection of plant pathogens isolated from infected samples. With this, our protocol is a substantial improvement within workflows used for plant microbiome and plant pathology studies as well as in plant breeding and engineering.
Rapid extraction of high yield, high quality DNA from tissue samples - Downlo...QIAGEN
Genetic and genomic analysis from tissue samples requires the extraction of high quality DNA. Mechanical disruption methods such as bead milling provide high yield from tissue samples, but cause damage to the nucleic acids. Purely enzymatic methods such as proteinase K digestion can extract nucleic acid without damage, but require long incubation times, often proceeding overnight, and without approaching the yields achieved by mechanical disruption techniques. Thus a method is needed which can provide a rapid extraction of high yield, high quality DNA from tissue samples. See the new method.
Critical Factors for Successful Real-Time PCR: Multiplex PCRQIAGEN
Multiplex end-point PCR is a powerful tool for genotyping and many other applications. QIAGEN’s multiplex PCR chemistry is optimized for reliable amplification of many different templates with high variability in copy numbers. Thus it enables very quick establishment of a new lab routine and instant success for your multiplex PCR strategy.
There is a set of critical factors which we recommend to be regarded for planning and performing this kind of PCR. These will be discussed in detail in the webinar. Additionally, our multiplex PCR chemistry has recently been gaining increasing popularity among scientists who are utilizing it for their next-generation sequencing workflows.
Practical hints and new solutions for successful real-time PCR studies QIAGEN
Part 1: Practical hints and new solutions for successful real-time PCR studies
In this webinar we will cover the following topics which are critical steps for efficient and precise gene expression studies using real-time PCR technology:
- Effect of RNA integrity on real-time PCR results – tips to achieve a true RNA profiling suitable for real-time PCR studies
- Improved methods for cDNA synthesis, optimized for real-time PCR
- Real-time PCR analysis
o Real-time PCR essentials and background information on different quantification strategies
o SYBR Green real-time PCR – factors influencing specificity
o Introduction to probe technology
o New, fast and efficient real-time PCR solutions
Part 2: Critical Factors for Successful Multiplex Real-Time PCR
Multiplex real-time PCR is a powerful tool for gene expression analysis, viral load monitoring, genotyping, and many other applications. The ability to amplify and detect several genomic DNA, cDNA, or RNA targets in the same reaction offers many benefits:
• Conservation of precious samples – more quantification data per sample
• Increased throughput – more targets analyzed per run on a cycler
• Reliable results – no well-to-well variability due to co-amplification of internal control
• Reduced costs – save time and reagents
The QuantiFast Multiplex PCR and RT-PCR kits are optimized for reliable amplification of many different templates despite a high variability in abundance. Thus they enable successful amplification of multiple targets on the first attempt without optimization.
This webinar explains the principles of the QIAGEN multiplex technologies and shows data demonstrating the exceptional multiplex real-time PCR performance of the QuantiFast Multiplex kits.
Overcome the challenges of Nucleic acid isolation from PCR inhibitor-rich mic...QIAGEN
This presentation will focus on nucleic acid extraction tools developed by QIAGEN that facilitate accurate non-biased community analysis and eliminate common amplification problems via the depletion of endogenous polymerase inhibitors using our patented Inhibitor Removal Technology.
RotorGene Q A Rapid, Automatable real-time PCR Instrument for Genotyping and...QIAGEN
QIAGEN has developed a selection of robust, novel chemistries to prevent PCR crosstalk. We can successfully measure target abundance and fold change in real-time assays, and perform sub-genotyping using a fast, high-throughput and powerful High-Resolution Melting (HRM) statistical analysis program. In this presentation, we will demonstrate these features and benefits with examples.
Reproducibility, Quality Control and Importance of AutomationQIAGEN
In this webinar, we will introduce you to the key sample quality parameters, discuss their respective impact on downstream applications and how to monitor them, and present the advantages of automating quality control along complex workflows.
Automated Nucleic Acid Purification from Diverse Sample types using dedicated...QIAGEN
This webinar will focus on the automation of QIAGEN’s new line of DNA and RNA sample prep kits for the microbiome. We will show how automation on the QIAcube enables efficient and reliable use of these samples for sensitive downstream applications such as qPCR and NGS. In addition, you will learn how to successfully use the CLC Microbial Genomics Module for metagenome sequencing and identification of microbial composition and diversity.
Dna Methylation Analysis in a Single Day - Download the SlidesQIAGEN
This webinar introduces the new PyroMark Q48 Autoprep system. Combined with the latest EpiTect Fast bisulfite conversion technology, the new PyroMark Q48 Autoprep can now provide highly automated methylation analysis in a single day.
Simultaneous Isolation of RNA & DNA from one FFPE SampleQIAGEN
Worldwide, there are millions of tissue samples archived in tissue biobanks and biorepositories. These samples are extremely valuable for pharmacological and biomedical research and companion diagnostics, due to the linkage to patient history. The vast majority of archived tissue samples are formalin-fixed and paraffin-embedded (FFPE), since formalin is the standard fixative for tissue samples.
FFPE blocks serve as an excellent source for histomorphology studies, but their use in molecular studies is challenging, due to crosslinking and fragmentation caused by fixation, processing, embedding, and storage conditions. For reliable comparison of genomic and transcriptomic data from heterogeneous samples and to spare sample material, purification of DNA and RNA from
the same sample is essential. This is particularly important when working with tumorous tissues, which contain a heterogeneous distribution of healthy and malignant cells.
Explore natural remedies for syphilis treatment in Singapore. Discover alternative therapies, herbal remedies, and lifestyle changes that may complement conventional treatments. Learn about holistic approaches to managing syphilis symptoms and supporting overall health.
Ethanol (CH3CH2OH), or beverage alcohol, is a two-carbon alcohol
that is rapidly distributed in the body and brain. Ethanol alters many
neurochemical systems and has rewarding and addictive properties. It
is the oldest recreational drug and likely contributes to more morbidity,
mortality, and public health costs than all illicit drugs combined. The
5th edition of the Diagnostic and Statistical Manual of Mental Disorders
(DSM-5) integrates alcohol abuse and alcohol dependence into a single
disorder called alcohol use disorder (AUD), with mild, moderate,
and severe subclassifications (American Psychiatric Association, 2013).
In the DSM-5, all types of substance abuse and dependence have been
combined into a single substance use disorder (SUD) on a continuum
from mild to severe. A diagnosis of AUD requires that at least two of
the 11 DSM-5 behaviors be present within a 12-month period (mild
AUD: 2–3 criteria; moderate AUD: 4–5 criteria; severe AUD: 6–11 criteria).
The four main behavioral effects of AUD are impaired control over
drinking, negative social consequences, risky use, and altered physiological
effects (tolerance, withdrawal). This chapter presents an overview
of the prevalence and harmful consequences of AUD in the U.S.,
the systemic nature of the disease, neurocircuitry and stages of AUD,
comorbidities, fetal alcohol spectrum disorders, genetic risk factors, and
pharmacotherapies for AUD.
New Directions in Targeted Therapeutic Approaches for Older Adults With Mantl...i3 Health
i3 Health is pleased to make the speaker slides from this activity available for use as a non-accredited self-study or teaching resource.
This slide deck presented by Dr. Kami Maddocks, Professor-Clinical in the Division of Hematology and
Associate Division Director for Ambulatory Operations
The Ohio State University Comprehensive Cancer Center, will provide insight into new directions in targeted therapeutic approaches for older adults with mantle cell lymphoma.
STATEMENT OF NEED
Mantle cell lymphoma (MCL) is a rare, aggressive B-cell non-Hodgkin lymphoma (NHL) accounting for 5% to 7% of all lymphomas. Its prognosis ranges from indolent disease that does not require treatment for years to very aggressive disease, which is associated with poor survival (Silkenstedt et al, 2021). Typically, MCL is diagnosed at advanced stage and in older patients who cannot tolerate intensive therapy (NCCN, 2022). Although recent advances have slightly increased remission rates, recurrence and relapse remain very common, leading to a median overall survival between 3 and 6 years (LLS, 2021). Though there are several effective options, progress is still needed towards establishing an accepted frontline approach for MCL (Castellino et al, 2022). Treatment selection and management of MCL are complicated by the heterogeneity of prognosis, advanced age and comorbidities of patients, and lack of an established standard approach for treatment, making it vital that clinicians be familiar with the latest research and advances in this area. In this activity chaired by Michael Wang, MD, Professor in the Department of Lymphoma & Myeloma at MD Anderson Cancer Center, expert faculty will discuss prognostic factors informing treatment, the promising results of recent trials in new therapeutic approaches, and the implications of treatment resistance in therapeutic selection for MCL.
Target Audience
Hematology/oncology fellows, attending faculty, and other health care professionals involved in the treatment of patients with mantle cell lymphoma (MCL).
Learning Objectives
1.) Identify clinical and biological prognostic factors that can guide treatment decision making for older adults with MCL
2.) Evaluate emerging data on targeted therapeutic approaches for treatment-naive and relapsed/refractory MCL and their applicability to older adults
3.) Assess mechanisms of resistance to targeted therapies for MCL and their implications for treatment selection
Lung Cancer: Artificial Intelligence, Synergetics, Complex System Analysis, S...Oleg Kshivets
RESULTS: Overall life span (LS) was 2252.1±1742.5 days and cumulative 5-year survival (5YS) reached 73.2%, 10 years – 64.8%, 20 years – 42.5%. 513 LCP lived more than 5 years (LS=3124.6±1525.6 days), 148 LCP – more than 10 years (LS=5054.4±1504.1 days).199 LCP died because of LC (LS=562.7±374.5 days). 5YS of LCP after bi/lobectomies was significantly superior in comparison with LCP after pneumonectomies (78.1% vs.63.7%, P=0.00001 by log-rank test). AT significantly improved 5YS (66.3% vs. 34.8%) (P=0.00000 by log-rank test) only for LCP with N1-2. Cox modeling displayed that 5YS of LCP significantly depended on: phase transition (PT) early-invasive LC in terms of synergetics, PT N0—N12, cell ratio factors (ratio between cancer cells- CC and blood cells subpopulations), G1-3, histology, glucose, AT, blood cell circuit, prothrombin index, heparin tolerance, recalcification time (P=0.000-0.038). Neural networks, genetic algorithm selection and bootstrap simulation revealed relationships between 5YS and PT early-invasive LC (rank=1), PT N0—N12 (rank=2), thrombocytes/CC (3), erythrocytes/CC (4), eosinophils/CC (5), healthy cells/CC (6), lymphocytes/CC (7), segmented neutrophils/CC (8), stick neutrophils/CC (9), monocytes/CC (10); leucocytes/CC (11). Correct prediction of 5YS was 100% by neural networks computing (area under ROC curve=1.0; error=0.0).
CONCLUSIONS: 5YS of LCP after radical procedures significantly depended on: 1) PT early-invasive cancer; 2) PT N0--N12; 3) cell ratio factors; 4) blood cell circuit; 5) biochemical factors; 6) hemostasis system; 7) AT; 8) LC characteristics; 9) LC cell dynamics; 10) surgery type: lobectomy/pneumonectomy; 11) anthropometric data. Optimal diagnosis and treatment strategies for LC are: 1) screening and early detection of LC; 2) availability of experienced thoracic surgeons because of complexity of radical procedures; 3) aggressive en block surgery and adequate lymph node dissection for completeness; 4) precise prediction; 5) adjuvant chemoimmunoradiotherapy for LCP with unfavorable prognosis.
Flu Vaccine Alert in Bangalore Karnatakaaddon Scans
As flu season approaches, health officials in Bangalore, Karnataka, are urging residents to get their flu vaccinations. The seasonal flu, while common, can lead to severe health complications, particularly for vulnerable populations such as young children, the elderly, and those with underlying health conditions.
Dr. Vidisha Kumari, a leading epidemiologist in Bangalore, emphasizes the importance of getting vaccinated. "The flu vaccine is our best defense against the influenza virus. It not only protects individuals but also helps prevent the spread of the virus in our communities," he says.
This year, the flu season is expected to coincide with a potential increase in other respiratory illnesses. The Karnataka Health Department has launched an awareness campaign highlighting the significance of flu vaccinations. They have set up multiple vaccination centers across Bangalore, making it convenient for residents to receive their shots.
To encourage widespread vaccination, the government is also collaborating with local schools, workplaces, and community centers to facilitate vaccination drives. Special attention is being given to ensuring that the vaccine is accessible to all, including marginalized communities who may have limited access to healthcare.
Residents are reminded that the flu vaccine is safe and effective. Common side effects are mild and may include soreness at the injection site, mild fever, or muscle aches. These side effects are generally short-lived and far less severe than the flu itself.
Healthcare providers are also stressing the importance of continuing COVID-19 precautions. Wearing masks, practicing good hand hygiene, and maintaining social distancing are still crucial, especially in crowded places.
Protect yourself and your loved ones by getting vaccinated. Together, we can help keep Bangalore healthy and safe this flu season. For more information on vaccination centers and schedules, residents can visit the Karnataka Health Department’s official website or follow their social media pages.
Stay informed, stay safe, and get your flu shot today!
Pulmonary Thromboembolism - etilogy, types, medical- Surgical and nursing man...VarunMahajani
Disruption of blood supply to lung alveoli due to blockage of one or more pulmonary blood vessels is called as Pulmonary thromboembolism. In this presentation we will discuss its causes, types and its management in depth.
Title: Sense of Taste
Presenter: Dr. Faiza, Assistant Professor of Physiology
Qualifications:
MBBS (Best Graduate, AIMC Lahore)
FCPS Physiology
ICMT, CHPE, DHPE (STMU)
MPH (GC University, Faisalabad)
MBA (Virtual University of Pakistan)
Learning Objectives:
Describe the structure and function of taste buds.
Describe the relationship between the taste threshold and taste index of common substances.
Explain the chemical basis and signal transduction of taste perception for each type of primary taste sensation.
Recognize different abnormalities of taste perception and their causes.
Key Topics:
Significance of Taste Sensation:
Differentiation between pleasant and harmful food
Influence on behavior
Selection of food based on metabolic needs
Receptors of Taste:
Taste buds on the tongue
Influence of sense of smell, texture of food, and pain stimulation (e.g., by pepper)
Primary and Secondary Taste Sensations:
Primary taste sensations: Sweet, Sour, Salty, Bitter, Umami
Chemical basis and signal transduction mechanisms for each taste
Taste Threshold and Index:
Taste threshold values for Sweet (sucrose), Salty (NaCl), Sour (HCl), and Bitter (Quinine)
Taste index relationship: Inversely proportional to taste threshold
Taste Blindness:
Inability to taste certain substances, particularly thiourea compounds
Example: Phenylthiocarbamide
Structure and Function of Taste Buds:
Composition: Epithelial cells, Sustentacular/Supporting cells, Taste cells, Basal cells
Features: Taste pores, Taste hairs/microvilli, and Taste nerve fibers
Location of Taste Buds:
Found in papillae of the tongue (Fungiform, Circumvallate, Foliate)
Also present on the palate, tonsillar pillars, epiglottis, and proximal esophagus
Mechanism of Taste Stimulation:
Interaction of taste substances with receptors on microvilli
Signal transduction pathways for Umami, Sweet, Bitter, Sour, and Salty tastes
Taste Sensitivity and Adaptation:
Decrease in sensitivity with age
Rapid adaptation of taste sensation
Role of Saliva in Taste:
Dissolution of tastants to reach receptors
Washing away the stimulus
Taste Preferences and Aversions:
Mechanisms behind taste preference and aversion
Influence of receptors and neural pathways
Impact of Sensory Nerve Damage:
Degeneration of taste buds if the sensory nerve fiber is cut
Abnormalities of Taste Detection:
Conditions: Ageusia, Hypogeusia, Dysgeusia (parageusia)
Causes: Nerve damage, neurological disorders, infections, poor oral hygiene, adverse drug effects, deficiencies, aging, tobacco use, altered neurotransmitter levels
Neurotransmitters and Taste Threshold:
Effects of serotonin (5-HT) and norepinephrine (NE) on taste sensitivity
Supertasters:
25% of the population with heightened sensitivity to taste, especially bitterness
Increased number of fungiform papillae
- Video recording of this lecture in English language: https://youtu.be/lK81BzxMqdo
- Video recording of this lecture in Arabic language: https://youtu.be/Ve4P0COk9OI
- Link to download the book free: https://nephrotube.blogspot.com/p/nephrotube-nephrology-books.html
- Link to NephroTube website: www.NephroTube.com
- Link to NephroTube social media accounts: https://nephrotube.blogspot.com/p/join-nephrotube-on-social-media.html
ARTIFICIAL INTELLIGENCE IN HEALTHCARE.pdfAnujkumaranit
Artificial intelligence (AI) refers to the simulation of human intelligence processes by machines, especially computer systems. It encompasses tasks such as learning, reasoning, problem-solving, perception, and language understanding. AI technologies are revolutionizing various fields, from healthcare to finance, by enabling machines to perform tasks that typically require human intelligence.
Tom Selleck Health: A Comprehensive Look at the Iconic Actor’s Wellness Journeygreendigital
Tom Selleck, an enduring figure in Hollywood. has captivated audiences for decades with his rugged charm, iconic moustache. and memorable roles in television and film. From his breakout role as Thomas Magnum in Magnum P.I. to his current portrayal of Frank Reagan in Blue Bloods. Selleck's career has spanned over 50 years. But beyond his professional achievements. fans have often been curious about Tom Selleck Health. especially as he has aged in the public eye.
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Introduction
Many have been interested in Tom Selleck health. not only because of his enduring presence on screen but also because of the challenges. and lifestyle choices he has faced and made over the years. This article delves into the various aspects of Tom Selleck health. exploring his fitness regimen, diet, mental health. and the challenges he has encountered as he ages. We'll look at how he maintains his well-being. the health issues he has faced, and his approach to ageing .
Early Life and Career
Childhood and Athletic Beginnings
Tom Selleck was born on January 29, 1945, in Detroit, Michigan, and grew up in Sherman Oaks, California. From an early age, he was involved in sports, particularly basketball. which played a significant role in his physical development. His athletic pursuits continued into college. where he attended the University of Southern California (USC) on a basketball scholarship. This early involvement in sports laid a strong foundation for his physical health and disciplined lifestyle.
Transition to Acting
Selleck's transition from an athlete to an actor came with its physical demands. His first significant role in "Magnum P.I." required him to perform various stunts and maintain a fit appearance. This role, which he played from 1980 to 1988. necessitated a rigorous fitness routine to meet the show's demands. setting the stage for his long-term commitment to health and wellness.
Fitness Regimen
Workout Routine
Tom Selleck health and fitness regimen has evolved. adapting to his changing roles and age. During his "Magnum, P.I." days. Selleck's workouts were intense and focused on building and maintaining muscle mass. His routine included weightlifting, cardiovascular exercises. and specific training for the stunts he performed on the show.
Selleck adjusted his fitness routine as he aged to suit his body's needs. Today, his workouts focus on maintaining flexibility, strength, and cardiovascular health. He incorporates low-impact exercises such as swimming, walking, and light weightlifting. This balanced approach helps him stay fit without putting undue strain on his joints and muscles.
Importance of Flexibility and Mobility
In recent years, Selleck has emphasized the importance of flexibility and mobility in his fitness regimen. Understanding the natural decline in muscle mass and joint flexibility with age. he includes stretching and yoga in his routine. These practices help prevent injuries, improve posture, and maintain mobilit
Report Back from SGO 2024: What’s the Latest in Cervical Cancer?bkling
Are you curious about what’s new in cervical cancer research or unsure what the findings mean? Join Dr. Emily Ko, a gynecologic oncologist at Penn Medicine, to learn about the latest updates from the Society of Gynecologic Oncology (SGO) 2024 Annual Meeting on Women’s Cancer. Dr. Ko will discuss what the research presented at the conference means for you and answer your questions about the new developments.
micro teaching on communication m.sc nursing.pdfAnurag Sharma
Microteaching is a unique model of practice teaching. It is a viable instrument for the. desired change in the teaching behavior or the behavior potential which, in specified types of real. classroom situations, tends to facilitate the achievement of specified types of objectives.
Building a large-scale missing persons ID SNP panel - Download the study
1. Building a large-scale missing
persons ID SNP panel
Christopher Phillips
A. Tillmar, T.J. Parsons, R. Huel, K. Kidd,
M.V. Lareu, K. Elliott, R. Samara, E. Lader
Forensic Genetics Unit, University of Santiago
de Compostela, Spain
2. Building a large-scale missing
persons ID SNP panel
Christopher Phillips
A. Tillmar, T.J. Parsons, R. Huel, K. Kidd,
M.V. Lareu, K. Elliott, R. Samara, E. Lader
Forensic Genetics Unit, University of Santiago
de Compostela, Spain
Yale
3. ICMP 2.0 aims to redefine the scope of its identification work worldwide
and adopt MPS as key technology, founded in the new headquarters
The Hague facilities will be well suited to
adoption of new forensic DNA technologies
Bioinformatics and computational
infrastructure is well founded
Qiagen and ICMP have agreed to develop
a complete missing persons ID pipeline
4. The rationale for SNP analysis in forensics
Microhaplotype genotyping becomes
possible with the introduction of MPS
Yale
5. The rationale for SNP analysis in forensics
Microhaplotype genotyping becomes
possible with the introduction of MPS
Yale
Constructed a new MPS panel
dedicated to missing persons
identification consisting of multiple-
allele SNPs and microhaplotypes
A collaborative project that makes
use of the QIAseq MPS system:
originally developed for sensitivity
to low-level mutation sequences
6. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
7. Single Nucleotide Polymorphisms can provide complimentary data to STRs
SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Yale
8. Single Nucleotide Polymorphisms can provide complimentary data to STRs
SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Yale
9. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
Yale
Single Nucleotide Polymorphisms can provide complimentary data to STRs
10. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
Yale
Single Nucleotide Polymorphisms can provide complimentary data to STRs
11. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
Yale
The amplified fragments used to
genotype SNPs can be very short
SNPs can be more successful than
STRs when the DNA is extremely
degraded
Single Nucleotide Polymorphisms can provide alternative tests to STRs
12. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
Yale
The amplified fragments used to
genotype SNPs can be very short
SNPs can be more successful than
STRs when the DNA is extremely
degraded
Single Nucleotide Polymorphisms can provide alternative tests to STRs
13. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
Yale
The amplified fragments used to
genotype SNPs can be very short
SNPforID 52-plex SNP panel
Innsbruck experimental Mini-STRs
NIST NC-01/NC-02 Mini-STRs
SNPs can be more successful than
STRs when the DNA is extremely
degraded
Single Nucleotide Polymorphisms can provide alternative tests to STRs
15. Yale
• Bode technologies used Orchid’s Snippet
system to genotype small-scale SNP multiplexes
• Compared to mtDNA analysis this early SNP
typing pilot study was largely unsuccessful
16. Single Nucleotide Polymorphisms are one form of short genetic variation
SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
The amplified fragments used to
genotype SNPs can be very short
Yale
SNPs can be more successful than
STRs when the DNA is extremely
degraded
17. The amplified fragments used to
genotype SNPs can be very short
SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
Yale
But new types of marker can now be genotyped with MPS
SNPs can be more successful than
STRs when the DNA is extremely
degraded
18. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
The amplified fragments used to
genotype SNPs can be very short
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
New types of SNP-based markers
But new types of marker can now be genotyped with MPS
SNPs can be more successful than
STRs when the DNA is extremely
degraded
19. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
The amplified fragments used to
genotype SNPs can be very short
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
New types of SNP-based markers
But new types of marker can now be genotyped with MPS
SNPs can be more successful than
STRs when the DNA is extremely
degraded
20. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
T G
C A
T A
The amplified fragments used to
genotype SNPs can be very short
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
New types of SNP-based markers
But new types of marker can now be genotyped with MPS
SNPs can be more successful than
STRs when the DNA is extremely
degraded
21. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
T G
C A
T A
The amplified fragments used to
genotype SNPs can be very short
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
New types of SNP-based markers
But new types of marker can now be genotyped with MPS
SNPs can be more successful than
STRs when the DNA is extremely
degraded
22. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
T G
C A
T A
Microhaplotypes
SNPs in and
around STRs
The amplified fragments used to
genotype SNPs can be very short
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
New types of SNP-based markers
But new types of marker can now be genotyped with MPS
SNPs can be more successful than
STRs when the DNA is extremely
degraded
23. SNPs in coding regions underlie a
large proportion of common genetic
variation
SNPs form the basis of forensic
trait-predictive tests of externally
visible characteristics (EVCs) and
much of the ancestry informative
markers in forensic ancestry tests
Ancestry Informative SNPsTrait-Predictive SNPs
Identity Testing SNPs Lineage SNPs
T G
C A
T A
Microhaplotypes
SNPs in and
around STRs
Both can now be
genotyped with MPS
The amplified fragments used to
genotype SNPs can be very short
SNPs in closely-linked sets can be
jointly genotyped by Massively Parallel
Sequencing techniques
Phased microhaplotypes have
multiple alleles - increasing their
discrimination power
New types of SNP-based markers
SNPs can be more successful than
STRs when the DNA is extremely
degraded
But new types of marker can now be genotyped with MPS
24. • Short amplicons give best
results even with low quants
• SNP dropout is random
rather than systematic
SNPs work well with
very degraded DNA
25. • SNP dropout is random
rather than systematic
No.ofreportableloci(46intotal)
DNAconcentrationng/μl
• Short amplicons give best
results even with low quants
SNPs work well with
very degraded DNA
26. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
27. Adaptor Barcode +
Adaptor
Barcode + Adaptor
Adaptor
Clonal amplification
by Bridge
Amplification
Add adaptors and
barcodes
Clean up
Library
Removes excess
reagents
Barcode + Adaptor
Adaptor
Repair ends and
add A overhang
Adaptor Barcode + Adaptor
Add adaptors and
barcodes
Clean up
Library
Removes excess
reagents
Barcode + Adaptor
Adaptor
Clonal amplification
by Bridge
Amplification
Create blunt ends
Adaptor Barcode + Adaptor
Clonal
amplification by
Emulsion PCR
DNA fragments bind
to Ion Sphere
Particles (ISPs)
I
S
P
I
S
P
D
Target amplification
DNA is
replicated
and new
fragments
bind to ISPs
Removes oil and
excess reagents
from PCR
Clean up
Clean up
Cluster generation
CLONAL
AMPLIFICATION
SEQUENCING
LIBRARY
PREPARATION
Hybridize fragments to
flow cell glass surface
Single-stranded DNA flexes
to form a bridge, then
extended by polymerase
Double-stranded bridge
denatured leaving two
single-stranded fragments
Process repeated to form
cluster of fragments
Reverse strands cleaved and
washed off to leave forward
strand clusters on flow cell
Spheres loaded into
individual wells on
surface of semi-
conductor chip
Chip surface is flushed
and drained with
successive dNTPs
Voltage
spikes
converted to
Ionogram
Nucleotide addition
causes a change in
pH with proportional
voltage charge
Sensor plate
Sensing layer
Sequencing primers added
Sequencing performed on
forward strands. As dNTPs
are added fluorescence is
emitted as light signals
measured by a detector
Regenerate DNA fragments for
reverse strand on flow cell
Sequencing is repeated on
reverse strands. Light
signals converted to
sequence read
PCR AMPLIFICATIONDNA EXTRACTION QUANTITATION
Replicate DNA with forensically relevant primer sets to target specific sitesMeasure amount of DNARelease DNA from cells
STANDARD
FORENSIC
TARGET DNA
PREPARATION
Dr. Linzi Wilson-Wilde
MPS
28. Adaptor Barcode +
Adaptor
Barcode + Adaptor
Adaptor
Clonal amplification
by Bridge
Amplification
Add adaptors and
barcodes
Clean up
Library
Removes excess
reagents
Barcode + Adaptor
Adaptor
Repair ends and
add A overhang
Adaptor Barcode + Adaptor
Add adaptors and
barcodes
Clean up
Library
Removes excess
reagents
Barcode + Adaptor
Adaptor
Clonal amplification
by Bridge
Amplification
Create blunt ends
Adaptor Barcode + Adaptor
Clonal
amplification by
Emulsion PCR
DNA fragments bind
to Ion Sphere
Particles (ISPs)
I
S
P
I
S
P
D
Target amplification
DNA is
replicated
and new
fragments
bind to ISPs
Removes oil and
excess reagents
from PCR
Clean up
Clean up
Cluster generation
CLONAL
AMPLIFICATION
SEQUENCING
LIBRARY
PREPARATION
Hybridize fragments to
flow cell glass surface
Single-stranded DNA flexes
to form a bridge, then
extended by polymerase
Double-stranded bridge
denatured leaving two
single-stranded fragments
Process repeated to form
cluster of fragments
Reverse strands cleaved and
washed off to leave forward
strand clusters on flow cell
Spheres loaded into
individual wells on
surface of semi-
conductor chip
Chip surface is flushed
and drained with
successive dNTPs
Voltage
spikes
converted to
Ionogram
Nucleotide addition
causes a change in
pH with proportional
voltage charge
Sensor plate
Sensing layer
Sequencing primers added
Sequencing performed on
forward strands. As dNTPs
are added fluorescence is
emitted as light signals
measured by a detector
Regenerate DNA fragments for
reverse strand on flow cell
Sequencing is repeated on
reverse strands. Light
signals converted to
sequence read
PCR AMPLIFICATIONDNA EXTRACTION QUANTITATION
Replicate DNA with forensically relevant primer sets to target specific sitesMeasure amount of DNARelease DNA from cells
STANDARD
FORENSIC
TARGET DNA
PREPARATION
MPS
Dr. Linzi Wilson-Wilde
29. Mixed marker MPS panels like the Illumina DNA Signature Kit sequence the
shortest possible amplicon lengths - here the bulk of SNPs are below 125 bp
Yale
STR fragment length ranges
SNP fragment sizes
30. • Tested a DNA extract from a
12th Century male skeleton in
duplicated MPS runs
• Volders mediaeval burial site
in Tyrol has 5th/6th Century
skeletons overlaid with later
12th/13th Century remains
A recent pilot study showed the sensitivity of MPS
31. N / NN
QUAL=0
SNPs
>100 x
coverage
SNPs
20-100 x
coverage
• Tested a DNA extract from a
12th Century male skeleton in
duplicated MPS runs
• Volders mediaeval burial site
in Tyrol has 5th/6th Century
skeletons overlaid with later
12th/13th Century remains
A recent pilot study showed the sensitivity of MPS
32. 47
68
108
130
45 SNPs were
removed from the
prototype set
60 SNPs have
reduced amplicon
sizes compared to the
prototype set (by an
average 57.5 bp)
64 SNPs retain the
original prototype
primer designs
120
123
137
117
119
99
Prototype SNP
panel amplicons
Final HID-Ion
AmpliSeq™ Identity
Panel amplicons
N / NN
QUAL=0
SNPs
>100 x
coverage
SNPs
20-100 x
coverage
33. 47
68
108
130
45 SNPs were
removed from the
prototype set
60 SNPs have
reduced amplicon
sizes compared to the
prototype set (by an
average 57.5 bp)
64 SNPs retain the
original prototype
primer designs
120
123
137
117
119
99
Prototype SNP
panel amplicons
Final HID-Ion
AmpliSeq™ Identity
Panel amplicons
N / NN
QUAL=0
SNPs
>100 x
coverage
SNPs
20-100 x
coverage
TFS Precision ID
Identification SNPs
Identification SNP
Prototype Panel
169
124
35. Adaptor Barcode +
Adaptor
Barcode + Adaptor
Adaptor
Clonal amplification
by Bridge
Amplification
Add adaptors and
barcodes
Clean up
Library
Removes excess
reagents
Barcode + Adaptor
Adaptor
Repair ends and
add A overhang
Adaptor Barcode + Adaptor
Add adaptors and
barcodes
Clean up
Library
Removes excess
reagents
Barcode + Adaptor
Adaptor
Clonal amplification
by Bridge
Amplification
Create blunt ends
Adaptor Barcode + Adaptor
Clonal
amplification by
Emulsion PCR
DNA fragments bind
to Ion Sphere
Particles (ISPs)
I
S
P
I
S
P
D
Target amplification
DNA is
replicated
and new
fragments
bind to ISPs
Removes oil and
excess reagents
from PCR
Clean up
Clean up
Cluster generation
CLONAL
AMPLIFICATION
SEQUENCING
LIBRARY
PREPARATION
Hybridize fragments to
flow cell glass surface
Single-stranded DNA flexes
to form a bridge, then
extended by polymerase
Double-stranded bridge
denatured leaving two
single-stranded fragments
Process repeated to form
cluster of fragments
Reverse strands cleaved and
washed off to leave forward
strand clusters on flow cell
Spheres loaded into
individual wells on
surface of semi-
conductor chip
Chip surface is flushed
and drained with
successive dNTPs
Voltage
spikes
converted to
Ionogram
Nucleotide addition
causes a change in
pH with proportional
voltage charge
Sensor plate
Sensing layer
Sequencing primers added
Sequencing performed on
forward strands. As dNTPs
are added fluorescence is
emitted as light signals
measured by a detector
Regenerate DNA fragments for
reverse strand on flow cell
Sequencing is repeated on
reverse strands. Light
signals converted to
sequence read
PCR AMPLIFICATIONDNA EXTRACTION QUANTITATION
Replicate DNA with forensically relevant primer sets to target specific sitesMeasure amount of DNARelease DNA from cells
Dr. Linzi Wilson-Wilde
STANDARD
FORENSIC
TARGET DNA
PREPARATION
The PCR and library
preparation steps build
more complex oligos in
the QIAseq chemistry
36. Yale
The capture PCR used as the preamble to MPS can produce artefacts
The capture PCR is prone to DNA synthesis errors (at a small
frequency but exacerbated when analysing low-level DNA)
Stochastic effects can be accelerated by big differences in GC
content and therefore Tm values for any one DNA fragment
37. Yale
568 amplicons
The capture PCR is prone to DNA synthesis errors (at a small
frequency but exacerbated when analysing low-level DNA)
PCR artefacts carrying through to MPS sequence output are
particularly prevalent in the much larger multiplexes typically
used in medical sequencing analyses
The capture PCR used as the preamble to MPS can produce artefacts
Stochastic effects can be accelerated by big differences in GC
content and therefore Tm values for any one DNA fragment
38. DNA Fragmentation
Library construction ligating Barcodes
- Sample indices - F-primer sequences
Sample indexing and
amplification
MPS-ready library
MB Molecular Barcode
GSP Gene-specific primer
UP Universal primer
FP Forward F-primer
SIP Sample index primer
~9 hours
Target enrichment by single primer extension
of F-primer & Gene (SNP/MH) specific primer
Ligation of extended oligos
Yale
QIAseq MPS chemistry combines a stepwise series of specific
sequences into a composite oligonucleotide by ligation
568 amplicons
39. DNA Fragmentation
Sample indexing and amplification
MPS-ready library
MB Molecular Barcode
GSP Gene-specific primer
UP Universal primer
FP Forward F-primer
SIP Sample index primer
~9 hours
Target enrichment by single primer extension
of F-primer & Gene (SNP/MH) specific primer
Ligation of extended oligos
Yale
Library construction ligating Barcodes -
Sample indices - F-primer sequences
QIAseq MPS chemistry combines a stepwise series of specific
sequences into a composite oligonucleotide by ligation
40. DNA Fragmentation
Sample indexing and amplification
MPS-ready library
MB Molecular Barcode
GSP Gene-specific primer
UP Universal primer
FP Forward F-primer
SIP Sample index primer
~9 hours
Target enrichment by single primer extension
of F-primer & Gene (SNP/MH) specific primer
Ligation of extended oligos
Yale
Library construction ligating Barcodes -
Sample indices - F-primer sequences
QIAseq MPS chemistry combines a stepwise series of specific
sequences into a composite oligonucleotide by ligation
Tag target DNA
fragments with
UMIs: Unique
Molecular Indices
Amplify
Correct errors
with UMIs
41. DNA Fragmentation
Library construction ligating Barcodes -
Sample indices - F-primer sequences
Sample indexing and amplification
MPS-ready library
MB Molecular Barcode
GSP Gene-specific primer
UP Universal primer
FP Forward F-primer
SIP Sample index primer
~9 hours
Target enrichment by single primer extension
of F-primer & Gene (SNP/MH) specific primer
Ligation of extended oligos
A typical QIAseq oligo prepared for MiSeq analysis
Yale
QIAseq MPS chemistry combines a stepwise series of specific
sequences into a composite oligonucleotide by ligation
42. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
43. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
44. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
• SNPs provide very short fragment PCR - markers of choice for degraded DNA
• But SNPs clearly have much less information per marker than STRs
• What happens when we use SNPs to examine a very distant kinship claim ?
45. Yale
Second cousins in a fully deficient pedigree is a challenging kinship
analysis to perform - in 2011 we opted to use an Affymetrix 6.0 SNP array
46. Yale
Second cousins in a fully deficient pedigree is a challenging kinship
analysis to perform - in 2011 we opted to use an Affymetrix 6.0 SNP array
49. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
• The PCR multiplex must be bigger than scales achieved in forensic MPS before
• We also decided to use more informative SNP-based markers:
• Tri-allelic SNPs (three possible nucleotide alleles per site = 6 genotypes)
• Microhaplotypes in realistically short sequence fragments
50. Yale
Simulated kinship test
LR distributions and
linkage adjustments
Microhaplotypes
The ICMP panel was developed from collaboration between five laboratories
and combines a large number of single-site SNPs and microhaplotype loci
QIAseq MPS
technology
Marker
selection
Optimisation of
panel and
Qiagen pipeline
52. Yale
Andreas Tillmar, Linkoping established the Qiagen 140-SNP ID
panel for the MiSeq system, USC adapted it for the Ion PGM
Qiagen first
generation MPS
technology
• Good sequence coverage and
MPS performance as a third-party
kit applicable to both platforms
• Qiagen now transitioned to
QIAseq chemistry to enable much
larger PCR multiplexes and use of
multiple primer sets for enhanced
sensitivity
53. Yale
With much larger marker panels applied to kinship tests allowance for linkage
becomes important: Linkoping-USC developed 'ILIR' to adjust for linked loci
Simulated kinship test LR
distributions and linkage
adjustments
QIAseq MPS
technology Rc value
estimation
54. Yale
With much larger marker panels applied to kinship tests allowance for linkage
becomes important: Linkoping-USC developed 'ILIR' to adjust for linked loci
Simulated kinship test LR
distributions and linkage
adjustments
QIAseq MPS
technology Rc value
estimation
• Not accounting for linkage
(here, 20 STRs + 52 SNPs)
has a marked effect on LR
calculations, particularly when
testing related individuals
56. SNP combinations in each haplotype and their phase can only be obtained
by sequencing the whole strand - made possible with MPS
Yale
57. GT, CT, CT
33 = 27 genotype
combinations
Yale
SNP combinations in each haplotype and their phase can only be obtained
by sequencing the whole strand - made possible with MPS
58. G C C
T T T
T C C
G T T
G T C
T C T
T T C
G C T
G C T
T T C
T C T
G T C
G T T
T C C
T T T
A A C
GT, CT, CT
33 = 27 genotype
combinations
36 combinations
of 8 haplotypes
Yale
SNP combinations in each haplotype and their phase can only be obtained
by sequencing the whole strand - made possible with MPS
59. G C C
T T T
T C C
G T T
G T C
T C T
T T C
G C T
G C T
T T C
T C T
G T C
G T T
T C C
T T T
A A C
GT, CT, CT
33 = 27 genotype
combinations
36 combinations
of 8 haplotypes
Yale
SNP combinations in each haplotype and their phase can only be obtained
by sequencing the whole strand - made possible with MPS
60. X
G C C
T T T
T C C
G T T
G T C
T C T
T T C
G C T
G C T
T T C
T C T
G T C
G T T
T C C
T T T
A A C
GT, CT, CT
33 = 27 genotype
combinations
36 combinations
of 8 haplotypes
Yale
SNP combinations in each haplotype and their phase can only be obtained
by sequencing the whole strand - made possible with MPS
61. Yale
Microhaplotypes
Marker
selection
Not all microhaplotypes were sufficiently informative - most 2-SNP
loci gave tri-allelic patterns with similar power to these loci
• Selected 46 from 130 microhaplotypes that
have high haplotype Heterozygosity values in
most or all 1000 Genomes population groups (i.e.
seeking ‘universal’ informativeness)
• Reduced the size of some microhaplotypes and
traded a reasonable reduction of informativeness
for efficiency with degraded DNA
64. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
65. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
• 1457 markers were incorporated after linkage screening
- only 6 sites were eliminated based on primer extension disqualification
• 1377 autosomal tri-alleleic SNPs
• 34 tri-allelic X chromosome SNPs
• 46 microhaplotypes with 2, 3, 4, and 5 SNP combinations
66. Why use single nucleotide polymorphisms rather
than tried-and-tested STRs for missing persons ID?
Forensic MPS as it currently stands and the
added advantages of QIAseq chemistry
Criteria for building the ICMP missing persons ID panel
- moving away from binary markers
Characteristics of the markers incorporated
into the ICMP missing persons ID panel
• 1457 markers were incorporated after linkage screening
- only 6 sites were eliminated based on primer extension disqualification
• 1377 autosomal tri-alleleic SNPs
• 34 tri-allelic X chromosome SNPs
• 2832 target enrichment extension primers
- 80% of sites with redundant targeting
• 46 microhaplotypes with 2, 3, 4, and 5 SNP combinations
FP1
FP2
67. EUR AFR E ASN S ASN AME
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
Typical tri-allelic SNPs for identification - 0.6-0.66 average Heterozygosity
EUR AFR E ASN S ASN AME EUR AFR E ASN S ASN AME
Yale
68. 1411 Tri-allelic SNPs
Ranked average Heterozygosity amongst five 1000 Genomes
populations (American-East Asian-South Asian-European-African)
Yale
We were able to combine 1411 tri-allelic SNPs with high levels of average
heterozygosity (averaged across five 1000 Genomes population groups)
Only 1.5% of tri-allelic SNPs have a lower
average Heterozygosity than the 0.5 bi-allelic
SNP maximum value (these SNPs had skewed
allele frequencies amongst different populations)
69. STR sequence variants and Microhaplotypes
Yale
This microhaplotype exemplifies trading size for informativeness
70. STR sequence variants and Microhaplotypes
Yale
This microhaplotype exemplifies trading size for informativeness
71. S ASN
EUR
AFR
Kiddlab microhaplotype sizes
ICMP panel microhaplotype sizes
46 Microhaplotypes ranked by descending size - as originally described in Kiddlab list of 130
Average size 128-NT
Average size 60.5-NT
16 microhaplotypes had identical sizes, 14 of these were at the
extreme size range with an average 55-nucleotide size
65% of Microhaplotypes adopted for the panel had their sizes
reduced by an average 67-nucleotides
Microhaplotypespaninnucleotides
Yale
72. MH-D01 39 NT 0.7330 MH-D07 66 NT 0.7434
MH-D50 62 NT 0.6887 MH-D62 59 NT 0.7074
AME
E ASN
AFR
S ASN
EUR
ACC ACT ATC ATT TCC TCT TTC TTT CAA CAG CTA CTG TAA TAG TTA
AAA ACA ACG CAA CCACAA CAT CGT TAA TGA
AME
E ASN
AFR
S ASN
EUR
Typical microhaplotypes for ID - 0.68-0.73 average Heterozygosity
Yale
73. 46 Microhaplotypes1411 Tri-allelic SNPs
Ranked average Heterozygosity amongst five 1000 Genomes
populations (American-East Asian-South Asian-European-African)
39% of microhaplotypes have
higher average Heterozygosity
than 0.66: the tri-allelic SNP
maximum value
Only 1.5% of tri-allelic SNPs have a lower
average Heterozygosity than the 0.5 bi-allelic
SNP maximum value (these SNPs had skewed
allele frequencies amongst different populations)
98.5% of tri-allelic SNPs and all 46 microhaplotypes are more
informative than the best binary SNPs with 0.5-0.5 alleles
Yale
75. Yale
• Approximately 7-10% of the tri-allelic sites in ICMP panel are actually segmental
duplications giving false three-allele patterns (often in one individual) - so these loci have
not been sufficiently well curated by 1000 Genomes at this stage and are now discarded
from the multiplex
In conclusion: One cautionary observation made during the optimisation of
this large panel for routine missing person identification
76. Yale
In conclusion: One cautionary observation made during the optimisation of
this large panel for routine missing person identification
• Approximately 7-10% of the tri-allelic sites in ICMP panel are actually segmental
duplications giving false three-allele patterns (often in one individual) - so these loci have
not been sufficiently well curated by 1000 Genomes at this stage and are now discarded
from the multiplex
Allelic diversity vs.
segmental duplication