Next-generation sequencing technologies like ion semiconductor sequencing and pyrosequencing have enabled applications of RNA-Seq. RNA-Seq involves sequencing cDNA to analyze transcriptomes, identify differentially expressed genes between conditions, and reconstruct transcripts through methods like genome-guided assembly or by building de Bruijn graphs from k-mers.
Transcriptomics is the study of RNA, single-stranded nucleic acid, which was not separated from the DNA world until the central dogma was formulated by Francis Crick in 1958, i.e., the idea that genetic information is transcribed from DNA to RNA and then translated from RNA into protein.
Transcriptomics is the study of RNA, single-stranded nucleic acid, which was not separated from the DNA world until the central dogma was formulated by Francis Crick in 1958, i.e., the idea that genetic information is transcribed from DNA to RNA and then translated from RNA into protein.
Tag-based transcript sequencing: Comparison of SAGE and CAGEMatthias Harbers
Talk given at the European Meeting on Next Generation Sequencing, August 29 to September 1, 2010, at Leiden University Medical Center, The Netherland.
Data have been published in: Hestand et al.: “Tissue-specific transcript annotation and expression profiling with complementary next-generation sequencing technologies.” Nucleic Acids Res. 2010 Sep;38(16):e165.
PMID: 20615900
Oxford Nanopore was founded in Oxford Nanolabs by Dr.Gordon Sanghera, Dr.Spike Willcocks and Professor Hagan Bayley. Nanopore sequencing has been around since the 1990s, when Church et al. and Deamer and Akeson separately proposed that it is possible to sequence DNA using nanopore sensors.
Next generation-sequencing.ppt-convertedShweta Tiwari
The advance version, sequences the whole genome efficiently with high speed and high throughput sequencing at reduce cost is termed as Next Generation Sequencing (NGS) or massively parallel sequencing (MPS).
AGRF in conjunction with EMBL Australia recently organised a workshop at Monash University Clayton. This workshop was targeted at beginners and biologists who are new to analysing Next-Gen Sequencing data. The workshop also aimed to provide users with a snapshot of bioinformatics and data analysis tips on how to begin to analyse project data. An introduction to RNA-seq data analysis was presented by AGRF Senior Bioinformatician Dr. Sonika Tyagi.
Presented: 1st August 2012
Tag-based transcript sequencing: Comparison of SAGE and CAGEMatthias Harbers
Talk given at the European Meeting on Next Generation Sequencing, August 29 to September 1, 2010, at Leiden University Medical Center, The Netherland.
Data have been published in: Hestand et al.: “Tissue-specific transcript annotation and expression profiling with complementary next-generation sequencing technologies.” Nucleic Acids Res. 2010 Sep;38(16):e165.
PMID: 20615900
Oxford Nanopore was founded in Oxford Nanolabs by Dr.Gordon Sanghera, Dr.Spike Willcocks and Professor Hagan Bayley. Nanopore sequencing has been around since the 1990s, when Church et al. and Deamer and Akeson separately proposed that it is possible to sequence DNA using nanopore sensors.
Next generation-sequencing.ppt-convertedShweta Tiwari
The advance version, sequences the whole genome efficiently with high speed and high throughput sequencing at reduce cost is termed as Next Generation Sequencing (NGS) or massively parallel sequencing (MPS).
AGRF in conjunction with EMBL Australia recently organised a workshop at Monash University Clayton. This workshop was targeted at beginners and biologists who are new to analysing Next-Gen Sequencing data. The workshop also aimed to provide users with a snapshot of bioinformatics and data analysis tips on how to begin to analyse project data. An introduction to RNA-seq data analysis was presented by AGRF Senior Bioinformatician Dr. Sonika Tyagi.
Presented: 1st August 2012
Course: Bioinformatics for Biomedical Research (2014).
Session: 4.1- Introduction to RNA-seq and RNA-seq Data Analysis.
Statistics and Bioinformatisc Unit (UEB) & High Technology Unit (UAT) from Vall d'Hebron Research Institute (www.vhir.org), Barcelona.
RNA-Seq To Identify Novel Markers For Research on Neural Tissue DifferentiationThermo Fisher Scientific
Neural tissue differentiated and cultured from derived stem cells is expected to revolutionize the treatment of brain and spinal injuries and diseases. Critical for these cellular therapies is accurate control and monitoring of differentiation but current methods for such cell typing are limited to qPCR and immunocytochemisty (ICC) which is not sufficient to discriminate between the numerous (likely >100,000) possible neural cell-types. Research using RNA sequencing (RNA-Seq) permits the characterization and discovery of much-needed novel markers. To define the temporal transcriptional signature of neural stem cells, cultured human embryonic stem cells (H9) were compared to induced neural stem cells (NSCs) at d0, d7 and d14. Total RNA was isolated over the time course from the undifferentiated and differentiated cells. Ion Torrent™ libraries were created to profile expression of miRNAs and whole transcriptomes for each cell population. Multiplexed Ion Proton™ sequencing and Torrent Suite™ Software analysis yielded ≥2.5 million small RNA reads and ≥29 million whole transcriptome reads per sample. Cluster analysis of the RNA-Seq profiles indicates that the cell populations have characteristic molecular signatures. Among genes that are decreased in induced cells are OCT4 (POU5F1), JARID2, NANOG, consistent with the differentiation of iPSCs into neurons. Among genes that showed increased expressions are NTRK2, POU3F2, and a number of HOX family genes. Recently, Ion AmpliSeq™ Transcriptome Human Gene Expression Kit has been launched, and the results from this analysis corroborated with whole transcriptome RNA-Seq results.
Examining gene expression and methylation with next gen sequencingStephen Turner
Slides on RNA-seq and methylation studies using next-gen sequencing given at the University of Miami Hussman Institute for Human Genomics "Genetic Analysis of Complex Human Diseases" course in 2012 (http://hihg.med.miami.edu/educational-programs/analysis-of-complex-human-diseases/genetic-analysis-of-complex-human-diseases/)
Summary table that compares the performance specifications of Illumina's new sequencers - HiSeq X and NextSeq with MiSeq and HiSeq 2500. Includes number of samples that can run on the different sequencers.
Guide to Molecular Cloning - Download the GuideQIAGEN
Molecular cloning can be sometimes tricky with significant challenges involved. Overcome the challenges with the essential knowledge and tips for successful cloning.
Apollo is a web-based application that supports and enables collaborative genome curation in real time, allowing teams of curators to improve on existing automated gene models through an intuitive interface. Apollo allows researchers to break down large amounts of data into manageable portions to mobilize groups of researchers with shared interests.
The i5K, an initiative to sequence the genomes of 5,000 insect and related arthropod species, is a broad and inclusive effort that seeks to involve scientists from around the world in their genome curation process, and Apollo is serving as the platform to empower this community.
This presentation is an introduction to Apollo for the members of the i5K Pilot Project working on species of the order Hemiptera.
The analysis of all transcripts within a cell is of essential importance. Molecular biology provides many approaches to clone RNA transcripts into cDNA. Large cDNA collections are in the public domain to serve the research community. Today, however, new high-speed sequencing methods allow a much deeper view into transcriptomes than possible by classical cloning.
Presented by Dr. Miller at the 40th Annual Symposium "Diagnostic and Clinical Challenges of 20th Century Microbes", held on Nov 18, 2010 in Philadelphia.
STS stands for sequence tagged site which is short DNA sequence, generally between 100 and 500 bp in length, that is easily recognizable and occurs only once in the chromosome or genome being studied.
It contains information about- DNA Sequencing; History and Era sequencing; Next Generation Sequencing- Introduction, Workflow, Illumina/Solexa sequencing, Roche/454 sequencing, Ion Torrent sequencing, ABI-SOLiD sequencing; Comparison between NGS & Sangers and NGS Platforms; Advantages and Applications of NGS; Future Applications of NGS.
Builder.ai Founder Sachin Dev Duggal's Strategic Approach to Create an Innova...Ramesh Iyer
In today's fast-changing business world, Companies that adapt and embrace new ideas often need help to keep up with the competition. However, fostering a culture of innovation takes much work. It takes vision, leadership and willingness to take risks in the right proportion. Sachin Dev Duggal, co-founder of Builder.ai, has perfected the art of this balance, creating a company culture where creativity and growth are nurtured at each stage.
"Impact of front-end architecture on development cost", Viktor TurskyiFwdays
I have heard many times that architecture is not important for the front-end. Also, many times I have seen how developers implement features on the front-end just following the standard rules for a framework and think that this is enough to successfully launch the project, and then the project fails. How to prevent this and what approach to choose? I have launched dozens of complex projects and during the talk we will analyze which approaches have worked for me and which have not.
UiPath Test Automation using UiPath Test Suite series, part 3DianaGray10
Welcome to UiPath Test Automation using UiPath Test Suite series part 3. In this session, we will cover desktop automation along with UI automation.
Topics covered:
UI automation Introduction,
UI automation Sample
Desktop automation flow
Pradeep Chinnala, Senior Consultant Automation Developer @WonderBotz and UiPath MVP
Deepak Rai, Automation Practice Lead, Boundaryless Group and UiPath MVP
GDG Cloud Southlake #33: Boule & Rebala: Effective AppSec in SDLC using Deplo...James Anderson
Effective Application Security in Software Delivery lifecycle using Deployment Firewall and DBOM
The modern software delivery process (or the CI/CD process) includes many tools, distributed teams, open-source code, and cloud platforms. Constant focus on speed to release software to market, along with the traditional slow and manual security checks has caused gaps in continuous security as an important piece in the software supply chain. Today organizations feel more susceptible to external and internal cyber threats due to the vast attack surface in their applications supply chain and the lack of end-to-end governance and risk management.
The software team must secure its software delivery process to avoid vulnerability and security breaches. This needs to be achieved with existing tool chains and without extensive rework of the delivery processes. This talk will present strategies and techniques for providing visibility into the true risk of the existing vulnerabilities, preventing the introduction of security issues in the software, resolving vulnerabilities in production environments quickly, and capturing the deployment bill of materials (DBOM).
Speakers:
Bob Boule
Robert Boule is a technology enthusiast with PASSION for technology and making things work along with a knack for helping others understand how things work. He comes with around 20 years of solution engineering experience in application security, software continuous delivery, and SaaS platforms. He is known for his dynamic presentations in CI/CD and application security integrated in software delivery lifecycle.
Gopinath Rebala
Gopinath Rebala is the CTO of OpsMx, where he has overall responsibility for the machine learning and data processing architectures for Secure Software Delivery. Gopi also has a strong connection with our customers, leading design and architecture for strategic implementations. Gopi is a frequent speaker and well-known leader in continuous delivery and integrating security into software delivery.
Let's dive deeper into the world of ODC! Ricardo Alves (OutSystems) will join us to tell all about the new Data Fabric. After that, Sezen de Bruijn (OutSystems) will get into the details on how to best design a sturdy architecture within ODC.
Search and Society: Reimagining Information Access for Radical FuturesBhaskar Mitra
The field of Information retrieval (IR) is currently undergoing a transformative shift, at least partly due to the emerging applications of generative AI to information access. In this talk, we will deliberate on the sociotechnical implications of generative AI for information access. We will argue that there is both a critical necessity and an exciting opportunity for the IR community to re-center our research agendas on societal needs while dismantling the artificial separation between the work on fairness, accountability, transparency, and ethics in IR and the rest of IR research. Instead of adopting a reactionary strategy of trying to mitigate potential social harms from emerging technologies, the community should aim to proactively set the research agenda for the kinds of systems we should build inspired by diverse explicitly stated sociotechnical imaginaries. The sociotechnical imaginaries that underpin the design and development of information access technologies needs to be explicitly articulated, and we need to develop theories of change in context of these diverse perspectives. Our guiding future imaginaries must be informed by other academic fields, such as democratic theory and critical theory, and should be co-developed with social science scholars, legal scholars, civil rights and social justice activists, and artists, among others.
PHP Frameworks: I want to break free (IPC Berlin 2024)Ralf Eggert
In this presentation, we examine the challenges and limitations of relying too heavily on PHP frameworks in web development. We discuss the history of PHP and its frameworks to understand how this dependence has evolved. The focus will be on providing concrete tips and strategies to reduce reliance on these frameworks, based on real-world examples and practical considerations. The goal is to equip developers with the skills and knowledge to create more flexible and future-proof web applications. We'll explore the importance of maintaining autonomy in a rapidly changing tech landscape and how to make informed decisions in PHP development.
This talk is aimed at encouraging a more independent approach to using PHP frameworks, moving towards a more flexible and future-proof approach to PHP development.
Key Trends Shaping the Future of Infrastructure.pdfCheryl Hung
Keynote at DIGIT West Expo, Glasgow on 29 May 2024.
Cheryl Hung, ochery.com
Sr Director, Infrastructure Ecosystem, Arm.
The key trends across hardware, cloud and open-source; exploring how these areas are likely to mature and develop over the short and long-term, and then considering how organisations can position themselves to adapt and thrive.
Neuro-symbolic is not enough, we need neuro-*semantic*Frank van Harmelen
Neuro-symbolic (NeSy) AI is on the rise. However, simply machine learning on just any symbolic structure is not sufficient to really harvest the gains of NeSy. These will only be gained when the symbolic structures have an actual semantics. I give an operational definition of semantics as “predictable inference”.
All of this illustrated with link prediction over knowledge graphs, but the argument is general.
DevOps and Testing slides at DASA ConnectKari Kakkonen
My and Rik Marselis slides at 30.5.2024 DASA Connect conference. We discuss about what is testing, then what is agile testing and finally what is Testing in DevOps. Finally we had lovely workshop with the participants trying to find out different ways to think about quality and testing in different parts of the DevOps infinity loop.
Smart TV Buyer Insights Survey 2024 by 91mobiles.pdf91mobiles
91mobiles recently conducted a Smart TV Buyer Insights Survey in which we asked over 3,000 respondents about the TV they own, aspects they look at on a new TV, and their TV buying preferences.
5. Pyrosequencing
• pyrophosphate (PPi) is translated into light
emission in two enzymatic steps
• ATP-sulfurylase converts PPi in ATP
• luciferase converts Luciferin in
oxiluciferin and emitted light (ATP is the
fuel for this reaction).
5
7. Cyclic reversible termination
• incorporation of modified nucleotides
• fluorescence imaging
• washing of fluorescent dye and removing of
terminating group
7
14. Workflow of RNA-Seq
• sample preparation ribo minus and strand specific libs
(paired end)
• seqeuncing
• read mapping
Fragmentation of cDNA
Purification
Adapter ligation
Size-based purification of ligation
products using
PCR of ligation products
Purification and sequencing of the
fragments
14
15. Workflow of RNA-Seq
• sample preparation ribo minus and strand specific libs
(paired end)
• seqeuncing
• read mapping
Fragmentation of cDNA
Purification
Adapter ligation
Size-based purification of ligation
products using
PCR of ligation products
Purification and sequencing of the
fragments
14
17. Read mapping
unspliced alignment match PERFECTLY the
•
genome
unspliced read alignment seed : after mapping, the seed is extended with
smith waterman method
• seed methods
Burrow-Wheeler: transform genome into
efficient data structure
• Burrows-Wheeler transform methods
• spliced aligners
• exon first
• seed and extend
16
18. Read mapping (spliced alignment
methods)
2 steps:
unspliced read aligner break reads into shorter
(k-mer-)seeds.
unmapped reads are
split into fragments seed-regions are evaluated
and aligned with sensitive alignment
independently methods (smith
waterman)
much slower method
pseudogene is dysfunctional
and does not contain
introns. in the first step of
exon-first approaches reads
are aligned to the genome
based on unspliced read
mappers. here it is detected
as gene and not as
pseudogene.
17
20. Transcriptome reconstruction
exon identification: is used for very
• genome guided reconstruction
short reads (36 bp)
first map reads to the genome. the
unmapped reads are tested for all
• exon identification
possibilities of exon-exon junctions
not able to identify full transcript
structures
• genome-guided assembly
• genome independent reconstruction
19
21. Transcriptome reconstruction
exon identification: is used for very
• genome guided reconstruction
short reads (36 bp)
first map reads to the genome. the
unmapped reads are tested for all
• exon identification
possibilities of exon-exon junctions
not able to identify full transcript
structures
• genome-guided assembly
• genome independent reconstruction
19
22. Transcriptome reconstruction
break reads into k-mers.
align reads to the genome build de-brujin graph.
and use spliced reads to
build transcript graph alignment to the genome
(node: read fragment, is because of annotation
edge: link bet ween purpose
fragments) one paht
through the graph
represents an isoform of
the transcript)
20
24. Differential gene expression for RNA-Seq
• comparing the count of reads for one gene
in different experiments (conditions)
• the problem is that one read cannot
assigned uniquely to one gene, because of
the overlap of genes and their different
isoforms
22