DNA sequencing determines the order of nucleotide bases in a DNA sample. The Sanger method, also known as dideoxy or chain termination method, was the first widely used DNA sequencing technique. It involves DNA polymerase, dNTPs, a primer, and ddNTPs to terminate DNA strand extension. The fragments are separated by gel electrophoresis and the sequence is read. Next-generation sequencing methods like pyrosequencing do not require electrophoresis or fragment separation. They detect incorporated nucleotides in real-time. Nanopore sequencing detects changes in electrical current as single-stranded DNA passes through a nanopore protein.
Sanger sequencing is a method of DNA sequencing based on the selective incorporation of chain-terminating dideoxynucleotides by DNA polymerase during in vitro DNA replication.
Sanger sequencing is a method of DNA sequencing based on the selective incorporation of chain-terminating dideoxynucleotides by DNA polymerase during in vitro DNA replication.
This presentation is explains about the genome sequencing, its traditional method and modern method. This basically focus on Next Generation Sequencing and its types.
The chain-termination method developed by Frederick Sanger and coworkers in 1977. This method used fewer toxic chemicals and lower amounts of radioactivity than the Maxam and Gilbert method. Because of its comparative ease, the Sanger method was soon automated and was the method used in the first generation of DNA sequencers.
This presentation is explains about the genome sequencing, its traditional method and modern method. This basically focus on Next Generation Sequencing and its types.
The chain-termination method developed by Frederick Sanger and coworkers in 1977. This method used fewer toxic chemicals and lower amounts of radioactivity than the Maxam and Gilbert method. Because of its comparative ease, the Sanger method was soon automated and was the method used in the first generation of DNA sequencers.
Intelligent Protocol Content Analysis - Efficient Data ExtractionBialogics
IPCA is a methodology designed for the extraction of Health Protocol content out of network traffic. The approach avoids network impacts, is entirely vendor neutral and does not require the building of expensive interfaces.
radiation regulatory bodies. ( international + indian )akshayonslideshar
Radiation is harmful . right ? but who is looking after that it is being used in correct manner in hospitals .I have tried to write about some international and indian regulatory bodies.
Presentation on legal and ethical issues in open access to research data given at the RECODE early career researcher workshop, University of Sheffield 14-15th May 2015
Ethical implication of Human genome project,
International ethical & legal issues connected with human genome diversity research,
Genetic studies of ethnic races.
Protocol Design & Development: What You Need to Know to Ensure a Successful S...Brook White, PMP
Solid protocol design is critical to clinical development. No matter how well executed a clinical study is, if the underlying design is flawed, it wasn’t worth doing. In this presentation, Dr. David Shoemaker, SVP R&D, and Dr. Karen Kesler, AVP Operations, will walk through the process of developing a protocol, explain the major considerations, and point out common mistakes and challenges.
DNA consists of a linear string of nucleotides, or bases, for simplicity, referred to by the first letters of their chemical names--A, T, C and G. The process of deducing the order of nucleotides in DNA is called DNA sequencing. Since the DNA sequence confers information that the cell uses to make RNA molecules and proteins, establishing the sequence of DNA is key for understanding how genomes work. The technology for DNA sequencing was made faster and less expensive as a part of the Human Genome Project. And recent developments have profoundly increased the efficiency of DNA sequencing even further.
DNA sequencing is the process of determining the nucleic acid sequence – the order of nucleotides in DNA. It includes any method or technology that is used to determine the order of the four bases: adenine, guanine, cytosine, and thymine. The advent of rapid DNA sequencing methods has greatly accelerated biological and medical research and discovery.
Knowledge of DNA sequences has become indispensable for basic biological research, DNA Genographic Projects and in numerous applied fields such as medical diagnosis, biotechnology, forensic biology, virology and biological systematics. Comparing healthy and mutated DNA sequences can diagnose different diseases including various cancers,characterize antibody repertoire, and can be used to guide patient treatment.[5Having a quick way to sequence DNA allows for faster and more individualized medical care to be administered, and for more organisms to be identified and cataloged.
The rapid speed of sequencing attained with modern DNA sequencing technology has been instrumental in the sequencing of complete DNA sequences, or genomes, of numerous types and species of life, including the human genome and other complete DNA sequences of many animal, plant, and microbial species.
The first DNA sequences were obtained in the early 1970s by academic researchers using laborious methods based on two-dimensional chromatography. Following the development of fluorescence-based sequencing methods with a DNA sequencer, DNA sequencing has become easier and orders of magnitude faster.
DNA sequencing refers to the general laboratory technique for determining the exact sequence of nucleotides, or bases, in a DNA molecule. The sequence of the bases (often referred to by the first letters of their chemical names: A, T, C, and G) encodes the biological information that cells use to develop and operate.Whole Genome Sequencing
•Allows doctors to closely analyze a patient's genes for mutations and health indicators.
•Can detect intellectual disabilities and developmental delays.
•WGS is currently available at Yale for patients in the NICU and PICU.
•Involves Genetics.Sequencing may be utilized to determine the order of nucleotides in small targeted genomic regions or entire genomes. Illumina sequencing enables a wide variety of applications, allowing researchers to ask virtually any question related to the genome, transcriptome, or epigenome of any organism.The spectrum of analysis of NGS can extend from a small number of genes to an entire genome, depending on the goal. Whole-genome sequencing (WGS) and whole-exome sequencing (WES) provide the sequence of DNA bases across the genome and exome, respectively.Capillary electrophoresis (CE) instruments are capable of performing both Sanger sequencing and fragment analysis. Fragment analysis is a method in which DNA fragments are fluorescently labeled, separated by CE, and sized by comparison to an internal standard. sanger and Maxam-Gilbert sequencing technologies were classified
1.This presentation contain information about DNA and the mehods used for their sequencing like whole genome sequencing and shotgun sequenencing.
2.Advantages and disadvantages of whole genome sequencing and shot gun sequencing are also mentioned .
3.And the most important one is the applications of DNA sequencing.
DNA sequencing is the process of determining the nucleic acid sequence – the order of nucleotides in DNA. It includes any method or technology that is used to determine the order of the four bases: adenine, guanine, cytosine, and thymine.
Nutraceutical market, scope and growth: Herbal drug technologyLokesh Patil
As consumer awareness of health and wellness rises, the nutraceutical market—which includes goods like functional meals, drinks, and dietary supplements that provide health advantages beyond basic nutrition—is growing significantly. As healthcare expenses rise, the population ages, and people want natural and preventative health solutions more and more, this industry is increasing quickly. Further driving market expansion are product formulation innovations and the use of cutting-edge technology for customized nutrition. With its worldwide reach, the nutraceutical industry is expected to keep growing and provide significant chances for research and investment in a number of categories, including vitamins, minerals, probiotics, and herbal supplements.
Seminar of U.V. Spectroscopy by SAMIR PANDASAMIR PANDA
Spectroscopy is a branch of science dealing the study of interaction of electromagnetic radiation with matter.
Ultraviolet-visible spectroscopy refers to absorption spectroscopy or reflect spectroscopy in the UV-VIS spectral region.
Ultraviolet-visible spectroscopy is an analytical method that can measure the amount of light received by the analyte.
What is greenhouse gasses and how many gasses are there to affect the Earth.moosaasad1975
What are greenhouse gasses how they affect the earth and its environment what is the future of the environment and earth how the weather and the climate effects.
Slide 1: Title Slide
Extrachromosomal Inheritance
Slide 2: Introduction to Extrachromosomal Inheritance
Definition: Extrachromosomal inheritance refers to the transmission of genetic material that is not found within the nucleus.
Key Components: Involves genes located in mitochondria, chloroplasts, and plasmids.
Slide 3: Mitochondrial Inheritance
Mitochondria: Organelles responsible for energy production.
Mitochondrial DNA (mtDNA): Circular DNA molecule found in mitochondria.
Inheritance Pattern: Maternally inherited, meaning it is passed from mothers to all their offspring.
Diseases: Examples include Leber’s hereditary optic neuropathy (LHON) and mitochondrial myopathy.
Slide 4: Chloroplast Inheritance
Chloroplasts: Organelles responsible for photosynthesis in plants.
Chloroplast DNA (cpDNA): Circular DNA molecule found in chloroplasts.
Inheritance Pattern: Often maternally inherited in most plants, but can vary in some species.
Examples: Variegation in plants, where leaf color patterns are determined by chloroplast DNA.
Slide 5: Plasmid Inheritance
Plasmids: Small, circular DNA molecules found in bacteria and some eukaryotes.
Features: Can carry antibiotic resistance genes and can be transferred between cells through processes like conjugation.
Significance: Important in biotechnology for gene cloning and genetic engineering.
Slide 6: Mechanisms of Extrachromosomal Inheritance
Non-Mendelian Patterns: Do not follow Mendel’s laws of inheritance.
Cytoplasmic Segregation: During cell division, organelles like mitochondria and chloroplasts are randomly distributed to daughter cells.
Heteroplasmy: Presence of more than one type of organellar genome within a cell, leading to variation in expression.
Slide 7: Examples of Extrachromosomal Inheritance
Four O’clock Plant (Mirabilis jalapa): Shows variegated leaves due to different cpDNA in leaf cells.
Petite Mutants in Yeast: Result from mutations in mitochondrial DNA affecting respiration.
Slide 8: Importance of Extrachromosomal Inheritance
Evolution: Provides insight into the evolution of eukaryotic cells.
Medicine: Understanding mitochondrial inheritance helps in diagnosing and treating mitochondrial diseases.
Agriculture: Chloroplast inheritance can be used in plant breeding and genetic modification.
Slide 9: Recent Research and Advances
Gene Editing: Techniques like CRISPR-Cas9 are being used to edit mitochondrial and chloroplast DNA.
Therapies: Development of mitochondrial replacement therapy (MRT) for preventing mitochondrial diseases.
Slide 10: Conclusion
Summary: Extrachromosomal inheritance involves the transmission of genetic material outside the nucleus and plays a crucial role in genetics, medicine, and biotechnology.
Future Directions: Continued research and technological advancements hold promise for new treatments and applications.
Slide 11: Questions and Discussion
Invite Audience: Open the floor for any questions or further discussion on the topic.
(May 29th, 2024) Advancements in Intravital Microscopy- Insights for Preclini...Scintica Instrumentation
Intravital microscopy (IVM) is a powerful tool utilized to study cellular behavior over time and space in vivo. Much of our understanding of cell biology has been accomplished using various in vitro and ex vivo methods; however, these studies do not necessarily reflect the natural dynamics of biological processes. Unlike traditional cell culture or fixed tissue imaging, IVM allows for the ultra-fast high-resolution imaging of cellular processes over time and space and were studied in its natural environment. Real-time visualization of biological processes in the context of an intact organism helps maintain physiological relevance and provide insights into the progression of disease, response to treatments or developmental processes.
In this webinar we give an overview of advanced applications of the IVM system in preclinical research. IVIM technology is a provider of all-in-one intravital microscopy systems and solutions optimized for in vivo imaging of live animal models at sub-micron resolution. The system’s unique features and user-friendly software enables researchers to probe fast dynamic biological processes such as immune cell tracking, cell-cell interaction as well as vascularization and tumor metastasis with exceptional detail. This webinar will also give an overview of IVM being utilized in drug development, offering a view into the intricate interaction between drugs/nanoparticles and tissues in vivo and allows for the evaluation of therapeutic intervention in a variety of tissues and organs. This interdisciplinary collaboration continues to drive the advancements of novel therapeutic strategies.
Comparing Evolved Extractive Text Summary Scores of Bidirectional Encoder Rep...University of Maribor
Slides from:
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Track: Artificial Intelligence
https://www.etran.rs/2024/en/home-english/
Multi-source connectivity as the driver of solar wind variability in the heli...Sérgio Sacani
The ambient solar wind that flls the heliosphere originates from multiple
sources in the solar corona and is highly structured. It is often described
as high-speed, relatively homogeneous, plasma streams from coronal
holes and slow-speed, highly variable, streams whose source regions are
under debate. A key goal of ESA/NASA’s Solar Orbiter mission is to identify
solar wind sources and understand what drives the complexity seen in the
heliosphere. By combining magnetic feld modelling and spectroscopic
techniques with high-resolution observations and measurements, we show
that the solar wind variability detected in situ by Solar Orbiter in March
2022 is driven by spatio-temporal changes in the magnetic connectivity to
multiple sources in the solar atmosphere. The magnetic feld footpoints
connected to the spacecraft moved from the boundaries of a coronal hole
to one active region (12961) and then across to another region (12957). This
is refected in the in situ measurements, which show the transition from fast
to highly Alfvénic then to slow solar wind that is disrupted by the arrival of
a coronal mass ejection. Our results describe solar wind variability at 0.5 au
but are applicable to near-Earth observatories.
THE IMPORTANCE OF MARTIAN ATMOSPHERE SAMPLE RETURN.Sérgio Sacani
The return of a sample of near-surface atmosphere from Mars would facilitate answers to several first-order science questions surrounding the formation and evolution of the planet. One of the important aspects of terrestrial planet formation in general is the role that primary atmospheres played in influencing the chemistry and structure of the planets and their antecedents. Studies of the martian atmosphere can be used to investigate the role of a primary atmosphere in its history. Atmosphere samples would also inform our understanding of the near-surface chemistry of the planet, and ultimately the prospects for life. High-precision isotopic analyses of constituent gases are needed to address these questions, requiring that the analyses are made on returned samples rather than in situ.
8. SANGER SEQUENCING
It involves following components:
a) 1. Primer
b) 2. DNA template
DNA polymerase
d) 4.. dNTPs(A,T,G,C)
e) 5. ddNTPs
4 Steps:
1. Denaturation
2. Primer attachment and extension of bases
3. Termination
4. Poly acrylamide gel electrophoresis
9. This method uses dideoxynucleotide triphosphates
(ddNTPs) chain terminators :
which have a H on the 3’ carbon of the ribose
sugar instead of the normal OH found in
deoxynucleotide triphosphates (dNTPs).
Therefore in a synthesis reaction, if a
dideoxynucleotide is added instead of the normal
deoxynucleotide, the synthesis stops at that point
because the 3’OH necessary for the addition
of the next nucleotide is absent
•ssDNA addition of dNTPs elongation
• ssDNA addition of ddNTPs elongation
stops
11. The sequence of a single-stranded DNA
molecule is determined by enzymatic
synthesis of complementary
polynucleotide chains.
These chains terminating at specific
nucleotide positions.
Separate by gel electrophoresis
Read DNA sequence
PRINCIPLE OF DNA SEQUENCING
12. In the Sanger method, the DNA strand to be analyzed is used as a
template and DNA polymerase is used, in a PCR reaction, to
generate complimentary strands using primers.
Four different PCR reaction mixtures are prepared, each containing
a certain percentage of dideoxynucleoside triphosphate (ddNTP)
analogs to one of the four nucleotides (ATP, CTP, GTP or TTP).
Synthesis of the new DNA strand continues until one of these
analogs is incorporated, at which time the strand is prematurely
truncated.
Each PCR reaction will end up containing a mixture of different
lengths of DNA strands, all ending with the nucleotide that was
dideoxy labeled for that reaction.
Gel electrophoresis is then used to separate the strands of the four
reactions, in four separate lanes, and determine the sequence of
the original template based on what lengths of strands end with
what nucleotide.
PROCEDURE
13.
14. • A M Maxam
and W Gilbert-
1977
• Chemical
sequencing
Treatment of
DNA with
certain
chemicals
DNA
cuts into
fragments
Monitoring
of
sequences
MAXAM AND GILBERT METHOD
15. CH
The partially cleaved DNA fragment is subjected to five separate
chemical base
Each of which is specific for a particular base.
The resulting fragment terminate at that specific
base followed by high resolution gel electrophoresis and detection of
the labeled fragments by autoradiography
PRINCIPLE
16. The method requires radioactive labelling at one end and
purification of the DNA fragment to be sequenced.
Chemical treatment generates breaks at a small proportion of one
or two of the four nucleotide bases in each of four reactions
(G, A+G, C, C+T).
Thus a series of labelled fragments is generated, from the
radiolabelled end to the first 'cut' site in each molecule.
The fragments in the four reactions are arranged side by side in geL
electrophoresis for size separation.
To visualize the fragments, the gel is exposed to X-ray film for
autoradiography, yielding a series of dark bands each
corresponding to a radiolabelled DNA fragment, from which the
sequence may be inferred.
PROCEDURE
17. Reagents Base Specific modification
Dimethyl Sulphate
(pH)
G Methylation of N7 renders the C8-C9 bond
susceptible to cleavage.
Piperidine formate A+G Weakens the glycosidic bond of adenine
and guanine residues by protonaing
nitrogen atoms in the purine rings resulting
in depurination.
Hydrazine C+T Opens pyrimidine rings, which recyclize in
a five membered form which is valnerable.
Hydrazine + 1.5 M Nacl C Only cytocin reacts with Hydrazine.
DIFFERENT TYPES OF BASE SPECIFIC REACTION USED IN THE CHEMICAL
DEGRADATION METHOD
20. Shotgun sequencing, also known as shotgun cloning, is a method used
for sequencing long DNA strands or the whole genome.
•In shotgun sequencing, DNA is broken up randomly into numerous small
segments and overlapping regions are identified between all the individual
sequences that are generated.
• Multiple overlapping reads for the target DNA are obtained by performing
several rounds of this fragmentation and sequencing.
•Computer programs then use the overlapping ends of different reads to
assemble them into a continuous sequence.
•The shotgun approach was first used successfully with the bacterium
Haemophilus influenzae.
•Craig Venter used this method to map the Human genome project in 2001.
21.
22.
23. PYROSEQUENCING
• Pyrosequencing is the second important type of DNA sequencing
methodology in use today.
• Reaction mixture contains
DNA sample to be sequenced
Primers
Deoxynucleotides
DNA polymerase
Sulfurylase enzyme
Advantages:
Accurate
Parallel processing
Easily automated
Eliminates the need for labeled
primers and nucleotides
No need for gel electrophoresis
DISADVANTAGES
Smaller sequences
Nonlinear light response after
more than 5-6 identical nucleotides
24. Do not require electrophoresis or any other fragment seperation.
Determine which of the 4 bases is incorporate at each step during the
copying of DNA templete.
ddNTPs are not required
.
As the new strand is being made, the order in which the dNTPs are
incorporated is detected.
So the sequence can be read as the reaction proceeds.
In reaction all 4 dNTPs are not added at one time.
Each dNTP is added individually in a sequential manner
25. If particular dNTP are not incorporated then it is rapidly
degraded by nucleotidase or by washed before addition of
next dNTP.
Incorporation leads release of pyrophosphate which is
detected in an enzyme cascade that emits light.
26.
27. •The system uses the Staphylococcus auereus toxin α-hemolysin, a
robust heptameric protein which normally forms holes in membranes.
•DNA and RNA can be electrophoretically driven through a nanopore of
suitable diameter (