There are two main methods of DNA sequencing: the chain termination method (Sanger sequencing) and fluorescent sequencing. Sanger sequencing uses dideoxynucleotides that terminate DNA synthesis, producing fragments of different lengths that can be resolved on a gel. Fluorescent sequencing labels each dideoxynucleotide with a different colored dye, then uses software to analyze electrophoresed fragments by color and size. Next-generation sequencing allows high-throughput parallel sequencing of multiple DNA segments. It can be used for whole genome sequencing, targeted exome sequencing, or custom panels. Metagenomics applies next-generation sequencing to study the genomes of multiple organisms within an environmental sample.
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.
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.
DNA sequencing is the process of determining the sequence of nucleotides (A, T, G, and C) in the DNA. It includes method or technology that is used to determine the order of the four bases: adenine, thymine, guanine and cytosine.
Gene Sequencing, a tool to analyze the exact order of nucleotide sequence in the DNA -Deoxyribonucleic Acid.
Focuses on Two methods:
a. Maxam-Gilbert (Chemical Degradation) Method
b. Sanger's Method (Dideoxy Chain termination Method)
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
DNA sequencing is the process of determining the sequence of nucleotides (A, T, G, and C) in the DNA. It includes method or technology that is used to determine the order of the four bases: adenine, thymine, guanine and cytosine.
Gene Sequencing, a tool to analyze the exact order of nucleotide sequence in the DNA -Deoxyribonucleic Acid.
Focuses on Two methods:
a. Maxam-Gilbert (Chemical Degradation) Method
b. Sanger's Method (Dideoxy Chain termination Method)
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
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 - DNA sequencing is like reading the instructions inside a cellAmitSamadhiya1
DNA sequencing is like reading the instructions inside a cell. It's figuring out the exact order of the building blocks that make up our DNA, represented by the letters A, T, C, and G. This order is like a code that tells our bodies how to function and grow.
By reading this code, scientists can understand genes, diagnose diseases, and even trace our ancestry. There are different ways to sequence DNA, kind of like having a few different ways to read a book. These techniques are constantly improving, making it faster and easier to unlock the secrets hidden in our DNA.
Sequencing is one of the major technological advancement that has taken shape in the last two or three decade. Starting from Sanger and Maxam-Gilbert sequencing methods to the latest high-throughput methods, sequencing technologies has changed the the landscape of biological sciences.
This slide takes a look a the major sequencing methods over time.
Note: Several images included here have been sourced from GOOGLE IMAGES. The content has been extracted from several SCIENTIFIC PAPERS and WEBSITES.
PLEASE DO CONTACT THE AUTHOR DIRECTLY IF ANY COPYRIGHT ISSUE ARISES.
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.
Recomendações da OMS sobre cuidados maternos e neonatais para uma experiência pós-natal positiva.
Em consonância com os ODS – Objetivos do Desenvolvimento Sustentável e a Estratégia Global para a Saúde das Mulheres, Crianças e Adolescentes, e aplicando uma abordagem baseada nos direitos humanos, os esforços de cuidados pós-natais devem expandir-se para além da cobertura e da simples sobrevivência, de modo a incluir cuidados de qualidade.
Estas diretrizes visam melhorar a qualidade dos cuidados pós-natais essenciais e de rotina prestados às mulheres e aos recém-nascidos, com o objetivo final de melhorar a saúde e o bem-estar materno e neonatal.
Uma “experiência pós-natal positiva” é um resultado importante para todas as mulheres que dão à luz e para os seus recém-nascidos, estabelecendo as bases para a melhoria da saúde e do bem-estar a curto e longo prazo. Uma experiência pós-natal positiva é definida como aquela em que as mulheres, pessoas que gestam, os recém-nascidos, os casais, os pais, os cuidadores e as famílias recebem informação consistente, garantia e apoio de profissionais de saúde motivados; e onde um sistema de saúde flexível e com recursos reconheça as necessidades das mulheres e dos bebês e respeite o seu contexto cultural.
Estas diretrizes consolidadas apresentam algumas recomendações novas e já bem fundamentadas sobre cuidados pós-natais de rotina para mulheres e neonatos que recebem cuidados no pós-parto em unidades de saúde ou na comunidade, independentemente dos recursos disponíveis.
É fornecido um conjunto abrangente de recomendações para cuidados durante o período puerperal, com ênfase nos cuidados essenciais que todas as mulheres e recém-nascidos devem receber, e com a devida atenção à qualidade dos cuidados; isto é, a entrega e a experiência do cuidado recebido. Estas diretrizes atualizam e ampliam as recomendações da OMS de 2014 sobre cuidados pós-natais da mãe e do recém-nascido e complementam as atuais diretrizes da OMS sobre a gestão de complicações pós-natais.
O estabelecimento da amamentação e o manejo das principais intercorrências é contemplada.
Recomendamos muito.
Vamos discutir essas recomendações no nosso curso de pós-graduação em Aleitamento no Instituto Ciclos.
Esta publicação só está disponível em inglês até o momento.
Prof. Marcus Renato de Carvalho
www.agostodourado.com
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!
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.
These simplified slides by Dr. Sidra Arshad present an overview of the non-respiratory functions of the respiratory tract.
Learning objectives:
1. Enlist the non-respiratory functions of the respiratory tract
2. Briefly explain how these functions are carried out
3. Discuss the significance of dead space
4. Differentiate between minute ventilation and alveolar ventilation
5. Describe the cough and sneeze reflexes
Study Resources:
1. Chapter 39, Guyton and Hall Textbook of Medical Physiology, 14th edition
2. Chapter 34, Ganong’s Review of Medical Physiology, 26th edition
3. Chapter 17, Human Physiology by Lauralee Sherwood, 9th edition
4. Non-respiratory functions of the lungs https://academic.oup.com/bjaed/article/13/3/98/278874
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.
Couples presenting to the infertility clinic- Do they really have infertility...Sujoy Dasgupta
Dr Sujoy Dasgupta presented the study on "Couples presenting to the infertility clinic- Do they really have infertility? – The unexplored stories of non-consummation" in the 13th Congress of the Asia Pacific Initiative on Reproduction (ASPIRE 2024) at Manila on 24 May, 2024.
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.
Acute scrotum is a general term referring to an emergency condition affecting the contents or the wall of the scrotum.
There are a number of conditions that present acutely, predominantly with pain and/or swelling
A careful and detailed history and examination, and in some cases, investigations allow differentiation between these diagnoses. A prompt diagnosis is essential as the patient may require urgent surgical intervention
Testicular torsion refers to twisting of the spermatic cord, causing ischaemia of the testicle.
Testicular torsion results from inadequate fixation of the testis to the tunica vaginalis producing ischemia from reduced arterial inflow and venous outflow obstruction.
The prevalence of testicular torsion in adult patients hospitalized with acute scrotal pain is approximately 25 to 50 percent
For Better Surat #ℂall #Girl Service ❤85270-49040❤ Surat #ℂall #Girls
DNA sequencing
1. Microbial genetics
Page 1 of 5
DNA Sequencing
DNA sequencing, process by which the precise order of nucleotides in a piece of DNA
can be determined.
Methods of DNA sequencing
A. Radioactive methods
The chemical degradation method (Maxam-Gilbert method), in which the
sequence of a double-stranded DNA molecule is determined by treatment
with chemicals that cut the molecule at specific nucleotide positions. Cleaves
DNA template at different nucleotide positions, G, A+G, T+C and C and labels
these cleaved fragments.Read A and T by interpreting double bands also
including G (with A) and C (with T) .Not used very much historically
The chain termination method (Sanger dideoxy (enzymatic) sequencing)
in which the sequence of a single-stranded DNA molecule is determined by
enzymatic synthesis of complementary polynucleotide chains, these chains
terminating at specific nucleotide positions;
Amount of time and labor per sequence, cost of isotope, failed gels or failed
reactions, etc., result in hand sequencing being somewhat more expensive
to do than to do automated sequencing, and less reliable!
The term dideoxy comes from a special modified nucleotide, called a dideoxynucleotide
triphosphate (generically, a ddNTP). This modified nucleotide is key to the Sanger
technique because of its ability to block continued DNA synthesis. Because a
dideoxynucleotide lacks the 3’ hydroxyl group, this reaction cannot take place, and
therefore DNA synthesis is blocked at the point of addition. The logic of dideoxy
sequencing is straightforward.
Consider a segment of DNA that is about 1000 base pairs long that we wish to sequence.
(1) The two DNA strands are separated. Heating to 100˚C to melt the base pairing
hydrogen bonds that hold the strands together does this.
(2) A short oligonucleotide (ca. 18 bases) designed to be complimentary to the end of
one of the strands is allowed to anneal to the single stranded DNA.
(3) DNA polymerase is added along with the four nucleotide precursors (dATP,
dGTP, dCTP, and dTTP). The mixture is then divided into four separate reactions
and to each reaction a small quantity different dideoxy nucleotide precursor is
added. Dideoxy nucleotide precursors are abbreviated ddATP, ddGTP, ddCTP,
and ddTTP.
(4) The polymerase reactions are allowed to proceed and, using one of a variety of
methods, radiolabel is incorporated into the newly synthesized DNA.
(5) After the DNA polymerase reactions are complete, the samples are melted and
run on a gel system that allows DNA strands of different lengths to be resolved.
The DNA sequence can be read from the gel by noting the positions of the
radiolabeled fragments.
Obtaining the complete nucleotide sequence of a segment of DNA is often an important
part of understanding the organization of a gene and its regulation, its relation to other
genes, or the function of its encoded RNA or protein. Indeed, for the most part, translating
the nucleic acid sequence of a cDNA to discover the amino sequence of its encoded
polypeptide chain is simpler than directly sequencing the polypeptide itself.
2. Microbial genetics
Page 2 of 5
Thus dideoxynuclotides can be incorporated into DNA, but once a dideoxynuclotide has
been incorporated further elongation stops because the resulting DNA will no longer have
a free 3’ OH end. Each of the four reactions contains one of the dideoxynuclotides added
at about 1% the concentration of the normal nucleotide precursors. Thus, for example, in
the reaction with added ddATP about 1% of the elongated chains will terminate at the
position of each A in the sequence. Once all of the elongating chains have been
terminated there will be a population of labeled chains that have terminated at the
position of each A in the sequence.
B- Automated fluorescent sequencing
Cycle-sequencing in a thermal cycler, in one tube, with a commercially prepared mixture,
Still uses ddNTP technology of Sanger method, but with ddNTPs labeled with a different
colored fluorescent dye
ddATPs labeled with a green dye ,ddCTPs labeled with a blue dye
ddGTPs labeled with a black dye ,ddTTPs labeled with a red dye
Fragments electrophoresed on a polyacrylamide gel. As samples electrophorese across
a slit or window, a laser "reads" the fluorescent peaks representing different-sized
fragments Subsequent analysis of intensity peaks, and their colors, against a known
standard, gives the sequence.
Much less time and cost in labor, fewer failed reactions, fewer problem points for
"trouble-shooting" or optimization; cost of sequencing is largest expense, cost of cycle-
sequencing is next one, but tiny volumes used make this relatively cheap
3. Microbial genetics
Page 3 of 5
Suppose we have determined the nucleotide sequence of DNA fragment. How can we
tell whether it contains one or more genes? The nucleotide sequence is fed into a
computer, which then scans all six reading frames (three in each direction) in the search
for possible protein-coding regions that begin with an ATG initiation codon, end with a
stop codon, and are long enough that an uninterrupted sequence of its length is unlikely
to have arisen by chance. These stretches are called open reading frames (ORFs).
They represent sequences that are candidate genes.
Genome sequencing
A single chain termination sequencing experiment produces about 750 bp of
sequence, Clearly a large number of sequencing experiments must be carried out in
order to determine the sequence of an entire genome.
Two different strategies have been developed for sequence assembly (Figure ):
The shotgun approach, in which the genome is randomly broken into short
fragments. The resulting sequences are examined for overlaps and these are
used to build up the contiguous genome sequence.
The clone contig approach, which involves a pre-sequencing phase during which a
series of overlapping clones is identified. This contiguous series is called a contig.
Each piece of cloned DNA is then sequenced, and this sequence placed at its
appropriate position on the contig map in order to gradually build up the
overlapping genome sequence.
Chromosomal Organization of Genes and Noncoding DNA
In the genomes of prokaryotes and most lower eukaryotes, which contain few
nonfunctional sequences, coding regions are densely arrayed along the genomic
DNA.
In contrast, vertebrate genomes contain many sequences that do not code for RNAs
or have any structural or regulatory function. Much of this nonfunctional DNA is
composed of repeated sequences. In humans, only about 1.5 percent of total DNA
(the exons) actually encodes proteins or functional RNAs.
Eukaryotic genomic DNA consists of three major classes of sequences: genes
encoding proteins and functional RNAs, including gene families and tandemly
repeated genes; repetitious DNA; and spacer DNA
4. Microbial genetics
Page 4 of 5
• Next-generation” sequencing (NGS) is a high-throughput technology that
enables simultaneous sequencing of multiple DNA segments in a sample. This
analysis is accomplished by parallel sequencing and aligning these to a reference
sequence.
Different platforms use different sequencing approaches, NGS can be used to sequence
the entire complement of DNA in a sample (whole genome sequencing—WGS) or
specific segments can be isolated for sequencing. These can include all of the exons
(whole exome sequencing—WES), or specific regions of interest, creating “panels” of
genes to be sequenced.
5. Microbial genetics
Page 5 of 5
Metagenomics is the study of genomes from multiple organisms that inhabit a particular
environment. Metagenomics is an NGS applied to complex biological samples containing
several or many different organisms (mostly micro-organisms) as they are found in
environmental samples or stool probes.
In order to obtain the list of organisms present in the sample as well as their relative
abundance, either a genomic region of high variability flanked by well-conserved regions
is amplified (such as 16S ribosomal RNA fragments) or simply all DNA fragments present
in the samples are amplified. In the second case, alignment of the sequences with all
potential reference organisms is necessary.