Genome organisation in eukaryotes...........!!!!!!!!!!!manish chovatiya
This document discusses the organization of eukaryotic genomes. It explains that eukaryotic genomes are much larger than prokaryotic genomes, with most of the DNA being non-coding. Eukaryotic genomes contain multiple linear chromosomes, introns, repetitive sequences, and both coding and non-coding RNA genes. The document also describes different types of repetitive elements like tandem repeats, transposons, retrotransposons, LINEs, SINEs and their roles in increasing genome size. Overall, the document provides an overview of the complex structure of eukaryotic genomes compared to simpler prokaryotic genomes.
DNA sequencing is a process to determine the order of nucleotides in a DNA molecule. It was discovered in the 1970s by scientists like Frederick Sanger who developed the chain termination method. This method involves DNA replication with modified nucleotides that cause the growing DNA strand to terminate at that point. The fragments are then separated by size to reveal the sequence. Automated sequencing now uses fluorescent dyes and capillary electrophoresis for faster and higher throughput sequencing. DNA sequencing has applications in medicine, forensics, and agriculture.
DNA sequencing refers to determining the order of nucleotides in a DNA molecule. The first DNA sequence was obtained in the 1970s using chromatography. Modern methods use dye-based sequencing and automation. The two main historical methods are the Maxam-Gilbert chemical degradation method and the Sanger dideoxy chain termination method. Next generation sequencing now allows millions of DNA molecules to be sequenced in parallel through massively parallel sequencing technologies.
Chloroplasts are organelles found in plant cells and algae that conduct photosynthesis. They contain their own DNA known as the chloroplast genome, which is typically 100-200kb in size and encodes genes for photosynthesis. The chloroplast genome is highly conserved and maternally inherited. It has been used for phylogenetic studies and shows potential for genetic engineering due to high transgene expression and maternal inheritance that prevents gene flow to other species.
The document discusses techniques for DNA sequencing, including early methods developed in the 1970s by Maxam and Gilbert as well as Sanger. It provides details on how both methods work, such as using specific chemical or enzymatic reactions to generate labeled DNA fragments of different lengths corresponding to nucleotide positions in the sequence. The document also describes how these methods were later automated, using fluorescent tags on dideoxynucleotides and capillary electrophoresis to simultaneously sequence multiple samples in a single gel. This allowed rapid determination of thousands of nucleotides and enabled large genome sequencing projects such as the Human Genome Project.
whole genome analysis
history
needs
steps involved
human genome data
NGS
pyrosequencing
illumina
SOLiD
Ion torrent
PacBio
applications
problems
benefits
This document discusses several types of PCR techniques and their applications. It begins by explaining standard PCR and its development. It then describes several specialized PCR techniques including allele-specific PCR, asymmetric PCR, assembly PCR, hot-start PCR, helicase-dependent amplification, in situ PCR, inverse PCR, ligation-mediated PCR, and multiplex ligation-dependent probe amplification. Each technique is explained and examples of its uses and applications are provided.
Genome organisation in eukaryotes...........!!!!!!!!!!!manish chovatiya
This document discusses the organization of eukaryotic genomes. It explains that eukaryotic genomes are much larger than prokaryotic genomes, with most of the DNA being non-coding. Eukaryotic genomes contain multiple linear chromosomes, introns, repetitive sequences, and both coding and non-coding RNA genes. The document also describes different types of repetitive elements like tandem repeats, transposons, retrotransposons, LINEs, SINEs and their roles in increasing genome size. Overall, the document provides an overview of the complex structure of eukaryotic genomes compared to simpler prokaryotic genomes.
DNA sequencing is a process to determine the order of nucleotides in a DNA molecule. It was discovered in the 1970s by scientists like Frederick Sanger who developed the chain termination method. This method involves DNA replication with modified nucleotides that cause the growing DNA strand to terminate at that point. The fragments are then separated by size to reveal the sequence. Automated sequencing now uses fluorescent dyes and capillary electrophoresis for faster and higher throughput sequencing. DNA sequencing has applications in medicine, forensics, and agriculture.
DNA sequencing refers to determining the order of nucleotides in a DNA molecule. The first DNA sequence was obtained in the 1970s using chromatography. Modern methods use dye-based sequencing and automation. The two main historical methods are the Maxam-Gilbert chemical degradation method and the Sanger dideoxy chain termination method. Next generation sequencing now allows millions of DNA molecules to be sequenced in parallel through massively parallel sequencing technologies.
Chloroplasts are organelles found in plant cells and algae that conduct photosynthesis. They contain their own DNA known as the chloroplast genome, which is typically 100-200kb in size and encodes genes for photosynthesis. The chloroplast genome is highly conserved and maternally inherited. It has been used for phylogenetic studies and shows potential for genetic engineering due to high transgene expression and maternal inheritance that prevents gene flow to other species.
The document discusses techniques for DNA sequencing, including early methods developed in the 1970s by Maxam and Gilbert as well as Sanger. It provides details on how both methods work, such as using specific chemical or enzymatic reactions to generate labeled DNA fragments of different lengths corresponding to nucleotide positions in the sequence. The document also describes how these methods were later automated, using fluorescent tags on dideoxynucleotides and capillary electrophoresis to simultaneously sequence multiple samples in a single gel. This allowed rapid determination of thousands of nucleotides and enabled large genome sequencing projects such as the Human Genome Project.
whole genome analysis
history
needs
steps involved
human genome data
NGS
pyrosequencing
illumina
SOLiD
Ion torrent
PacBio
applications
problems
benefits
This document discusses several types of PCR techniques and their applications. It begins by explaining standard PCR and its development. It then describes several specialized PCR techniques including allele-specific PCR, asymmetric PCR, assembly PCR, hot-start PCR, helicase-dependent amplification, in situ PCR, inverse PCR, ligation-mediated PCR, and multiplex ligation-dependent probe amplification. Each technique is explained and examples of its uses and applications are provided.
The document summarizes DNA sequencing methods. It discusses the DNA double helix structure and how the four nitrogenous bases form complementary pairs between strands. It then describes the two main historical DNA sequencing methods: the Maxam-Gilbert method which uses chemical degradation, and the Sanger method which is based on chain termination using dideoxynucleotides. The Sanger method is now the most common approach and involves sequencing in four separate reactions with one of the four ddNTPs added to each.
DNA methylation is an epigenetic mechanism that involves the addition of a methyl group to cytosine residues in DNA. It is catalyzed by DNA methyltransferase enzymes and plays a key role in gene expression and cellular differentiation. Aberrant DNA methylation, including both hypermethylation and hypomethylation, has been associated with cancer development by disrupting gene expression. Detection of DNA methylation patterns can provide insights into cancer biology and may have applications as a diagnostic tool.
DNA sequencing is the process of determining the order of nucleotides in a DNA molecule. The Sanger method, developed in 1977, was the most widely used sequencing technique for 25 years. It utilizes chain termination with dideoxynucleotides which lack a 3' OH group, preventing formation of a phosphodiester bond and terminating strand elongation. Four reactions are run in parallel with each dideoxynucleotide labeled with a different color. Gel electrophoresis separates the terminated fragments by size, allowing the DNA sequence to be read by matching fragment sizes to nucleotide colors.
454 pyrosequencing is a DNA sequencing method based on detecting pyrophosphate release during nucleotide incorporation. It sequences a single strand of DNA 400-500 base pairs in length. The process involves emulsion PCR to amplify DNA fragments on beads, which are then loaded into wells along with enzymes and substrates. Upon addition of a nucleotide, DNA polymerase incorporates it into the growing strand and releases pyrophosphate, triggering a light-emitting reaction proportional to the amount incorporated and identifying the nucleotide.
This document discusses nucleotide probes, which are single-stranded DNA or RNA fragments that are labeled and complementary to a target DNA sequence. Probes can range in size from 15 base pairs to several hundred kilobases. They are used to identify a specific DNA fragment through base pairing. Probes must be labeled to be detected, typically through radioactive labeling or fluorescent tags. Labeling can occur on the end of the probe or through polymerase-based incorporation of multiple labeled nucleotides during DNA synthesis. Probes have various uses, including searching DNA libraries and diagnosing genetic disorders through techniques like Southern and Northern blotting.
Ion Torrent (Proton/PGM) and SOLiD sequencing are two types of next-generation sequencing technologies. Ion Torrent uses semiconductor sequencing to detect hydrogen ions released during DNA synthesis, while SOLiD uses ligation of octamer probes and fluorescent dyes to determine sequences in color space. Both have advantages such as fast run times and high throughput but also limitations including errors in homopolymers for Ion Torrent and issues with palindromic sequences for SOLiD.
Mitochondria contain their own circular genome that is 16.5kb in size and located in the mitochondrial matrix. The mitochondrial genome contains 37 genes that encode 13 proteins, 22 tRNAs, and 2 rRNAs. These genes help produce enzymes and proteins that are crucial for oxidative phosphorylation and energy production in mitochondria. The control region of mitochondrial DNA contains signals that regulate mitochondrial DNA and RNA synthesis.
Gene expression and transcript profiling involves determining the pattern of genes expressed at the transcriptional level under specific circumstances by measuring the expression of thousands of genes simultaneously. This allows one to understand cellular function. Common techniques for profiling include DNA microarrays, RNA sequencing, and EST tags. DNA microarrays involve hybridizing cDNA or cRNA samples to probes on a chip to determine relative abundance of sequences. RNA sequencing uses next-generation sequencing to reveal presence and quantity of RNA in a sample.
1) DNA sequencing refers to determining the order of nucleotide bases (A, G, C, T) in a DNA molecule. This provides essential genetic information for growth and development.
2) Two major early methods for DNA sequencing were the chemical cleavage method developed by Maxam and Gilbert in 1977 and the chain termination method developed by Sanger. Sanger's method became more popular due to fewer toxic chemicals.
3) Modern DNA sequencing often uses fluorescent dye-labeled chain terminators and capillary electrophoresis. Each dye fluoresces at a different wavelength, allowing all four reactions to occur in one tube. This high-throughput automated approach has accelerated genomic research.
The document discusses DNA sequencing techniques. It defines DNA sequencing as determining the exact order of nucleotides within a DNA molecule. The first DNA sequences were obtained in the 1970s using 2D chromatography. Sanger and Maxam-Gilbert sequencing were the first generation techniques, with Sanger using DNA polymerase and Maxam-Gilbert using chemical degradation. Next generation sequencing allows millions of reactions in parallel and produces short reads quickly and at low cost without electrophoresis. It utilizes cluster generation and sequencing methods like pyrosequencing, reversible terminators, semiconductor, and ligation. Data analysis involves separating reads, clustering, pairing strands, and aligning to reference genomes.
This document discusses primer design for PCR. It begins by defining a primer as a short DNA sequence that is complementary to the target sequence and needed to initiate DNA replication. It notes that primers are essential for PCR, acting like tires on a car. The document then outlines general rules for effective primer design, including having a length of 18-30 nucleotides, a melting temperature of 55-65°C, less than 5°C difference between primer pairs, avoiding primer dimers and secondary structures, having a GC content of 40-60%, and targeting a product size between 150bp to 10kbp.
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.
Ribozymes are RNA molecules that can catalyze biochemical reactions like protein enzymes. The first ribozyme was discovered in 1980. There are two main classes of natural ribozymes - self-cleaving ribozymes like hammerhead and hairpin ribozymes, and self-splicing ribozymes like group I and group II introns and RNase P. Ribozymes are being investigated for their potential in gene therapy applications by specifically cleaving target mRNA molecules.
DNA methylation is a biological process where methyl groups are added to DNA, changing gene expression without altering the DNA sequence. It is essential for normal development in mammals and is associated with processes like genomic imprinting and carcinogenesis. DNA methyltransferases are enzymes that catalyze the addition of methyl groups to DNA from S-adenosyl methionine. DNA methylation plays important roles in gene silencing, X-chromosome inactivation, and suppressing viral genomes and repetitive elements incorporated into the host genome. Abnormal DNA methylation is also associated with cancer by transcriptionally silencing tumor suppressor genes.
The human mitochondrial genome is much smaller than the nuclear genome, consisting of 16,569 base pairs. It contains 37 genes, 13 of which code for proteins involved in cellular respiration. Mitochondrial DNA is inherited solely from the mother and encodes for transfer RNA, ribosomal RNA and proteins that are critical subunits of the oxidative phosphorylation complexes. The human mitochondrial genome has a highly condensed structure with minimal non-coding regions and some overlapping genes. It also differs slightly from the standard genetic code.
Chloroplasts are organelles found in plant cells that capture light energy through photosynthesis. This document outlines the history and organization of chloroplast genomes, methods for chloroplast transformation, and applications in biotechnology. Specifically, it details a case study where the chloroplast genome of rapeseed was successfully transformed to be resistant to the antibiotic spectinomycin, demonstrating the potential for chloroplast transformation in important crop species. Overall, chloroplast transformation allows for high levels of protein expression and containment of transgenes, representing promising applications in biotechnology and molecular biology research.
The document summarizes Ion Torrent sequencing technology. It detects hydrogen ions released during DNA polymerization rather than using optics. The sequencing occurs on semiconductor chips patterned through photolithography into wells, each sequencing a different template. As nucleotides are incorporated, hydrogen ions change the pH detected by ion sensors below each well. This allows massively parallel sequencing that is faster, cheaper and simpler than previous technologies.
Pyrosequencing is a sequencing by synthesis technique that uses a luciferase enzyme system to monitor DNA synthesis. It works by adding DNA polymerase and a single nucleotide to the DNA fragments, generating pyrophosphate that is converted to light. The light is detected and identifies the nucleotide incorporated. Pyrosequencing has applications in cDNA analysis, mutation detection, re-sequencing of disease genes, and identifying single nucleotide polymorphisms and typing bacteria and viruses.
This document summarizes two important tumor suppressor genes - PRB and P53. It provides background on tumor suppressor genes, noting that they function through loss of function to regulate cell cycle and suppress uncontrolled cell proliferation. For PRB, it describes its role in retinoblastoma cancer and cell cycle regulation. For P53, it discusses its role as the "guardian of the genome" in DNA repair and apoptosis, as well as its structure and functions in halting the cell cycle when damage is detected.
Polymerase chain reaction (PCR) is a technique developed by Kerry Mullis in 1984 that uses thermal cycling to amplify a specific DNA across several orders of magnitude, generating millions of copies of the target DNA segment. It involves repeated cycles of separating DNA strands through heating, annealing primers to the strands through cooling, and extending the primers with a thermostable DNA polymerase through heating. This allows for rapid and efficient amplification of targeted DNA regions.
This document discusses polymerase chain reaction (PCR), a technique used to amplify a specific segment of DNA. It provides background on PCR's history and development in the 1980s. The key components of PCR are described, including DNA template, primers, DNA polymerase, nucleotides, and a thermal cycler. The basic steps of PCR are explained as denaturation, annealing and extension, which are repeated in cycles to exponentially amplify the target DNA sequence. Various applications and types of PCR are also outlined, along with its advantages of being fast, sensitive and not requiring radioactivity, though it can be prone to contamination.
The document summarizes DNA sequencing methods. It discusses the DNA double helix structure and how the four nitrogenous bases form complementary pairs between strands. It then describes the two main historical DNA sequencing methods: the Maxam-Gilbert method which uses chemical degradation, and the Sanger method which is based on chain termination using dideoxynucleotides. The Sanger method is now the most common approach and involves sequencing in four separate reactions with one of the four ddNTPs added to each.
DNA methylation is an epigenetic mechanism that involves the addition of a methyl group to cytosine residues in DNA. It is catalyzed by DNA methyltransferase enzymes and plays a key role in gene expression and cellular differentiation. Aberrant DNA methylation, including both hypermethylation and hypomethylation, has been associated with cancer development by disrupting gene expression. Detection of DNA methylation patterns can provide insights into cancer biology and may have applications as a diagnostic tool.
DNA sequencing is the process of determining the order of nucleotides in a DNA molecule. The Sanger method, developed in 1977, was the most widely used sequencing technique for 25 years. It utilizes chain termination with dideoxynucleotides which lack a 3' OH group, preventing formation of a phosphodiester bond and terminating strand elongation. Four reactions are run in parallel with each dideoxynucleotide labeled with a different color. Gel electrophoresis separates the terminated fragments by size, allowing the DNA sequence to be read by matching fragment sizes to nucleotide colors.
454 pyrosequencing is a DNA sequencing method based on detecting pyrophosphate release during nucleotide incorporation. It sequences a single strand of DNA 400-500 base pairs in length. The process involves emulsion PCR to amplify DNA fragments on beads, which are then loaded into wells along with enzymes and substrates. Upon addition of a nucleotide, DNA polymerase incorporates it into the growing strand and releases pyrophosphate, triggering a light-emitting reaction proportional to the amount incorporated and identifying the nucleotide.
This document discusses nucleotide probes, which are single-stranded DNA or RNA fragments that are labeled and complementary to a target DNA sequence. Probes can range in size from 15 base pairs to several hundred kilobases. They are used to identify a specific DNA fragment through base pairing. Probes must be labeled to be detected, typically through radioactive labeling or fluorescent tags. Labeling can occur on the end of the probe or through polymerase-based incorporation of multiple labeled nucleotides during DNA synthesis. Probes have various uses, including searching DNA libraries and diagnosing genetic disorders through techniques like Southern and Northern blotting.
Ion Torrent (Proton/PGM) and SOLiD sequencing are two types of next-generation sequencing technologies. Ion Torrent uses semiconductor sequencing to detect hydrogen ions released during DNA synthesis, while SOLiD uses ligation of octamer probes and fluorescent dyes to determine sequences in color space. Both have advantages such as fast run times and high throughput but also limitations including errors in homopolymers for Ion Torrent and issues with palindromic sequences for SOLiD.
Mitochondria contain their own circular genome that is 16.5kb in size and located in the mitochondrial matrix. The mitochondrial genome contains 37 genes that encode 13 proteins, 22 tRNAs, and 2 rRNAs. These genes help produce enzymes and proteins that are crucial for oxidative phosphorylation and energy production in mitochondria. The control region of mitochondrial DNA contains signals that regulate mitochondrial DNA and RNA synthesis.
Gene expression and transcript profiling involves determining the pattern of genes expressed at the transcriptional level under specific circumstances by measuring the expression of thousands of genes simultaneously. This allows one to understand cellular function. Common techniques for profiling include DNA microarrays, RNA sequencing, and EST tags. DNA microarrays involve hybridizing cDNA or cRNA samples to probes on a chip to determine relative abundance of sequences. RNA sequencing uses next-generation sequencing to reveal presence and quantity of RNA in a sample.
1) DNA sequencing refers to determining the order of nucleotide bases (A, G, C, T) in a DNA molecule. This provides essential genetic information for growth and development.
2) Two major early methods for DNA sequencing were the chemical cleavage method developed by Maxam and Gilbert in 1977 and the chain termination method developed by Sanger. Sanger's method became more popular due to fewer toxic chemicals.
3) Modern DNA sequencing often uses fluorescent dye-labeled chain terminators and capillary electrophoresis. Each dye fluoresces at a different wavelength, allowing all four reactions to occur in one tube. This high-throughput automated approach has accelerated genomic research.
The document discusses DNA sequencing techniques. It defines DNA sequencing as determining the exact order of nucleotides within a DNA molecule. The first DNA sequences were obtained in the 1970s using 2D chromatography. Sanger and Maxam-Gilbert sequencing were the first generation techniques, with Sanger using DNA polymerase and Maxam-Gilbert using chemical degradation. Next generation sequencing allows millions of reactions in parallel and produces short reads quickly and at low cost without electrophoresis. It utilizes cluster generation and sequencing methods like pyrosequencing, reversible terminators, semiconductor, and ligation. Data analysis involves separating reads, clustering, pairing strands, and aligning to reference genomes.
This document discusses primer design for PCR. It begins by defining a primer as a short DNA sequence that is complementary to the target sequence and needed to initiate DNA replication. It notes that primers are essential for PCR, acting like tires on a car. The document then outlines general rules for effective primer design, including having a length of 18-30 nucleotides, a melting temperature of 55-65°C, less than 5°C difference between primer pairs, avoiding primer dimers and secondary structures, having a GC content of 40-60%, and targeting a product size between 150bp to 10kbp.
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.
Ribozymes are RNA molecules that can catalyze biochemical reactions like protein enzymes. The first ribozyme was discovered in 1980. There are two main classes of natural ribozymes - self-cleaving ribozymes like hammerhead and hairpin ribozymes, and self-splicing ribozymes like group I and group II introns and RNase P. Ribozymes are being investigated for their potential in gene therapy applications by specifically cleaving target mRNA molecules.
DNA methylation is a biological process where methyl groups are added to DNA, changing gene expression without altering the DNA sequence. It is essential for normal development in mammals and is associated with processes like genomic imprinting and carcinogenesis. DNA methyltransferases are enzymes that catalyze the addition of methyl groups to DNA from S-adenosyl methionine. DNA methylation plays important roles in gene silencing, X-chromosome inactivation, and suppressing viral genomes and repetitive elements incorporated into the host genome. Abnormal DNA methylation is also associated with cancer by transcriptionally silencing tumor suppressor genes.
The human mitochondrial genome is much smaller than the nuclear genome, consisting of 16,569 base pairs. It contains 37 genes, 13 of which code for proteins involved in cellular respiration. Mitochondrial DNA is inherited solely from the mother and encodes for transfer RNA, ribosomal RNA and proteins that are critical subunits of the oxidative phosphorylation complexes. The human mitochondrial genome has a highly condensed structure with minimal non-coding regions and some overlapping genes. It also differs slightly from the standard genetic code.
Chloroplasts are organelles found in plant cells that capture light energy through photosynthesis. This document outlines the history and organization of chloroplast genomes, methods for chloroplast transformation, and applications in biotechnology. Specifically, it details a case study where the chloroplast genome of rapeseed was successfully transformed to be resistant to the antibiotic spectinomycin, demonstrating the potential for chloroplast transformation in important crop species. Overall, chloroplast transformation allows for high levels of protein expression and containment of transgenes, representing promising applications in biotechnology and molecular biology research.
The document summarizes Ion Torrent sequencing technology. It detects hydrogen ions released during DNA polymerization rather than using optics. The sequencing occurs on semiconductor chips patterned through photolithography into wells, each sequencing a different template. As nucleotides are incorporated, hydrogen ions change the pH detected by ion sensors below each well. This allows massively parallel sequencing that is faster, cheaper and simpler than previous technologies.
Pyrosequencing is a sequencing by synthesis technique that uses a luciferase enzyme system to monitor DNA synthesis. It works by adding DNA polymerase and a single nucleotide to the DNA fragments, generating pyrophosphate that is converted to light. The light is detected and identifies the nucleotide incorporated. Pyrosequencing has applications in cDNA analysis, mutation detection, re-sequencing of disease genes, and identifying single nucleotide polymorphisms and typing bacteria and viruses.
This document summarizes two important tumor suppressor genes - PRB and P53. It provides background on tumor suppressor genes, noting that they function through loss of function to regulate cell cycle and suppress uncontrolled cell proliferation. For PRB, it describes its role in retinoblastoma cancer and cell cycle regulation. For P53, it discusses its role as the "guardian of the genome" in DNA repair and apoptosis, as well as its structure and functions in halting the cell cycle when damage is detected.
Polymerase chain reaction (PCR) is a technique developed by Kerry Mullis in 1984 that uses thermal cycling to amplify a specific DNA across several orders of magnitude, generating millions of copies of the target DNA segment. It involves repeated cycles of separating DNA strands through heating, annealing primers to the strands through cooling, and extending the primers with a thermostable DNA polymerase through heating. This allows for rapid and efficient amplification of targeted DNA regions.
This document discusses polymerase chain reaction (PCR), a technique used to amplify a specific segment of DNA. It provides background on PCR's history and development in the 1980s. The key components of PCR are described, including DNA template, primers, DNA polymerase, nucleotides, and a thermal cycler. The basic steps of PCR are explained as denaturation, annealing and extension, which are repeated in cycles to exponentially amplify the target DNA sequence. Various applications and types of PCR are also outlined, along with its advantages of being fast, sensitive and not requiring radioactivity, though it can be prone to contamination.
PCR (polymerase chain reaction) is a technique used to amplify a single copy of a DNA segment across orders of magnitude, generating thousands to millions of copies. It involves repeated cycles of heating and cooling of the DNA sample to separate and copy the DNA strands. The key components are DNA primers, a DNA polymerase enzyme, nucleotides, and a thermocycler. During each cycle, the DNA is denatured, the primers anneal to the DNA, and the polymerase extends the primers to copy the DNA. This process is repeated many times to exponentially amplify the target DNA segment. PCR is a widely used technique in research and clinical labs due to its speed, low cost, and sensitivity.
DNA amplification techniques include in vivo cloning and in vitro PCR. PCR was independently proposed in the 1970s and 1980s and allows selective amplification of DNA segments using a thermostable DNA polymerase. Key components of PCR include a template DNA, primers, DNA polymerase, nucleotides, and magnesium. During cycling, the DNA is denatured, primers anneal, and the polymerase extends the DNA. PCR has revolutionized molecular biology due to its ability to rapidly amplify specific DNA regions.
The document discusses polymerase chain reaction (PCR), including its basic components, procedure, and applications. PCR is a technique used to amplify DNA sequences. It involves repeated cycles of heating and cooling of the DNA sample to separate and copy the DNA strands. The key steps are initial denaturation, denaturation, annealing of primers, and extension of new strands by DNA polymerase. PCR can generate millions of copies of target DNA sequences and is widely used for applications like infectious disease diagnosis, genetic testing, forensics, and molecular biology research. Recent developments include using magneto-plasmonic nanoparticles to develop nanoPCR for faster COVID-19 diagnosis within 20 minutes at the point-of-care.
Polymerase chain reaction (PCR) was invented in 1984 by Kary Mullis and revolutionized biological research. PCR amplifies a specific DNA sequence using heat-stable DNA polymerase and primers. The process involves denaturation of DNA, annealing of primers, and extension of primers to exponentially amplify the target sequence. Variations of PCR include real-time PCR, reverse transcription PCR, asymmetric PCR, and site-directed mutagenesis PCR, which are used for applications such as detecting pathogens and inducing mutations.
The document describes a lecture on polymerase chain reaction (PCR). PCR is a technique used to amplify a specific segment of DNA across several orders of magnitude, generating thousands to millions of copies of a particular DNA sequence. The key components required for PCR are a DNA template, primers, DNA polymerase, dNTPs, buffer and magnesium chloride. The process involves repeated cycles of heating and cooling of the reaction to denature and extend DNA. Applications of PCR include DNA fingerprinting, prenatal diagnosis of genetic diseases, diagnosis of viral infections, and studying ancient DNA samples.
The advent of the polymerase chain reaction (PCR) radically transformed biological science from the time it was first discovered (Mullis, 1990). For the first time, it allowed for specific detection and production of large amounts of DNA. PCR-based strategies have propelled huge scientific endeavors such as the Human Genome Project. The technique is currently widely used by clinicians and researchers to diagnose diseases, clone and sequence genes, and carry out sophisticated quantitative and genomic studies in a rapid and very sensitive manner. One of the most important medical applications of the classical PCR method is the detection of pathogens. In addition, the PCR assay is used in forensic medicine to identify criminals. Because of its widespread use, it is important to understand the basic principles of PCR and how its use can be modified to provide for sophisticated analysis of genes and the genome
This document discusses the polymerase chain reaction (PCR) technique. It begins by defining PCR and explaining its basic principles, which involve using DNA polymerase to amplify a targeted segment of DNA. It then describes the three main steps of PCR - denaturation, annealing of primers, and extension. The document provides details on how these steps are used to exponentially amplify DNA copies. It also discusses applications of PCR such as diagnosing diseases, genetic testing, forensics, and research.
4. Brief introduction to Polymerase Chain Reaction.pptxHarshadaa bafna
The document provides an introduction to polymerase chain reaction (PCR), which is a technique for amplifying specific DNA sequences. PCR allows for generating billions of copies of a target DNA fragment. It involves repeated cycles of denaturation, annealing of primers to the DNA templates, and extension of the primers by DNA polymerase. This process amplifies the target DNA exponentially. Key requirements for PCR are a DNA template, primers, DNA polymerase, and nucleotides. Automated PCR machines allow for rapid and precise amplification through repeated temperature changes. PCR has many applications in medicine, infectious disease detection, forensics, and molecular genetics research.
The document discusses polymerase chain reaction (PCR), its history, the basic steps and components involved. PCR is a technique used to amplify a specific region of DNA through repeated cycles of heating and cooling. It allows for exponential amplification of DNA, enabling small amounts of genetic material to be analyzed. The document outlines the key aspects of setting up a PCR reaction and factors important for optimal results such as primer design, annealing temperature and polymerase used. Several types of PCR are also described briefly, including real-time PCR, asymmetric PCR and nested PCR.
The document discusses various types of polymerase chain reaction (PCR) techniques. It begins by explaining what PCR is and how it works to exponentially amplify DNA sequences. It then covers the history of PCR's invention and describes the basic components and steps of a PCR reaction. The document proceeds to discuss different PCR techniques like real-time PCR, asymmetric PCR, colony PCR, and nested PCR. It concludes by noting some applications and limitations of PCR.
Polymerase chain reaction (PCR) is a technique used to amplify a specific region of DNA across multiple cycles. It involves denaturing the double-stranded DNA target, annealing primers to the single strands, and extending the primers with a DNA polymerase to synthesize new strands. Repeating this process results in exponential amplification of the target DNA sequence. PCR requires a DNA template, primers, nucleotides, and a thermostable DNA polymerase. It is used for applications like prenatal diagnosis of genetic diseases, detection of infectious diseases, cancer diagnosis, and forensics.
this section helps students how to prepare master mix solution and how to pcr. specially life life science fields such as biotechnology, biology, and medical laboratory
Polymerase chain reaction (PCR) is a laboratory technique used to amplify specific DNA sequences. It involves repeated cycles of heating and cooling of the DNA sample in the presence of primers and DNA polymerase. During each cycle, the DNA strands are separated by heating, then primers allow the polymerase to make copies of the target sequence. This results in exponential amplification, producing millions of copies of the target DNA. PCR is commonly used in research, forensics, medicine and many other applications to detect and analyze DNA.
Gene cloning and polymerase chain reaction Abhay jha
In these you are able to know about the gene cloning basic steps and Polymerase chain reaction process also there is an brief description about the ideal property shown by vectors which are lambda and M13 phases and there are lots of things in these slides
Polymerase chain reaction is a technique used in molecular biology to amplify a single copy or a few copies of a segment of DNA across several orders of magnitude, generating thousands to millions of copies of a particular DNA sequence
Pcr, Polymerase chain reaction principle of PCR, #PCRRAHUL SINWER
Polymerase chain reaction (PCR) is a technique used to amplify a specific region of DNA across multiple cycles. It involves denaturing DNA into single strands, annealing primers to the target sequence, and extending the primers with a DNA polymerase. Each cycle doubles the amount of target DNA. PCR can generate billions of copies of the target sequence, allowing it to be analyzed. It is used in various applications including DNA cloning, diagnosis, forensics, and sequencing.
PCR (polymerase chain reaction) is a technique used to amplify a specific sequence of DNA. It involves cycling between heating and cooling steps to denature and copy the DNA. During each cycle, the amount of target DNA doubles, allowing millions of copies to be produced in a few hours. It uses primers that are complementary to the target sequence and a thermostable DNA polymerase to copy the target. The basic steps involve denaturing the DNA, annealing the primers, and extending the primers to copy the target. Nested PCR and other variations allow amplification of rare sequences or detection of gene expression.
Similar to Multiple displacement amplification (20)
Candidate young stellar objects in the S-cluster: Kinematic analysis of a sub...Sérgio Sacani
Context. The observation of several L-band emission sources in the S cluster has led to a rich discussion of their nature. However, a definitive answer to the classification of the dusty objects requires an explanation for the detection of compact Doppler-shifted Brγ emission. The ionized hydrogen in combination with the observation of mid-infrared L-band continuum emission suggests that most of these sources are embedded in a dusty envelope. These embedded sources are part of the S-cluster, and their relationship to the S-stars is still under debate. To date, the question of the origin of these two populations has been vague, although all explanations favor migration processes for the individual cluster members. Aims. This work revisits the S-cluster and its dusty members orbiting the supermassive black hole SgrA* on bound Keplerian orbits from a kinematic perspective. The aim is to explore the Keplerian parameters for patterns that might imply a nonrandom distribution of the sample. Additionally, various analytical aspects are considered to address the nature of the dusty sources. Methods. Based on the photometric analysis, we estimated the individual H−K and K−L colors for the source sample and compared the results to known cluster members. The classification revealed a noticeable contrast between the S-stars and the dusty sources. To fit the flux-density distribution, we utilized the radiative transfer code HYPERION and implemented a young stellar object Class I model. We obtained the position angle from the Keplerian fit results; additionally, we analyzed the distribution of the inclinations and the longitudes of the ascending node. Results. The colors of the dusty sources suggest a stellar nature consistent with the spectral energy distribution in the near and midinfrared domains. Furthermore, the evaporation timescales of dusty and gaseous clumps in the vicinity of SgrA* are much shorter ( 2yr) than the epochs covered by the observations (≈15yr). In addition to the strong evidence for the stellar classification of the D-sources, we also find a clear disk-like pattern following the arrangements of S-stars proposed in the literature. Furthermore, we find a global intrinsic inclination for all dusty sources of 60 ± 20◦, implying a common formation process. Conclusions. The pattern of the dusty sources manifested in the distribution of the position angles, inclinations, and longitudes of the ascending node strongly suggests two different scenarios: the main-sequence stars and the dusty stellar S-cluster sources share a common formation history or migrated with a similar formation channel in the vicinity of SgrA*. Alternatively, the gravitational influence of SgrA* in combination with a massive perturber, such as a putative intermediate mass black hole in the IRS 13 cluster, forces the dusty objects and S-stars to follow a particular orbital arrangement. Key words. stars: black holes– stars: formation– Galaxy: center– galaxies: star formation
Discovery of An Apparent Red, High-Velocity Type Ia Supernova at 𝐳 = 2.9 wi...Sérgio Sacani
We present the JWST discovery of SN 2023adsy, a transient object located in a host galaxy JADES-GS
+
53.13485
−
27.82088
with a host spectroscopic redshift of
2.903
±
0.007
. The transient was identified in deep James Webb Space Telescope (JWST)/NIRCam imaging from the JWST Advanced Deep Extragalactic Survey (JADES) program. Photometric and spectroscopic followup with NIRCam and NIRSpec, respectively, confirm the redshift and yield UV-NIR light-curve, NIR color, and spectroscopic information all consistent with a Type Ia classification. Despite its classification as a likely SN Ia, SN 2023adsy is both fairly red (
�
(
�
−
�
)
∼
0.9
) despite a host galaxy with low-extinction and has a high Ca II velocity (
19
,
000
±
2
,
000
km/s) compared to the general population of SNe Ia. While these characteristics are consistent with some Ca-rich SNe Ia, particularly SN 2016hnk, SN 2023adsy is intrinsically brighter than the low-
�
Ca-rich population. Although such an object is too red for any low-
�
cosmological sample, we apply a fiducial standardization approach to SN 2023adsy and find that the SN 2023adsy luminosity distance measurement is in excellent agreement (
≲
1
�
) with
Λ
CDM. Therefore unlike low-
�
Ca-rich SNe Ia, SN 2023adsy is standardizable and gives no indication that SN Ia standardized luminosities change significantly with redshift. A larger sample of distant SNe Ia is required to determine if SN Ia population characteristics at high-
�
truly diverge from their low-
�
counterparts, and to confirm that standardized luminosities nevertheless remain constant with redshift.
ESA/ACT Science Coffee: Diego Blas - Gravitational wave detection with orbita...Advanced-Concepts-Team
Presentation in the Science Coffee of the Advanced Concepts Team of the European Space Agency on the 07.06.2024.
Speaker: Diego Blas (IFAE/ICREA)
Title: Gravitational wave detection with orbital motion of Moon and artificial
Abstract:
In this talk I will describe some recent ideas to find gravitational waves from supermassive black holes or of primordial origin by studying their secular effect on the orbital motion of the Moon or satellites that are laser ranged.
JAMES WEBB STUDY THE MASSIVE BLACK HOLE SEEDSSérgio Sacani
The pathway(s) to seeding the massive black holes (MBHs) that exist at the heart of galaxies in the present and distant Universe remains an unsolved problem. Here we categorise, describe and quantitatively discuss the formation pathways of both light and heavy seeds. We emphasise that the most recent computational models suggest that rather than a bimodal-like mass spectrum between light and heavy seeds with light at one end and heavy at the other that instead a continuum exists. Light seeds being more ubiquitous and the heavier seeds becoming less and less abundant due the rarer environmental conditions required for their formation. We therefore examine the different mechanisms that give rise to different seed mass spectrums. We show how and why the mechanisms that produce the heaviest seeds are also among the rarest events in the Universe and are hence extremely unlikely to be the seeds for the vast majority of the MBH population. We quantify, within the limits of the current large uncertainties in the seeding processes, the expected number densities of the seed mass spectrum. We argue that light seeds must be at least 103 to 105 times more numerous than heavy seeds to explain the MBH population as a whole. Based on our current understanding of the seed population this makes heavy seeds (Mseed > 103 M⊙) a significantly more likely pathway given that heavy seeds have an abundance pattern than is close to and likely in excess of 10−4 compared to light seeds. Finally, we examine the current state-of-the-art in numerical calculations and recent observations and plot a path forward for near-future advances in both domains.
BIRDS DIVERSITY OF SOOTEA BISWANATH ASSAM.ppt.pptxgoluk9330
Ahota Beel, nestled in Sootea Biswanath Assam , is celebrated for its extraordinary diversity of bird species. This wetland sanctuary supports a myriad of avian residents and migrants alike. Visitors can admire the elegant flights of migratory species such as the Northern Pintail and Eurasian Wigeon, alongside resident birds including the Asian Openbill and Pheasant-tailed Jacana. With its tranquil scenery and varied habitats, Ahota Beel offers a perfect haven for birdwatchers to appreciate and study the vibrant birdlife that thrives in this natural refuge.
Mending Clothing to Support Sustainable Fashion_CIMaR 2024.pdfSelcen Ozturkcan
Ozturkcan, S., Berndt, A., & Angelakis, A. (2024). Mending clothing to support sustainable fashion. Presented at the 31st Annual Conference by the Consortium for International Marketing Research (CIMaR), 10-13 Jun 2024, University of Gävle, Sweden.
Mechanisms and Applications of Antiviral Neutralizing Antibodies - Creative B...Creative-Biolabs
Neutralizing antibodies, pivotal in immune defense, specifically bind and inhibit viral pathogens, thereby playing a crucial role in protecting against and mitigating infectious diseases. In this slide, we will introduce what antibodies and neutralizing antibodies are, the production and regulation of neutralizing antibodies, their mechanisms of action, classification and applications, as well as the challenges they face.
PPT on Sustainable Land Management presented at the three-day 'Training and Validation Workshop on Modules of Climate Smart Agriculture (CSA) Technologies in South Asia' workshop on April 22, 2024.
2. Introduction
Multiple displacement amplification is a non-pcr based
amplification technique.
It does not require multiple cycles for the amplification of
template DNA.
An specific type of DNA polymerase is required i.e Φ29 DNA
polymerase.
Even minute quantity of DNA from single cell can be amplified
with high fidelity rate.
Whole amplification process requires a constant temperature of
30°C .
3. Φ29 DNA polymerase
Φ29 DNA polymerase is obtained from bacteriophage Φ29.
It has been extensively used in the field of molecular
biology.
It comprises of two domains:
A C-terminal domain (polymerase domain).
An N-terminal domain (3'-5’ exonuclease domain).
As compared to other type of polymerases, It has a higher
processitivity and proofreading ability.
8. Pros :
It provides higher processitivity , More than 70 kb of DNA
can be obtained.
Higher fidelity rate, Φ29 DNA polymerase has a 3’–5'
proofreading activity that provides amplification error rate
to 1 in 106−107 bases compared to
conventional Taq polymerase.
DNA from a single cell can be used for amplification.
Does not require a thermo cycler .
9. Cons :
Over amplification of a certain region may occur.
Primer dimer formation is common occurrence.
Allelic Dropout may occur in case of heterozygous sample.
Requires constant temperature.
10. Applications of MDA
Single cell genome sequencing.
Forensic Analysis.
SNP genotyping
Southern Blotting etc.