Colony blot hybridization pptx
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Colony blot hybridization is a procedure where colonies grown on a nutrient medium are transferred to a membrane, the cells are lysed to release DNA or protein, and the membrane is treated to allow binding of the DNA or protein. The membrane can then be exposed to a hybridization probe to identify colonies containing a specific nucleic acid sequence or an antibody to identify colonies containing a specific protein. The procedure involves blotting microbial colonies onto a membrane, lysing the cells to release nucleic acids, fixing the DNA or RNA to the membrane, hybridizing it with a labeled probe, washing away unbound probe, and visualizing the results. Colony blot hybridization can be used to screen clones or bacterial isolates.
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History
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Yeast Artificial Chromosome (BAC)
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Hybridization techniques involve immobilizing target DNA/RNA on a membrane and probing it with a labeled DNA/RNA probe. The key steps are immobilizing the target nucleic acid, preparing a labeled probe with complementary sequence, hybridizing the probe to the target, and detecting the hybridized probe. Membrane hybridization does not proceed to completion as some nucleic acids are embedded in the membrane and inaccessible to the probe. Pre-hybridization and hybridization buffers contain components like rate enhancers and blocking agents to promote hybridization while preventing non-specific binding.
Blotting techniques in Biotechnology
This document provides an overview of blotting techniques, including transfer methods, membranes used, and applications of Southern blotting, Northern blotting, and Western blotting. It discusses how blotting involves transferring proteins, DNA, or RNA from a gel onto a membrane. It describes the capillary, electro-blotting, and vacuum blot transfer processes and membranes like nitrocellulose, nylon, and PVDF. It then explains the specific methods and applications of Southern blotting for detecting DNA, Northern blotting for detecting RNA, and Western blotting for detecting proteins.
Blotting techniques
These slides include all the 3 blotting techniques i,e Southern,northern and western.Their principle,procedure and application.
Monoclonal antibodies production(1)
The document describes the hybridoma technique for producing monoclonal antibodies. It involves immunizing an animal with an antigen, fusing the animal's antibody-producing B cells with myeloma cells to create hybridomas, and using HAT selection medium to select hybridomas that produce the desired monoclonal antibody. The hybridomas are then screened using ELISA or radioimmunoassay to identify clones that secrete antibodies targeting the specific antigen. Clones that test positive can then be cloned using limiting dilution or soft agar methods to generate stable monoclonal antibody-producing cell lines. Monoclonal antibodies have various applications in areas like diagnostics, imaging, pregnancy testing, and organ transplantation.
Blotting techniques
This document provides information on immunoblotting techniques such as Western blotting. It discusses the basic workflow of Western blotting which includes sample preparation, gel electrophoresis, protein transfer, blocking and antibody probing, detection, and analysis. It also summarizes other blotting techniques like Southern blot which is used to detect specific DNA sequences. The applications of Western and Southern blotting in research and clinical diagnosis are also briefly mentioned.
Multiplex analysis as tools in Biological science research
Multiplex tools- Microarray techniques / Microassay techiniques and Multidimentional Liquid Chromatography
unit 4 immunoblotting technique complete.pptx
Immunoblotting techniques such as Western blotting are used to detect specific proteins separated by electrophoresis using labeled antibodies. Western blotting involves separating proteins by gel electrophoresis, transferring them to a membrane, and using antibodies to detect the target protein bands on the membrane. ELISA (enzyme-linked immunosorbent assay) is a technique used to detect the presence of antibodies, antigens, or other substances in a liquid sample using enzymes to produce a detectable color change when the target substance is present. Microbial biotransformation uses microorganisms or enzyme systems to structurally modify chemical compounds, maintaining the original carbon skeleton while increasing polarity, and is useful for applications like steroid transformation and pollutant degradation.
Blotting
A blot, in molecular biology and genetics, is a method of transferring proteins, DNA or RNA, onto a carrier (for example, a nitrocellulose, polyvinylidene fluoride (PVDF) or nylon membrane). In many instances, this is done after a gel electrophoresis, transferring the molecules from the gel onto the blotting membrane, and other times adding the samples directly onto the membrane. After the blotting, the transferred proteins, DNA or RNA are then visualized by colorant staining (for example, silver staining of proteins), autoradiographic visualization of radioactive labelled molecules (performed before the blot), or specific labelling of some proteins or nucleic acids. The latter is done with antibodies or hybridization probes that bind only to some molecules of the blot and have an enzyme joined to them. After proper washing, this enzymatic activity (and so, the molecules we search in the blot) is visualized by incubation with proper reactive, rendering either a colored deposit on the blot or a chemiluminiscent reaction which is registered by photographic film.
Southern blot
A Southern blot is a method routinely used in molecular biology for detection of a specific DNA sequence in DNA samples. Southern blotting combines transfer of electrophoresis-separated DNA fragments to a filter membrane and subsequent fragment detection by probe hybridization.[1]
Western blot
A Western blot is used for the detection of specific proteins in complex samples. Proteins are first separated by size using electrophoresis before being transferred to an appropriate blotting matrix (usually PVDF or nitrocellulose) and subsequent detection with antibodies.
Eastern blot
The Eastern blot is used for the detection of specific posttranslational modifications of proteins. Proteins are separated by gel electrophoresis before being transferred to a blotting matrix whereupon posttranslational modifications are detected by specific substrates (cholera toxin, concanavalin, phosphomolybdate, etc.) or antibodies.
Western Blotting
Western blotting is a technique used to detect specific proteins in a sample. It involves separating proteins by gel electrophoresis, transferring them to a membrane, and using antibodies to identify a target protein. The key steps are sample preparation, gel electrophoresis to separate proteins by size, transferring proteins to a membrane, blocking the membrane to reduce background noise, probing the membrane with antibodies to detect the target protein, washing unbound antibodies, and detecting the target protein to analyze its presence and quantity. Western blotting is widely used in research and medical diagnostics to study protein expression and identify proteins of interest.
Blotting techniques1
This document discusses different blotting techniques used to detect DNA, RNA, and proteins. Southern blotting is used to detect DNA and was developed by Edwin Southern in 1975. It involves separating DNA fragments by size, transferring them to a membrane, and using a probe to detect specific sequences. Northern blotting detects RNA and was developed in 1979. It similarly separates RNA and uses a probe to detect sequences of interest. Western blotting detects proteins and was developed in 1981. It separates proteins by SDS-PAGE gel electrophoresis, transfers them to a membrane, and uses an antibody probe to detect the target protein. These techniques are widely used in research and diagnostics.
blooting image.pptx
1. DNA or proteins are separated by gel electrophoresis.
2. The molecules are then transferred to a membrane through either southern or western blotting.
3. For southern blotting, DNA is detected through hybridization with a labeled probe. For western blotting, specific proteins are detected through interaction with labeled antibodies.
4. These techniques are used for diagnostic purposes and research applications such as identifying DNA sequences or proteins of interest.
IMMUNOBLOTTING.pptx
This document discusses various techniques used in immunoblotting and blotting. It begins by defining blotting as techniques used to visualize specific DNA, RNA, and proteins among contaminants. It then describes three main types of blotting - western blotting for proteins, northern blotting for RNA, and southern blotting for DNA. The document focuses on western blotting and immunoblotting. It provides details on tissue preparation, gel electrophoresis, protein transfer, blocking, detection, analysis, and applications of western blotting and immunoblotting techniques.
Western blotting
Western blotting is a technique used to detect specific proteins in a sample. It involves separating proteins by electrophoresis, transferring them to a membrane, and using antibodies to locate the target protein. The key steps are sample preparation, SDS-PAGE gel electrophoresis to separate proteins by size, transferring proteins from the gel to a membrane, blocking the membrane to prevent nonspecific antibody binding, probing the membrane with antibodies to detect the target protein, washing unbound antibodies, and detecting the bound antibodies to analyze and image the results. Western blotting allows identification and quantification of proteins in a given sample.
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Selection & Screening of Recombinant cells & expression of recombinant (2) (1)
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Multiplex analysis as tools in Biological science research
Multiplex analysis as tools in Biological science research
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Orders of class osteichthyes
This document summarizes the key characteristics of 18 orders of bony fishes (class Osteichthyes) based on their anatomical features and examples. It describes traits such as fin structure, scale type, presence of swim bladders and their connections to other organs, body shape adaptations, and example fish from each order. The orders discussed include Crossopterygii, Dipnoi, Polypteriformes, Acipenseriformes, and others.
Mammals stomach and dentition
The document discusses the origin and evolution of mammals. It describes two main theories for the ancestry of mammals - descent from amphibians or reptiles. While Huxley proposed an amphibian ancestry, most evidence supports a reptilian ancestry, with mammals evolving from mammal-like reptiles called therapsids. True mammals first appeared in the Jurassic period and were small, nocturnal creatures. After the extinction of dinosaurs, mammals underwent an adaptive radiation, diversifying to fill new ecological niches. Limbs and teeth evolved adaptations for different diets and modes of locomotion like burrowing, swimming and climbing. Distantly related groups sometimes converged on similar forms when occupying similar habitats.
Origin and Evolution of Mammals
The document discusses the origin and evolution of mammals. It describes two main theories for the ancestry of mammals - through amphibians or reptiles. While amphibian ancestry was proposed, reptilian ancestry is now widely accepted based on fossil evidence. Many characteristics of early mammal-like reptiles called therapsids were mammalian. True mammals first appeared in the Jurassic period but remained small until after the extinction of dinosaurs. When ecological niches opened up in the Cenozoic era, mammals underwent adaptive radiations into various forms through modifications of limbs, teeth and other features for different habitats like trees, ground, burrows, water and air. Convergent evolution also led to similarities between unrelated mammals adapting to the same nic
Draco volans
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Digestive system of vertebrates
1. The document describes the digestive system of vertebrates, including the development and anatomy of key structures like the mouth, pharynx, esophagus, stomach, intestines, liver and gallbladder.
2. It notes that the digestive tract develops from the embryonic archenteron and surrounding mesoderm. Structures vary between aquatic and terrestrial vertebrates.
3. Accessory organs like the tongue, teeth and salivary glands are described. The stomach and intestines show diversity between species related to diet. The liver and gallbladder function to produce and store bile.
Common peafowl
1. The document discusses the common peafowl, specifically the Indian peafowl. It describes the classification, appearance, habitat, and distribution of male peacocks and female peahens.
2. Male peacocks have iridescent blue-green plumage and elongated tail feathers called a "train" with eye-like patterns. Females are duller in coloration and lack the elongated tail feathers.
3. Peafowl are found in forests, farmland, bushlands and rainforests, preferring areas with open spaces for displaying and dust bathing. Their historic range was northern India but they are now protected in several national parks and reserves in Nepal.
Brain of vertebrates
The document summarizes the development and comparative anatomy of the vertebrate brain. It describes how the embryonic brain divides into three primary vesicles - the prosencephalon, mesencephalon, and rhombencephalon. It then discusses the specific structures that develop from each of these regions across different vertebrate groups, including differences in brain structure between cyclostomes, fish, amphibians, reptiles, birds, and mammals. Key differences include the size of regions like the cerebrum, cerebellum, and olfactory bulbs across the groups.
Integument
This document discusses the comparative anatomy of the integument and its derivatives in vertebrates. It describes the general structure of skin, which consists of an outer epidermis and inner dermis layers. The epidermis is stratified squamous epithelium that provides protection, while the dermis contains connective tissue, blood vessels, and sensory structures. Major derivatives of the integument discussed include epidermal glands, scales, claws/nails/hooves, horns, feathers, and hair. The integument and its derivatives have evolved to suit the environment and provide functions like protection, thermoregulation, sensation, and identification.
Water
Water is essential for life and exists in three states: liquid, solid, and gas. It has unique properties like being a universal solvent and exhibiting surface tension due to hydrogen bonding between molecules. Water is used for many important purposes including household use, agriculture, industry, recreation, hydropower, and more. As the global population increases, proper management of water resources and conservation efforts will be needed to ensure adequate fresh water supplies for the future.
Translation
Translation is the process by which the genetic code in mRNA is used to direct the synthesis of proteins. It involves three main steps - initiation, elongation, and termination. Initiation requires the small and large ribosomal subunits to assemble around an mRNA molecule along with initiator tRNA and other initiation factors. Elongation then adds amino acids one by one to the growing polypeptide chain according to the mRNA codons. Termination occurs when a stop codon is reached, causing the ribosome to dissociate and release the complete protein.
The structure of dna and genome ogranization
The document discusses the structure of DNA and genome organization. It describes that DNA is usually composed of two polynucleotide chains twisted around each other in a double helix structure. The backbone of each strand is made up of alternating sugar and phosphate residues, while the bases project inward. The two strands are linked by hydrogen bonds between complementary bases, with adenine pairing with thymine and guanine pairing with cytosine. Together, the sugar, phosphate, and base components make up the nucleotides that serve as the fundamental building blocks of DNA.
Plasmodium
This document discusses the life cycle and pathogenesis of Plasmodium vivax, the parasite that causes vivax malaria. P. vivax has a digenetic life cycle involving two hosts. In humans, it undergoes asexual reproduction in the liver and red blood cells. Mosquitoes ingest the parasite's gametocyte stages, where sexual reproduction occurs leading to sporozoite formation. Sporozoites are then transmitted to humans to continue the cycle, causing symptoms of malaria including fever and anemia. Diagnosis involves blood smear microscopy and treatment involves chloroquine and primaquine to prevent relapses.
Comparative study of vertebrates
The document describes the development and anatomy of the human eye and compares it to eyes in other organisms. It discusses that in the third week of human development, optic vesicles form and later develop into the optic cup, which will become the retina. By week six, tissues surrounding the eye differentiate into the sclera and choroid. The document then compares key features of eyes across vertebrate groups like fish, amphibians, reptiles, birds, and mammals.
The nucleic acids
Friedrich Miescher first isolated nucleic acids in 1869 and called them "nuclein" due to their acidic properties. The discovery of DNA's double helix structure in 1953 by Watson and Crick was monumental. Nucleic acids function to store and transmit genetic information through DNA and RNA. DNA is made of two strands bound together by complementary nucleotide bases, with adenine pairing with thymine and guanine pairing with cytosine. The discovery of DNA's structure explained the mechanism of heredity.
Honey bee
This document provides information about honey bees. It begins with the classification of honey bees, then discusses their social behavior and advantages. It provides examples of social insects including honey bees, wasps, termites and ants. The document focuses on honey bees, describing their appearance, distribution, and the different types of bees within a hive: the queen, workers, and drones. It explains the key roles and characteristics of each type of honey bee.
Genetic engineering of humans
This document discusses genetic engineering of humans through various methods such as germline genetic modification and somatic genetic modification. It notes both the promising applications for improving health through gene therapy but also the ethical concerns regarding enhancing traits and the possibility of genetic inequality. Specific examples discussed include using CRISPR to genetically modify human embryos to potentially treat disease as well as concerns about "designer babies". The document examines issues around testing for non-disease traits, gene doping in athletes, and the potential benefits and risks of human gene editing technologies.
Hazards of genetic engineering
Genetic engineering and genetically modified organisms have potential risks and downsides according to the document. While genetic engineering has applications in agriculture and medicine, it can also have detrimental effects on humanity. Introducing foreign genes into other species can have uncontrolled and unpredictable consequences. Studies have shown genetically modified foods can be toxic and cause health issues in animals. Genetically engineered crops also have negative impacts on biodiversity and the environment by promoting the growth of pesticide-resistant superweeds and superbugs. The long-term effects of genetic engineering are uncertain due to its new and crude nature, making it a controversial technology that manipulates nature in ways bypassing normal evolution.
Azotobacter spp.
The document describes isolating and identifying Azotobacter species from a soil sample. It discusses enriching the soil in Ashby's Mannitol Broth for a week to activate nitrogen-fixing bacteria. Colonies were then streaked on the broth and incubated for 4-6 days. Gram staining identified gram-negative cells with a red color and capsule staining showed transparent capsules around violet cells, indicating motility. The colonies were creamy white, raised, circular and mucoid, consistent with Azotobacter.
Biological membrane
The document summarizes the fluid mosaic model of the cell membrane. It describes the main components of the membrane - phospholipids, cholesterol, proteins, and carbohydrates. The phospholipid bilayer forms the main structure of the membrane, with phospholipids arranged in two layers with their hydrophobic tails facing each other and hydrophilic heads facing outwards. Proteins embedded within or attached to the membrane carry out important functions like transport and cell signaling. Carbohydrates attached to proteins and lipids on the outer surface of the membrane help with cell recognition. The fluid nature of the membrane allows it to maintain selective permeability and flexibility. Passive and active transport mechanisms allow movement of substances across the membrane.
Atomic radius
1) Atomic radius is the distance from the nucleus to the outermost shell of electrons or the point where electron density is highest.
2) There are three types of atomic radii: covalent, van der Waals, and metallic.
3) Covalent radius is half the distance between bonded atoms. Van der Waals radius is half the distance between adjacent non-bonded atoms. Metallic radius is half the distance between metal ions in a lattice.
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Receptor Discordance in Breast Carcinoma During the Course of Life
Definition:
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Causes:
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Treatment Effects:
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Acne is a common skin condition that occurs when hair follicles become clogged with oil and dead skin cells. It typically manifests as pimples, blackheads, or whiteheads, often on the face, chest, shoulders, or back. Acne can range from mild to severe and may cause emotional distress and scarring in some cases.
**Causes:**
1. **Excess Oil Production:** Hormonal changes during adolescence or certain times in adulthood can increase sebum (oil) production, leading to clogged pores.
2. **Clogged Pores:** When dead skin cells and oil block hair follicles, bacteria (usually Propionibacterium acnes) can thrive, causing inflammation and acne lesions.
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Treatment depends on the severity and type of acne but may include:
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Colony blot hybridization pptx
- 1. Colony blot hybridization By- Sanju sah St. Xavier’s college, Maitighar, Kathmandu Department of Microbiology
- 2. Introduction A procedure in which an imprint of the colonies grown on the surface of a solid nutrient medium is transferred to a membrane and the Cells lysed or disrupted, allowing the DNA or protein to bind to the membrane Following appropriate treatment, the membrane can be exposed to either a hybridization probe for the identification of colonies containing a specific nucleic acid sequence or to an antibody for the identification of colonies containing a specific protein.
- 3. Procedure • Colony blot hybridization is applied to DNA or RNA released from blotted microbial colonies. • The microbial colonies are transferred (blotted) to a membrane. • The cells are lysed in place to release the nucleic acids. • The RNA or DNA (after denaturation) is fixed to the filter and hybridized with a labelled probe. • Blocking reagent may be added prior to the probe to prevent unspecific binding. • Excess probe is washed away and the membrane is visualized by UV or autoradiography. • Colony blot hybridization can be used for screening clones or bacterial isolates.