Introduction to novel antibody sequencing technique
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Antibody sequencing is a technique that determines the amino acid sequence of an antibody. Discovering the structures and functions of antibodies in living organism is an important tool in understanding cellular processes, which allow drugs that target specific metabolic pathways to be invented more easily.
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Genome editing uses engineered nucleases to insert, delete, or replace sections of the genome. CRISPR/Cas9 is a popular genome editing technique that uses guide RNA to direct nucleases to specific DNA sequences. While promising for treating disease, human germline editing raises ethical concerns. Early studies editing human embryos and non-viable embryos demonstrated proof-of-concept but had low efficiencies and off-target mutations. Later studies improved targeting and showed potential for correcting genetic defects. However, regulation is needed as the first claimed use of CRISPR to genetically edit human babies was unapproved.
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Genome editing is a type of genetic engineering in which DNA is inserted, replaced, or removed from a genome using artificially engineered nucleases, or "molecular scissors”. Identified CRISPR-associated protein 9 from Streptococcus pyogenes (Cas9) in (2012). Genome engineering by CRISPR/Cas system (16 February 2012). New application of this technology to combating viral infection by destroying invading virus DNA has now become possible in plants.
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CRISPR Cas9 is a genome editing technology that allows genetic material to be added, removed, or altered from a genome. It originated as a bacterial immune system but can now be directed to make precise edits to DNA. The technology has wide applications for gene therapy, agriculture, research, and more, but also raises ethical concerns if misused. CRISPR offers promising possibilities but also challenges that must be addressed regarding safety, accuracy, and societal effects.
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Genome editing allows specific changes to be made to the DNA of cells and organisms. Key techniques include zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the CRISPR-Cas9 system. ZFNs and TALENs use a DNA-binding domain fused to a nuclease to cut the DNA at a targeted location. CRISPR-Cas9 uses a guide RNA to target the Cas9 nuclease. Double-strand breaks caused by these nucleases can be repaired by non-homologous end joining or homology-directed repair, allowing genes to be knocked out, knocked down, or replaced. Genome editing holds promise for applications like disease
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Genome editing allows specific changes to be made to the DNA of cells and organisms. Key techniques include zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the CRISPR-Cas9 system. ZFNs and TALENs use a DNA-binding domain fused to a nuclease to cut the DNA at a targeted location. CRISPR-Cas9 uses a guide RNA to target the Cas9 nuclease. Double-strand breaks caused by these nucleases can be repaired by non-homologous end joining or homology-directed repair, allowing genes to be knocked out, knocked down, or replaced. Genome editing holds promise for applications like disease
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CRISPR, CAS9, Genome Editing, ZFNS, TALENS, Spacer DNA, tracrRNA, crRNA, sgRNA, PAM, CRISPAR-CAS9, CAS GENES, Gene editing, gene silencing, gene knockout, CAS protein, enzyme, nuclease, restriction enzyme, CRISPR Array, Genetic Engineering, Applications of CRISPR, Advantages of CRISPR, LIMITATIONS OF CRISPR, DISADVANTAGES OF CRISPR, BENEFITS OF CRISPR, Future of CRISPR, CRISPR and Cancer, Ethical Issues.
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Cas9 (CRISPR-associated protein 9) is an enzyme that uses CRISPR sequences as a guide to recognize and cleave specific strands of DNA that are complementary to the CRISPR sequence. Cas9 enzymes together with CRISPR sequences form the basis of a technology known as CRISPR-Cas9 that can be used to edit genes within organisms.This editing process has a wide variety of applications including basic biological research, development of biotechnology products, and treatment of diseases.
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The most talked about gene editing tool- CRISPR Cas9 and its applications in all the possible spheres of science and research is talked about in brief in this presentation.
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Gene therapy involves introducing genes into cells to treat disease. CRISPR is a gene editing technique using Cas9 protein and guide RNA. It allows DNA cutting and modification. Luxturna is a gene therapy for an inherited eye disease caused by RPE65 gene mutations. It uses AAV vector to insert a healthy RPE65 gene into retinal cells. In clinical trials, Luxturna improved patients' visual function and mobility in low light after 1 year as measured by a mobility test.
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The last decade has seen the fields of molecular biology and genetics transformed by the development of CRISPR-based gene editing technologies. These technologies were derived from bacterial defense systems that protect against viral invasion. In the past few years, a variety of phages and other mobile genetic elements have been shown to encode anti-CRISPR proteins (Acrs) that interact directly (in a sequence-independent manner) with components of the CRISPR-Cas system and inactivate it. The discovery of anti-CRISPR proteins has opened up a new area of phage research and has provided a valuable addition to the CRISPR toolbox as an ‘off switch’’ for Cas9 activity. But, most of the CRISPR-Cas systems still have no known inhibitors, suggesting that many anti-CRISPR protein families are yet to be discovered which can be used as regulators for genome engineering and other biotechnological applications.
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The document summarizes key aspects of CRISPR genome editing technology. It describes how CRISPR uses Cas9 and guide RNA to precisely target and edit DNA sequences. It provides a brief history of CRISPR discovery and outlines its components and mechanism of action. The document also discusses several medical applications of CRISPR including treating Duchenne muscular dystrophy, beta-thalassemia, and testing for viruses. It concludes that CRISPR is a flexible and accurate gene editing tool being explored for various applications in agriculture, biotechnology, and medicine.
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The document summarizes a seminar on the role of CRISPR-Cas9 in antimicrobial resistance. It first describes the components and mechanism of the CRISPR-Cas9 system, including how it was discovered and can be used for gene editing. It then discusses how CRISPR-Cas9 can be programmed to target and cleave DNA sequences, including antibiotic resistance genes. Specifically, a CRISPR-Cas9 system can be built into a bacteriophage to target and disrupt bacteria carrying resistance genes. Studies have shown this approach can successfully target genes conferring resistance to antibiotics like carbapenem, kanamycin, ceftazidime, and methicillin.
CRISPR-Cas system
CRISPR (clustered regularly interspaced short palindromic repeats) is a family of DNA sequences found within the genomes of prokaryotic organisms such as bacteria and archaea. These sequences are derived from DNA fragments of bacteriophages that have previously infected the prokaryote and are used to detect and destroy DNA from similar phages during subsequent infections. Hence these sequences play a key role in the antiviral defense system of prokaryotes.
Cas9 (CRISPR-associated protein 9) is an enzyme that uses CRISPR sequences as a guide to recognize and cleave specific strands of DNA that are complementary to the CRISPR sequence. Cas9 enzymes together with CRISPR sequences form the basis of a technology known as CRISPR-Cas9 that can be used to edit genes within organisms.This editing process has a wide variety of applications including basic biological research, development of biotechnology products, and treatment of diseases.
The CRISPR-Cas system is a prokaryotic immune system that confers resistance to foreign genetic elements such as those present within plasmids and phages that provides a form of acquired immunity. RNA harboring the spacer sequence helps Cas (CRISPR-associated) proteins recognize and cut foreign pathogenic DNA. Other RNA-guided Cas proteins cut foreign RNA. CRISPR are found in approximately 50% of sequenced bacterial genomes and nearly 90% of sequenced archaea.
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We use an error-prone PCR integrated DNA-shuffling approach to mutate mainly CDR regions during sub-library construction. If the potential of introducing immunogenic mutations to framework positions is not a concern, we usually use this approach to create mutations at completely random positions across the entire VH and VL fragments. In these cases, the genetic diversity of the sub-library is further increased via passage through our proprietary bacterial mutator strain, CD-affi™.
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This document summarizes a presentation on CRISPR/Cas genome editing. It defines CRISPR/Cas as a type of genetic engineering that uses artificially engineered nucleases to make specific cuts in DNA. It describes the CRISPR/Cas system's origins and components, including Cas9, guide RNA, and PAM sequences. Applications discussed include genome editing in animals and plants, as well as concerns over off-target effects. Companies offering CRISPR services or kits are also mentioned.
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CRISPR-Cas9 is a unique technology that enables geneticists and medical researchers to edit parts of the genome by removing, adding or altering sections of the DNA sequence.
It is currently the simplest, most versatile and precise method of genetic manipulation and is therefore causing a buzz in the science world.
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Many bacterial clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated (Cas) systems employ the dual RNA–guided DNA endonuclease Cas9 to defend against invading phages and conjugative plasmids by introducing site-specific double-stranded breaks in target DNA. Target recognition strictly requires the presence of a short protospacer adjacent motif (PAM) flanking the target site, and subsequent R-loop formation and strand scission are driven by complementary base pairing between the guide RNA and target DNA, Cas9–DNA interactions, and associated conformational changes. The use of CRISPR–Cas9 as an RNA-programmable
DNA targeting and editing platform is simplified by a synthetic single-guide RNA (sgRNA) mimicking the natural dual trans-activating CRISPR RNA (tracrRNA)–CRISPR RNA (crRNA) structure
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CRISPR/Cas9 gene editing shows promise for correcting diseases through gene therapy and targeted changes. Current applications include editing T-cells to produce CAR T-cells for cancer therapy and editing retinal cells in rats to treat retinitis pigmentosa. Additional areas of focus include eliminating HIV from cells, correcting the Fah mutation in mice, and overcoming issues like delivery efficiency and off-target effects. Looking ahead, cell therapies and local treatments for eye/ear diseases appear promising applications for CRISPR's precise gene editing capabilities.
CRISPR technology -A revolutionary discovery
CRISPR technology allows for genome editing using a prokaryotic immune system called CRISPR/Cas. The system works by adapting spacers from viral DNA, producing CRISPR RNA, and targeting matching sequences. It is being applied in industry to make bacterial cultures virus-resistant, in labs for genetic engineering, and in medicine for treating genetic diseases and developing more specific antibiotics.
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This document discusses various methods for mutation detection including:
- Single-stranded conformation polymorphism (SSCP) which detects differences in DNA secondary structure between mutant and wild-type sequences.
- Denaturing gradient gel electrophoresis (DGGE) which separates DNA fragments based on their melting properties.
- Oligonucleotide ligation assay (OLA) which detects mutations by hybridizing PCR primers and ligating probes.
- Restriction fragment length polymorphism (RFLP) which identifies polymorphisms by cleaving DNA with restriction enzymes.
31931 31941
This document provides an overview of Serial Analysis of Gene Expression (SAGE). SAGE allows for the digital analysis of overall gene expression patterns in a sample by producing a snapshot of the mRNA population. It provides a quantitative and comprehensive expression profile. The document outlines the key principles and steps of the SAGE methodology, including isolating mRNA, synthesizing cDNA, ligating linkers, releasing tags, concatenating tags, and sequencing. It also discusses various applications and advances of SAGE, such as LongSAGE, CAGE, and SuperSAGE. SAGE is a powerful tool for studying gene expression, but it has some limitations regarding transcript identification and quantitation bias.
PROTEIN MICROARRAYS
it will help you to understand how the protein microarrays are made, what are the different types and what all purposes they are used for. its very useful ppt
Congreso de Biotecnología Arequipa Perú June 2011
This document summarizes research to bioengineer a new red fluorescent protein tag from a cyanobacteriochrome found in Thermosynechococcus elongatus. The researchers aim to develop a small, photostable red tag as an alternative to commonly used green and yellow fluorescent proteins from jellyfish. They use molecular biology techniques like PCR, site-directed mutagenesis, and transfection of E. coli and mammalian cells to express and purify the mutated protein, which they characterize through spectroscopy and microscopy. The goal is to create a useful new tool for cellular imaging applications.
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For research use only. Not intended for any clinical use. No products from Creative Biolabs may be resold, modified for resale or used to manufacture commercial products without prior written approval from Creative Biolabs.
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Anti-CGRP Antagonist Antibody scFv fragment G1 binds to a C-terminal epitope with F37 and G33 being the most important residues. G1 does not bind to CGRP when an extra amino acid residue (alanine) is added at the C-terminal.
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This document summarizes a recombinant monoclonal antibody that targets CEACAM5, a potential cancer treatment. It is a single domain antibody derived from a llama VHH library against the extracellular region of human CEACAM5. The antibody is intended for research use only and not for diagnostic or therapeutic human use.
30 kda protein
This product is a mouse monoclonal antibody that recognizes the 30 kDa protein of Chlamydia trachomatis. It is an IgG antibody derived from a mouse immunized with C. trachomatis and specifically reacts with the 30 kDa protein of C. trachomatis. This antibody is intended for ELISA applications and research use only.
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Recombinant Mouse Antibody (3E6) is capable of binding to Coagulation factor VIII, expressed in Chinese Hamster Ovary cells (CHO).
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This mouse monoclonal antibody binds to the 65 kDa late antigen (LA) of cytomegalovirus (CMV). It was derived from a mouse immunized with the 65 kDa CMV late antigen. Applications for this antibody include western blot, immunofluorescence, and ELISA for research purposes only.
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Recombinant Human Antibody (dAb HEL4) is capable of binding to Chicken LYZ, expressed in HEK 293 cells. Expressed as the combination of a heavy chain (HC) containing VH from anti-Chicken LYZ mAb and CH1-3 region of human IgG1 and a light chain (LC) encoding VL from anti-Chicken LYZ mAb and CL of human kappa light chain. Exists as a disulfide linked dimer of the HC and LC hetero-dimer under non-reducing condition. HEL4 is highly soluble at concentrations of> or =3 mM, essentially monomeric and resistant to aggregation upon thermodenaturation at concentrations as high as 56 microM.
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This antibody-cytokine fusion protein was achieved by conjugating/fusing the Anti-CD3 IgG to IL2. It was expressed in CHO and purified with affinity chromatography. The immunocytokine retains the ability to bind the CD3 as well as the biological activity of IL2. under in vitro conditions, IL2-FuP efficiently initiated T-cell activation and stimulated LAKs as well as CTLs. Its potency was superior but not qualitatively different from that of a mixture of anti-EGFR and IL2 This immunocytokine was designed for treating Hodgkin's lymphoma.
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Introduction to novel antibody sequencing technique
- 1. Introduction to Novel Antibody Sequencing Technique
- 2. Brief Introduction Antibody sequencing is a bio-technique that determines the amino acid sequence of an antibody. Discovering the structures and functions of antibodies in living organism is an important tool in understanding cellular processes, which allow drugs that target specific metabolic pathways to be invented more easily. This next generation antibody sequencing technique has now become a routine measurement that can be completed within 10 working days for IgGs. The method is applicable for all antibody formats, namely IgMs, fluorochrome conjugates, immobilized antibodies and mixtures. Creative Biolabs, an emerging qualified service provider in this field, provides researchers their unique technique in antibody sequencing.
- 3. With the development of biotechnology, scientists of Creative Biolabs have developed a novel strategy to determine the complete sequence of an antibody with unparalleled speed and accuracy. The technique which combines all the advantages of existing approaches is named “Database Assisted Shotgun Sequencing” (DASS). In the first step, the antibody is fragmented to peptides by a special technique, which generates up to 5000 different peptides per chain. This set of peptides will be analyzed by high end mass spectrometers to generate extensive sequence information. MS/MS spectra are then de novo sequenced by the latest algorithms and matched against a database with related sequences. Special “in house” data mining tools allow Creative Biolabs to extract the sequence information from ten thousands of MS/MS spectra within hours.
- 4. De Novo Sequencing of the CDR3 Region While the CDR3 of the light chain is mostly encoded by the germline sequences, the CDR3 of the heavy chain is usually not available in databases. It is encoded by the so called D-segments, but these are modified by nucleases and terminal transferases. Typically, only 1-4 AA of a D-segment remain in the matured antibody. The rest of the D-segment is “artificial” and has to go through de novo sequence. Their method generates many overlapping peptides during the fragmentation process, enabling them to sequence very long stretches of unknown amino acids. The high quality of MS/MS spectra in combination with intelligent data mining, allows them to read the CDR3 like a book. The technique is so powerful, that they were able to sequence a 20 kDa protein, which had no homologue in the database.
- 5. Sequencing of the V and J and C segments by "Database Assisted Shotgun Sequencing“ The V and J and C gene segments of antibodies are available in public databases. During the maturation of an antibody, the B-cell introduces hypermutations into the sequence to optimize the affinity. Creative Biolabs’s mapping algorithm is error tolerant and can match the “mutated” peptides to the corresponding germline reliably. Because of the great number of peptides, Creative Biolabs is able to get sequence information for ever bond in the antibody. Typically, 20-70 different MS/MS spectra are generated for each amino acid (AA) position. Hence, even the hardest sequences of proline and arginine rich peptides can be resolved.
- 6. E-mail: marketing@creative-biolabs.com Tel: 1-631-619-7922 Website: http://www.creative-biolabs.com
- 7. E-mail: marketing@creative-biolabs.com Tel: 1-631-619-7922 Website: http://www.creative-biolabs.com