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Off-target Effects Of Genome Editing
Off-target genome editing refers to nonspecific and unintended genetic modifications that can arise through the use of engineered nuclease technologies such as: clustered, regularly interspaced, short palindromic repeats (CRISPR)- Cas9, transcription activator-like effector nucleases (TALEN), meganucleases, and zinc finger nucleases (ZFN). These tools use different mechanisms to bind a predetermined sequence of DNA (“target”), which they cleave (or "cut"), creating a double-stranded chromosomal break (DSB) that summons the cell's DNA repair mechanisms (non-homologous end joining ( NHEJ) and homologous recombination ( HR)) and leads to site-specific modifications. If these complexes do not bind at the target, often a result of homologous sequences and/or mismatch tolerance, they will cleave off-target DSB and cause non-specific genetic modifications. Specifically, off-target effects consist of unintended point mutations, deletions, insertions inversions, and translocations ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Genome Editing
Genome editing, or genome engineering, or gene editing, is a type of genetic engineering in which DNA is inserted, deleted, modified or replaced in the genome of a living organism. Unlike early genetic engineering techniques that randomly inserts genetic material into a host genome, genome editing targets the insertions to site-specific locations. The basic mechanism involved in genetic manipulations through programmable nucleases is the recognition of target genomic loci and binding of effector DNA-binding domain (DBD), double-strand breaks (DSBs) in target DNA by the restriction endonucleases ( FokI and Cas), and the repair of DSBs through homology-directed recombination (HDR) or non-homologous end joining (NHEJ). History Genome editing was pioneered in the 1990s, before the advent of the common current nuclease-based gene editing platforms, however, its use was limited by low efficiencies of editing. Genome editing with engineered nucleases, i.e. all three major classes of ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
ChIP
Chromatin immunoprecipitation (ChIP) is a type of immunoprecipitation experimental technique used to investigate the interaction between proteins and DNA in the cell. It aims to determine whether specific proteins are associated with specific genomic regions, such as transcription factors on promoter (biology), promoters or other DNA binding sites, and possibly define cistromes. ChIP also aims to determine the specific location in the genome that various histone modifications are associated with, indicating the target of the histone modifiers. ChIP is crucial for the advancements in the field of epigenomics and learning more about epigenetic phenomena. Briefly, the conventional method is as follows: # DNA and associated proteins on chromatin in living cells or tissues are crosslinked (this step is omitted in Native ChIP). # The DNA-protein complexes (chromatin-protein) are then sheared into ~500 bp DNA fragments by sonication or nuclease digestion. # Crosslinking of DNA, Cross-linked ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Flavobacterium Okeanokoites
''Flavobacterium'' is a genus of Gram-negative, nonmotile and motile, rod-shaped bacteria that consists of 130 recognized species. Flavobacteria are found in soil and fresh water in a variety of environments. Several species are known to cause disease in freshwater fish. '' Flavobacterium psychrophilum'' causes the bacterial cold water disease on salmonids and the rainbow trout fry disease on rainbow trout. '' F. columnare'' causes the cotton-wool disease on freshwater fishes. '' F. branchiophilum'' causes the bacterial gill disease on trout. Another member of this genus, '' F. okeanokoites'' is the original source for the type IIs restriction endonuclease ''Fok''I, used in Zinc finger nucleases and TALENs. Nylon-eating bacteria are a strain of ''Flavobacterium'' that is capable of digesting certain by-products of nylon 6 manufacture. Species who are a part of the genus ''Flavobacterium'' are most likely found scattered along in nature. These microbes are mostly found in a ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
FokI
The restriction endonuclease ''Fok''1, naturally found in ''Flavobacterium okeanokoites'', is a bacterial type IIS restriction endonuclease consisting of an N-terminal DNA-binding domain and a non-specific DNA cleavage domain at the C-terminal. Once the protein is bound to duplex DNA via its DNA-binding domain at the 5'-GGATG-3' recognition site, the DNA cleavage domain is activated and cleaves, without further sequence specificity, the first strand 9 nucleotides downstream and the second strand 13 nucleotides upstream of the nearest nucleotide of the recognition site. Its molecular mass is 65.4 kDa, being composed of 587 amino acids. DNA-binding domain The recognition domain contains three subdomains (D1, D2 and D3) that are evolutionarily related to the DNA-binding domain of the catabolite gene activator protein which contains a helix-turn-helix. DNA-cleavage domain DNA cleavage is mediated through the non-specific cleavage domain which also includes the dimerisation su ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Dimer (chemistry)
A dimer () ('' di-'', "two" + ''-mer'', "parts") is an oligomer consisting of two monomers joined by bonds that can be either strong or weak, covalent or intermolecular. Dimers also have significant implications in polymer chemistry, inorganic chemistry, and biochemistry. The term ''homodimer'' is used when the two molecules are identical (e.g. A–A) and ''heterodimer'' when they are not (e.g. A–B). The reverse of dimerization is often called dissociation. When two oppositely charged ions associate into dimers, they are referred to as ''Bjerrum pairs'', after Niels Bjerrum. Noncovalent dimers Anhydrous carboxylic acids form dimers by hydrogen bonding of the acidic hydrogen and the carbonyl oxygen. For example, acetic acid forms a dimer in the gas phase, where the monomer units are held together by hydrogen bonds. Under special conditions, most OH-containing molecules form dimers, e.g. the water dimer. Excimers and exciplexes are excited structures with a short lifetime. ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
