Recombinase
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Recombinase
Recombinases are genetic recombination enzymes. Site specific recombinases DNA recombinases are widely used in multicellular organisms to manipulate the structure of genomes, and to control gene expression. These enzymes, derived from bacteria (bacteriophages) and fungi, catalyze directionally sensitive DNA exchange reactions between short (30–40 nucleotides) target site sequences that are specific to each recombinase. These reactions enable four basic functional modules: excision/insertion, inversion, translocation and cassette exchange, which have been used individually or combined in a wide range of configurations to control gene expression. Types include: * Cre recombinase * Hin recombinase * Tre recombinase * FLP recombinase Homologous recombination Recombinases have a central role in homologous recombination in a wide range of organisms. Such recombinases have been described in archaea, bacteria, eukaryotes and viruses. Archaea The archaeon ''Sulfolobus solfataricus ...
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Site-specific Recombination
Site-specific recombination, also known as conservative site-specific recombination, is a type of genetic recombination in which DNA strand exchange takes place between segments possessing at least a certain degree of sequence homology. Enzymes known as site-specific recombinases (SSRs) perform rearrangements of DNA segments by recognizing and binding to short, specific DNA sequences (sites), at which they cleave the DNA backbone, exchange the two DNA helices involved, and rejoin the DNA strands. In some cases the presence of a recombinase enzyme and the recombination sites is sufficient for the reaction to proceed; in other systems a number of accessory proteins and/or accessory sites are required. Many different genome modification strategies, among these recombinase-mediated cassette exchange (RMCE), an advanced approach for the targeted introduction of transcription units into predetermined genomic loci, rely on SSRs. Site-specific recombination systems are highly specific, fa ...
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Cre Recombinase
Cre recombinase is a tyrosine recombinase enzyme derived from the P1 bacteriophage. The enzyme uses a topoisomerase I-like mechanism to carry out site specific recombination events. The enzyme (38kDa) is a member of the integrase family of site specific recombinase and it is known to catalyse the site specific recombination event between two DNA recognition sites ( LoxP sites). This 34 base pair (bp) loxP recognition site consists of two 13 bp palindromic sequences which flank an 8bp spacer region. The products of Cre-mediated recombination at loxP sites are dependent upon the location and relative orientation of the loxP sites. Two separate DNA species both containing loxP sites can undergo fusion as the result of Cre mediated recombination. DNA sequences found between two loxP sites are said to be " floxed". In this case the products of Cre mediated recombination depends upon the orientation of the loxP sites. DNA found between two loxP sites oriented in the same direction wi ...
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FLP Recombinase
In genetics, Flp-''FRT'' recombination is a site-directed recombination technology, increasingly used to manipulate an organism's DNA under controlled conditions ''in vivo''. It is analogous to Cre-''lox'' recombination but involves the recombination of sequences between short flippase recognition target (''FRT'') sites by the recombinase flippase (''Flp'') derived from the 2 µ plasmid of baker's yeast ''Saccharomyces cerevisiae''. The 34bp minimal FRT site sequence has the sequence ::5'3' for which flippase (Flp) binds to both 13-bp 5'-GAAGTTCCTATTC-3' arms flanking the 8 bp spacer, i.e. the site-specific recombination (region of crossover) in reverse orientation. ''FRT''-mediated cleavage occurs just ahead from the asymmetric 8bp core region (5''3') on the top strand and behind this sequence on the bottom strand. Several variant ''FRT'' sites exist, but recombination can usually occur only between two ''identical'' ''FRT''s but generally not among ''non-identical'' ("h ...
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DMC1 (gene)
Meiotic recombination protein DMC1/LIM15 homolog is a protein that in humans is encoded by the ''DMC1'' gene. Meiotic recombination protein Dmc1 is a homolog of the bacterial strand exchange protein RecA. Dmc1 plays the central role in homologous recombination in meiosis by assembling at the sites of programmed DNA double strand breaks and carrying out a search for allelic DNA sequences located on homologous chromatids. The name "Dmc" stands for "disrupted meiotic cDNA" and refers to the method used for its discovery which involved using clones from a meiosis-specific cDNA library to direct knock-out mutations of abundantly expressed meiotic genes. The Dmc1 protein is one of two homologs of RecA found in eukaryotic cells, the other being Rad51. In budding yeast, Rad51 serves as a strand exchange protein in mitosis where it is critical for the repair of DNA breaks. Rad51 is converted to an accessory factor for Dmc1 during meiosis by inhibition of its strand exchange activity. Homo ...
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Rad51
DNA repair protein RAD51 homolog 1 is a protein encoded by the gene ''RAD51''. The enzyme encoded by this gene is a member of the RAD51 protein family which assists in repair of DNA double strand breaks. RAD51 family members are homologous to the bacterial RecA, Archaeal RadA and yeast Rad51. The protein is highly conserved in most eukaryotes, from yeast to humans. Variants Two alternatively spliced transcript variants of this gene, which encode distinct proteins, have been reported. Transcript variants utilizing alternative polyA signals exist. Family In mammals, seven recA-like genes have been identified: Rad51, Rad51L1/B, Rad51L2/C, Rad51L3/D, XRCC2, XRCC3, and DMC1/Lim15. All of these proteins, with the exception of meiosis-specific DMC1, are essential for development in mammals. Rad51 is a member of thRecA-like NTPases Function In humans, RAD51 is a 339-amino acid protein that plays a major role in homologous recombination of DNA during double strand break r ...
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Homologous Recombination
Homologous recombination is a type of genetic recombination in which genetic information is exchanged between two similar or identical molecules of double-stranded or single-stranded nucleic acids (usually DNA as in cellular organisms but may be also RNA in viruses). Homologous recombination is widely used by cells to accurately DNA repair harmful breaks that occur on both strands of DNA, known as double-strand breaks (DSB), in a process called homologous recombinational repair (HRR). Homologous recombination also produces new combinations of DNA sequences during meiosis, the process by which eukaryotes make gamete cells, like sperm and egg cells in animals. These new combinations of DNA represent genetic variation in offspring, which in turn enables populations to adapt during the course of evolution. Homologous recombination is also used in horizontal gene transfer to exchange genetic material between different strains and species of bacteria and viruses. Horizontal ...
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Tre Recombinase
Tre recombinase is an experimental enzyme that in lab tests has removed DNA inserted by HIV from infected cells. Through selective mutation, Cre recombinase which recognizes loxP sites are modified to identify HIV long terminal repeats (loxLTR) instead. As a result, instead of performing Cre-Lox recombination Cre-Lox recombination is a site-specific recombinase technology, used to carry out deletions, insertions, translocations and inversions at specific sites in the DNA of cells. It allows the DNA modification to be targeted to a specific cell type ..., the new enzyme performs recombination at HIV provirus sites. The structure of Tre in complex with loxLTR has been resolved (), allowing for analyzing the roles of individual mutations. References External links * {{Antiretroviral drug Genetics Molecular biology ...
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Genetic Recombination
Genetic recombination (also known as genetic reshuffling) is the exchange of genetic material between different organisms which leads to production of offspring with combinations of traits that differ from those found in either parent. In eukaryotes, genetic recombination during meiosis can lead to a novel set of genetic information that can be further passed on from parents to offspring. Most recombination occurs naturally and can be classified into two types: (1) ''interchromosomal'' recombination, occurring through independent assortment of alleles whose loci are on different but homologous chromosomes (random orientation of pairs of homologous chromosomes in meiosis I); & (2) ''intrachromosomal'' recombination, occurring through crossing over. During meiosis in eukaryotes, genetic recombination involves the pairing of homologous chromosomes. This may be followed by information transfer between the chromosomes. The information transfer may occur without physical exchange (a se ...
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Hin Recombinase
Hin recombinase is a 21kD protein composed of 198 amino acids that is found in the bacteria ''Salmonella''. Hin belongs to the serine recombinase family (B2) of DNA invertases in which it relies on the active site serine to initiate DNA cleavage and recombination. The related protein, gamma-delta resolvase shares high similarity to Hin, of which much structural work has been done, including structures bound to DNA and reaction intermediates. Hin functions to invert a 900 base pair ( bp) DNA segment within the salmonella genome that contains a promoter for downstream flagellar genes, fljA and fljB. Inversion of the intervening DNA alternates the direction of the promoter and thereby alternates expression of the flagellar genes. This is advantageous to the bacterium as a means of escape from the host immune response. Hin functions by binding to two 26bp imperfect inverted repeat sequences as a homodimer. These hin binding sites flank the invertible segment which not only encodes t ...
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D-loop
In molecular biology, a displacement loop or D-loop is a DNA structure where the two strands of a double-stranded DNA molecule are separated for a stretch and held apart by a third strand of DNA. An R-loop is similar to a D-loop, but in this case the third strand is RNA rather than DNA. The third strand has a base sequence which is complementary to one of the main strands and pairs with it, thus displacing the other complementary main strand in the region. Within that region the structure is thus a form of triple-stranded DNA. A diagram in the paper introducing the term illustrated the D-loop with a shape resembling a capital "D", where the displaced strand formed the loop of the "D". D-loops occur in a number of particular situations, including in DNA repair, in telomeres, and as a semi-stable structure in mitochondrial circular DNA molecules. In mitochondria Researchers at Caltech discovered in 1971 that the circular mitochondrial DNA from growing cells included a short ...
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DNA Damage (naturally Occurring)
DNA damage is an alteration in the chemical structure of DNA, such as a break in a strand of DNA, a nucleobase missing from the backbone of DNA, or a chemically changed base such as 8-OHdG. DNA damage can occur naturally or via environmental factors, but is distinctly different from mutation, although both are types of error in DNA. DNA damage is an abnormal chemical structure in DNA, while a mutation is a change in the sequence of base pairs. DNA damages cause changes in the structure of the genetic material and prevents the replication mechanism from functioning and performing properly. The DNA damage response (DDR) is a complex signal transduction pathway which recognizes when DNA is damaged and initiates the cellular response to the damage. DNA damage and mutation have different biological consequences. While most DNA damages can undergo DNA repair, such repair is not 100% efficient. Un-repaired DNA damages accumulate in non-replicating cells, such as cells in the brains o ...
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Gene Cassette
In biology, a gene cassette is a type of mobile genetic element that contains a gene and a recombination site. Each cassette usually contains a single gene and tends to be very small; on the order of 500–1000 base pairs. They may exist incorporated into an integron or freely as circular DNA. Gene cassettes can move around within an organism's genome or be transferred to another organism in the environment via horizontal gene transfer. These cassettes often carry antibiotic resistance genes. An example would be the '' kanMX'' cassette which confers kanamycin (an antibiotic) resistance upon bacteria. Integrons Integrons are genetic structures in bacteria which express and are capable of acquiring and exchanging gene cassettes. The integron consists of a promoter, an attachment site, and an integrase gene that encodes a site-specific recombinase There are three classes of integrons described. The mobile units that insert into integrons are gene cassettes. For cassettes that ca ...
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