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DNA REPLICATION

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DNA replication is a biological process that occurs in all living organisms and copies their DNA; it is the basis for biological inheritance. The process starts with one double-stranded DNA molecule and produces two identical copies of the molecule. Each strand of the original double-stranded DNA molecule serves as template for the production of the complementary strand. Cellular proofreading and error toe-checking mechanisms ensure near perfect fidelity for DNA replication.living organismsDNAbiological inheritanceproofreadingnear perfect In a cell, DNA replication begins at specific locations in the genome, called "origins". Unwinding of DNA at the origin, and synthesis of new strands, forms a replication fork. In addition to DNA polymerase, the enzyme that synthesizes the new DNA by adding nucleotides matched to the template strand, a number of other proteins are associated with the fork and assist in the initiation and continuation of DNA synthesis.celloriginsreplication forkDNA polymeraseenzymenucleotidesproteins DNA replication can also be performed in vitro (artificially, outside a cell). DNA polymerases, isolated from cells, and artificial DNA primers are used to initiate DNA synthesis at known sequences in a template molecule. The polymerase chain reaction (PCR), a common laboratory technique, employs such artificial synthesis in a cyclic manner to amplify a specific target DNA fragment from a pool of DNA.in vitroDNA polymerasespolymerase chain reaction

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DNA STRUCTURE DNA usually exists as a double-stranded structure, with both strands coiled together to form the characteristic double-helix.double-helix Each single strand of DNA is a chain of four types of nucleotides having the bases: adenine, cytosine, guanine, and thymine.nucleotidesadeninecytosineguaninethymine A nucleotide is a mono-, di-, or triphosphate deoxyribonucleoside; that is, a deoxyribose sugar is attached to one, two, or three phosphates. Chemical interaction of these nucleotides forms phosphodiester linkages, creating the phosphate-deoxyribose backbone of the DNA double helix with the bases pointing inward.phosphodiester linkages Nucleotides (bases) are matched between strands through hydrogen bonds to form base pairs. Adenine pairs with thymine, and cytosine pairs with guanine.hydrogen bondsbase pairs

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a)Key features of DNA b) Partial chemical structure c) Space-filling model structure

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DNA strands have a directionality, and the different ends of a single strand are called the "3' (three-prime) end" and the "5' (five-prime) end". These terms refer to the carbon atom in deoxyribose to which the next phosphate in the chain attaches. In addition to being complementary, the two strands of DNA are antiparallel. They are orientated in opposite directions. This directionality has consequences in DNA synthesis, because DNA polymerase can synthesize DNA in only one direction by adding nucleotides to the 3' end of a DNA strand. The pairing of bases in DNA through hydrogen bonding means that the information contained within each strand is redundant. The nucleotides on a single strand can be used to reconstruct nucleotides on a newly synthesized partner strand

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DNA REPLICATION AND REPAIR The relationship between structure and function is manifest in the double helix. The idea that there is specific pairing of nitrogenous bases in DNA was the flash of inspiration that led Watson and Crick to the correct double helix. At the same time, they saw the functional significance of the base- pairing rules. They ended their classic paper with this wry statement : It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic mechanism for the genetic material.

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The figure below is an illustrates by Watson n Cricks basic idea. To make it easier to follow, we show only a short section of double helix in untwisted form. The two strands are complementary; each stores the information necessary to reconstruct the other. When a cell copies a DNA molecule, each strand serves as a template for ordering nucleotides into a new strand. Nucleotides line up along the template strand according to the base-pairing rules and are linked to form the new strand. Where there was one double-stranded DNA molecule at the beginning of the process, there are soon two, each an exact replica of the parent molecule. The copying mechanism is analogous to using a photographic negative to make a positive image, which can in turn be used to make another positive, and so on.

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This model of DNA replication remained untested for several years following publication of the DNA structure. The requisite experiments were simple in concept but difficult to perform. Watson and Cricks model predicts that when a double helix replicates, each of the two daughter molecules will have one old strand, derived from the parent molecules, and one newly made strand. This semiconservative model can be distinguish from a conservative model of replication, in which the two parent strands somehow come back together after the process( that is, the parent molecule is conserved). In yet a third model, called the dispersive model, all four strands of DNA following replication have a mixture of old and new DNA. Although mechanisms for conservatives or dispersive DNA replication are not easy devise, these models remained possibilities until they could be ruled out.

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STEPS OF REPLICATION

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The replication of DNA molecule begins at special sites called origin of replication. Multiple replication bubbles form and eventually fuse, thus speeding up the copying of the very long DNA molecules. At each end of a replication bubble is a replication fork, a Y-shaped region where the parental strands of DNA are being unwound. Helicases are enzymes that untwist the double helix at the replication forks, separating the two parental strands and making them available as template strands. After parental strands separation, single-strand binding proteins bind to the unpaired DNA strands, stabilizing them. The untwisting of the double helix causes tighter twisting and strain ahead of the replication fork. Topoisomerase helps releive this strain by breaking, swiveling, and rejoining DNA strands.

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The unwound sections of parental DNA strands are now available to serve as templates for the synthesis of new complementary DNA strands. However, the enzyme that synthesize DNA cannot initiate the synthesis of a polynucleotide; they can only add nucleotides to the end of an already existing chain that is base-paired with the template strand. The initial nucleotide chin that is produced during DNA synthesis ia actually a short stretch of RNA, not DNA. This RNA chain is called primer and is synthesized by the enzyme primase. Primase starts an RNA chain from a single nucleotide, adding RNA nucleotides one at a time, using the parental DNA strand as a template. The completed primer, generally 5 to 10 nucleotides long, is thus base- paired to the template strand. The new DNA strand will start from the 3end of the RNA primer.

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Synthesis of leading strand

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Synthesis of lagging strand

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