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In
molecular biology Molecular biology is the branch of biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, P ...
, DNA replication is the
biological process Biological processes are those processes that are vital for an organism to live, and that shape its capacities for interacting with its environment. Biological processes are made of many chemical reactions or other events that are involved in th ...
of producing two identical replicas of DNA from one original
DNA Deoxyribonucleic acid (; DNA) is a molecule A scanning tunneling microscopy image of pentacene molecules, which consist of linear chains of five carbon rings. A molecule is an electrically Electricity is the set of physical ...

DNA
molecule. DNA replication occurs in all
living organisms In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, Physiology, physiological ...

living organisms
acting as the most essential part for
biological inheritance Heredity, also called inheritance or biological inheritance, is the passing on of traits Trait may refer to: * Phenotypic trait in biology, which involve genes and characteristics of organisms * Trait (computer programming), a model for st ...

biological inheritance
. This is essential for cell division during growth and repair of damaged tissues, while it also ensures that each of the new cells receives its own copy of the DNA. The cell possesses the distinctive property of division, which makes replication of DNA essential. DNA is made up of a
double helix In molecular biology Molecular biology is the branch of biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, ...
of two
complementary A complement is often something that completes something else, or at least adds to it in some useful way. Thus it may be: * Complement (linguistics), a word or phrase having a particular syntactic role ** Subject complement, a word or phrase addi ...
strands. The double helix describes the appearance of a double-stranded DNA which is thus composed of two linear strands that run opposite to each other and twist together to form. During replication, these strands are separated. Each strand of the original DNA molecule then serves as a template for the production of its counterpart, a process referred to as
semiconservative replication Semiconservative replication describes the mechanism of DNA replication in all known cells. DNA replication occurs on multiple Origin of replication, origins of replication along the DNA template strand. As the DNA double helix is unwound by helicas ...

semiconservative replication
. As a result of semi-conservative replication, the new helix will be composed of an original DNA strand as well as a newly synthesized strand. Cellular
proofreading Proofreading is the reading Reading is the process of taking in the sense or meaning of letters, symbols, ''etc.'', especially by sight or touch. For educators and researchers, reading is a multifaceted process involving such areas as wor ...
and error-checking mechanisms ensure near perfect
fidelity Fidelity is the quality of faithfulness Eugene Santos or Faithfulness is the concept of unfailingly remaining loyal to someone or something, and putting that loyalty into consistent practice regardless of extenuating circumstances. It may be ex ...

fidelity
for DNA replication.Imperfect DNA replication results in
mutation In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, Physiology, physiological mechan ...
s.
In a
cell Cell most often refers to: * Cell (biology) The cell (from Latin Latin (, or , ) is a classical language belonging to the Italic languages, Italic branch of the Indo-European languages. Latin was originally spoken in the area around ...
, DNA replication begins at specific locations, or origins of replication, in the
genome In the fields of molecular biology Molecular biology is the branch of biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, M ...

genome
which contains the genetic material of an organism. Unwinding of DNA at the origin and synthesis of new strands, accommodated by an
enzyme Enzymes () are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates in ...

enzyme
known as
helicase Helicases are a class of enzyme Enzymes () are protein Proteins are large s and s that comprise one or more long chains of . Proteins perform a vast array of functions within organisms, including , , , providing and , and from one ...

helicase
, results in
replication fork In molecular biology, DNA replication is the biological process of producing two identical replicas of DNA from one original DNA molecule. DNA replication occurs in all life, living organisms acting as the most essential part for heredity, biologi ...

replication fork
s growing bi-directionally from the origin. A number of
protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture solved by X-ray crystallography by Max Perutz and Sir John Cowdery Kendrew in 1958, for which they received a No ...

protein
s are associated with the replication fork to help in the initiation and continuation of
DNA synthesis DNA synthesis is the natural or artificial creation of deoxyribonucleic acid Deoxyribonucleic acid (; DNA) is a molecule composed of two polynucleotide chains that coil around each other to form a Nucleic acid double helix, double helix ca ...
. Most prominently,
DNA polymerase A DNA polymerase is a member of a family of enzyme Enzymes () are s that act as s (biocatalysts). Catalysts accelerate . The molecules upon which enzymes may act are called , and the enzyme converts the substrates into different molecules ...

DNA polymerase
synthesizes the new strands by adding
nucleotide Nucleotides are organic molecules , CH4; is among the simplest organic compounds. In chemistry, organic compounds are generally any chemical compounds that contain carbon-hydrogen chemical bond, bonds. Due to carbon's ability to Catenation, ...

nucleotide
s that complement each (template) strand. DNA replication occurs during the S-stage of
interphase Interphase is the portion of the cell cycle The cell cycle, or cell-division cycle, is the series of events that take place in a cell that cause it to divide into two daughter cells. These events include the duplication of its DNA (DNA ...

interphase
. DNA replication (DNA amplification) can also be performed ''
in vitro ''In vitro'' (meaning in glass, or ''in the glass'') studies Study or studies may refer to: General * Education **Higher education * Clinical trial * Experiment * Observational study * Research * Study skills, abilities and approaches applie ...

in vitro
'' (artificially, outside a cell). DNA polymerases isolated from cells and artificial DNA primers can be used to start DNA synthesis at known sequences in a template DNA molecule.
Polymerase chain reaction Polymerase chain reaction (PCR) is a method widely used to rapidly make millions to billions of copies (complete copies or partial copies) of a specific DNA Deoxyribonucleic acid (; DNA) is a molecule File:Pentacene on Ni(111) STM. ...

Polymerase chain reaction
(PCR),
ligase chain reaction The ligase chain reaction (LCR) is a method of DNA Deoxyribonucleic acid (; DNA) is a molecule File:Pentacene on Ni(111) STM.jpg, A scanning tunneling microscopy image of pentacene molecules, which consist of linear chains of five carbon ...
(LCR), and
transcription-mediated amplification Transcription-mediated amplification (TMA) is an isothermal (does not change the nucleic acid temperature), single-tube nucleic acid Nucleic acids are biopolymers, macromolecules, essential to all Organism, known forms of life. They are composed ...

transcription-mediated amplification
(TMA) are examples. In March 2021, researchers reported evidence suggesting that a preliminary form of
transfer RNA Transfer RNA (abbreviated tRNA and formerly referred to as sRNA, for soluble RNA) is an adaptor molecule A scanning tunneling microscopy image of pentacene molecules, which consist of linear chains of five carbon rings. A molecule is an e ...
, a necessary component of
translation Translation is the communication of the meaning Meaning most commonly refers to: * Meaning (linguistics), meaning which is communicated through the use of language * Meaning (philosophy), definition, elements, and types of meaning discusse ...

translation
, the biological synthesis of new
protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture solved by X-ray crystallography by Max Perutz and Sir John Cowdery Kendrew in 1958, for which they received a No ...

protein
s in accordance with the
genetic code The genetic code is the set of rules used by living cells Cell most often refers to: * Cell (biology), the functional basic unit of life Cell may also refer to: Closed spaces * Monastic cell, a small room, hut, or cave in which a monk or rel ...

genetic code
, could have been a replicator molecule itself in the very early development of life, or
abiogenesis In evolutionary biology Evolutionary biology is the subfield of biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molec ...
.


DNA structure

DNA exists as a double-stranded structure, with both strands coiled together to form the characteristic
double-helix In molecular biology, the term double helix refers to the structure formed by double-stranded molecules of nucleic acids such as DNA. The double helical structure of a nucleic acid complex arises as a consequence of its secondary structure ...
. Each single strand of DNA is a chain of four types of
nucleotide Nucleotides are organic molecules , CH4; is among the simplest organic compounds. In chemistry, organic compounds are generally any chemical compounds that contain carbon-hydrogen chemical bond, bonds. Due to carbon's ability to Catenation, ...

nucleotide
s. Nucleotides in DNA contain a
deoxyribose Deoxyribose, or more precisely 2-deoxyribose, is a monosaccharide Monosaccharides (from Greek#REDIRECT Greek Greek may refer to: Greece Anything of, from, or related to Greece Greece ( el, Ελλάδα, , ), officially the Hellenic Rep ...

deoxyribose
sugar, a
phosphate In chemistry, a phosphate is an anion, salt (chemistry), salt, functional group or ester derived from a phosphoric acids and phosphates, phosphoric acid. It most commonly means orthophosphate, a derivative of phosphoric acid, orthophosphoric a ...

phosphate
, and a
nucleobase Nucleobases, also known as ''nitrogenous bases'' or often simply ''bases'', are nitrogen-containing biological compounds that form nucleosides Nucleosides are glycosylamines that can be thought of as nucleotide Nucleotides are organic mo ...
. The four types of
nucleotide Nucleotides are organic molecules , CH4; is among the simplest organic compounds. In chemistry, organic compounds are generally any chemical compounds that contain carbon-hydrogen chemical bond, bonds. Due to carbon's ability to Catenation, ...

nucleotide
correspond to the four
nucleobase Nucleobases, also known as ''nitrogenous bases'' or often simply ''bases'', are nitrogen-containing biological compounds that form nucleosides Nucleosides are glycosylamines that can be thought of as nucleotide Nucleotides are organic mo ...
s
adenine Adenine (A, Ade) is a nucleobase 230px, Pyrimidine nucleobases are simple ring molecules. Nucleobases, also known as ''nitrogenous bases'' or often simply ''bases'', are nitrogen-containing biological compounds that form nucleosides Nucleos ...

adenine
,
cytosine Cytosine () (symbol A symbol is a mark, sign, or word In linguistics, a word of a spoken language can be defined as the smallest sequence of phonemes that can be uttered in isolation with semantic, objective or pragmatics, practical ...

cytosine
,
guanine Guanine () (symbol A symbol is a mark, sign, or word In linguistics, a word of a spoken language can be defined as the smallest sequence of phonemes that can be uttered in isolation with semantic, objective or pragmatics, practical me ...

guanine
, and
thymine Thymine () (symbol A symbol is a mark, sign, or word In linguistics, a word of a spoken language can be defined as the smallest sequence of phonemes that can be uttered in isolation with semantic, objective or pragmatics, practical m ...

thymine
, commonly abbreviated as A, C, G and T. Adenine and guanine are
purine Purine is a heterocyclic 125px, Pyridine, a heterocyclic compound A heterocyclic compound or ring structure is a cyclic compound that has atoms of at least two different chemical element, elements as members of its ring(s). Heterocyclic chemi ...

purine
bases, while cytosine and thymine are
pyrimidine Pyrimidine is an aromatic In chemistry Chemistry is the scientific Science () is a systematic enterprise that builds and organizes knowledge Knowledge is a familiarity or awareness, of someone or something, such as facts ...

pyrimidine
s. These nucleotides form
phosphodiester bonds A phosphodiester bond occurs when exactly two of the hydroxyl groups A hydroxy or hydroxyl group is a functional group with the chemical formula -OH and composed of one oxygen Oxygen is the chemical element Image:Simple Periodic Ta ...
, creating the phosphate-deoxyribose backbone of the DNA double helix with the nucleobases pointing inward (i.e., toward the opposing strand). Nucleobases are matched between strands through
hydrogen bonds A hydrogen bond (or H-bond) is a primarily Electrostatics, electrostatic force of attraction between a hydrogen Hydrogen is the chemical element Image:Simple Periodic Table Chart-blocks.svg, 400px, Periodic table, The periodic table of ...
to form
base pair A base pair (bp) is a fundamental unit of double-stranded nucleic acids Nucleic acids are biopolymer Biopolymers are natural polymers produced by the cells of Organism, living organisms. Biopolymers consist of monomeric units that are Covalent_ ...
s. Adenine pairs with thymine (two hydrogen bonds), and guanine pairs with cytosine (three
hydrogen bonds A hydrogen bond (or H-bond) is a primarily Electrostatics, electrostatic force of attraction between a hydrogen Hydrogen is the chemical element Image:Simple Periodic Table Chart-blocks.svg, 400px, Periodic table, The periodic table of ...
). 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". By convention, if the base sequence of a single strand of DNA is given, the left end of the sequence is the 5′ end, while the right end of the sequence is the 3′ end. The strands of the double helix are anti-parallel with one being 5′ to 3′, and the opposite strand 3′ to 5′. These terms refer to the carbon atom in deoxyribose to which the next phosphate in the chain attaches. 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 complementary bases in DNA (through
hydrogen bonding A hydrogen bond (or H-bond) is a primarily Electrostatics, electrostatic force of attraction between a hydrogen Hydrogen is the chemical element Image:Simple Periodic Table Chart-blocks.svg, 400px, Periodic table, The periodic table of ...
) means that the information contained within each strand is redundant. Phosphodiester (intra-strand) bonds are stronger than hydrogen (inter-strand) bonds. The actual job of the phosphodiester bonds is where in DNA polymers connect the 5' carbon atom of one nucleotide to the 3' carbon atom of another nucleotide, while the hydrogen bonds stabilize DNA double helices across the helix axis but not in the direction of the axis 1. This allows the strands to be separated from one another. The nucleotides on a single strand can therefore be used to reconstruct nucleotides on a newly synthesized partner strand.


DNA polymerase

DNA polymerase A DNA polymerase is a member of a family of enzyme Enzymes () are s that act as s (biocatalysts). Catalysts accelerate . The molecules upon which enzymes may act are called , and the enzyme converts the substrates into different molecules ...

DNA polymerase
s are a family of
enzyme Enzymes () are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates in ...

enzyme
s that carry out all forms of DNA replication. DNA polymerases in general cannot initiate synthesis of new strands, but can only extend an existing DNA or RNA strand paired with a template strand. To begin synthesis, a short fragment of RNA, called a primer, must be created and paired with the template DNA strand. DNA polymerase adds a new strand of DNA by extending the 3′ end of an existing nucleotide chain, adding new
nucleotide Nucleotides are organic molecules , CH4; is among the simplest organic compounds. In chemistry, organic compounds are generally any chemical compounds that contain carbon-hydrogen chemical bond, bonds. Due to carbon's ability to Catenation, ...

nucleotide
s matched to the template strand one at a time via the creation of
phosphodiester bond A phosphodiester bond occurs when exactly two of the hydroxyl groups A hydroxy or hydroxyl group is a functional group with the chemical formula -OH and composed of one oxygen atom Chemical bond , covalently bonded to one hydrogen atom. In or ...
s. The energy for this process of DNA polymerization comes from hydrolysis of the
high-energy phosphate High-energy phosphate can mean one of two things: * The phosphate In chemistry, a phosphate is an anion, salt (chemistry), salt, functional group or ester derived from a phosphoric acids and phosphates, phosphoric acid. It most commonly means ...
(phosphoanhydride) bonds between the three phosphates attached to each unincorporated
base Base or BASE may refer to: Brands and enterprises *Base (mobile telephony provider) Base (stylized as BASE) is the third largest of Belgium Belgium ( nl, België ; french: Belgique ; german: Belgien ), officially the Kingdom of Belgium, ...

base
. Free bases with their attached phosphate groups are called
nucleotide Nucleotides are organic molecules , CH4; is among the simplest organic compounds. In chemistry, organic compounds are generally any chemical compounds that contain carbon-hydrogen chemical bond, bonds. Due to carbon's ability to Catenation, ...

nucleotide
s; in particular, bases with three attached phosphate groups are called
nucleoside triphosphate Nucleosides are glycosylamine 120px, Cyclic hemiaminal ether bond derived from an aldehyde Glycosylamines are a class of biochemical compounds consisting of a Glycosyl, glycosyl group attached to an amine, amino group, -NR2. They are also known ...
s. When a nucleotide is being added to a growing DNA strand, the formation of a phosphodiester bond between the proximal phosphate of the nucleotide to the growing chain is accompanied by hydrolysis of a high-energy phosphate bond with release of the two distal phosphate groups as a
pyrophosphate In chemistry Chemistry is the scientific Science () is a systematic enterprise that builds and organizes knowledge Knowledge is a familiarity or awareness, of someone or something, such as facts A fact is an occurrence in th ...
. Enzymatic hydrolysis of the resulting
pyrophosphate In chemistry Chemistry is the scientific Science () is a systematic enterprise that builds and organizes knowledge Knowledge is a familiarity or awareness, of someone or something, such as facts A fact is an occurrence in th ...
into inorganic phosphate consumes a second high-energy phosphate bond and renders the reaction effectively irreversible.The
energetics Energetics is the study of energy In physics Physics is the that studies , its , its and behavior through , and the related entities of and . "Physical science is that department of knowledge which relates to the order of nature, ...
of this process may also help explain the directionality of synthesis—if DNA were synthesized in the 3′ to 5′ direction, the energy for the process would come from the 5′ end of the growing strand rather than from free nucleotides. The problem is that if the high energy triphosphates were on the growing strand and not on the free nucleotides, proof-reading by removing a mismatched terminal nucleotide would be problematic: Once a nucleotide is added, the triphosphate is lost and a single phosphate remains on the backbone between the new nucleotide and the rest of the strand. If the added nucleotide were mismatched, removal would result in a DNA strand terminated by a monophosphate at the end of the "growing strand" rather than a high energy triphosphate. So strand would be stuck and wouldn't be able to grow anymore. In actuality, the high energy triphosphates hydrolyzed at each step originate from the free nucleotides, not the polymerized strand, so this issue does not exist.
In general, DNA polymerases are highly accurate, with an intrinsic error rate of less than one mistake for every 107 nucleotides added. In addition, some DNA polymerases also have proofreading ability; they can remove nucleotides from the end of a growing strand in order to correct mismatched bases. Finally, post-replication mismatch repair mechanisms monitor the DNA for errors, being capable of distinguishing mismatches in the newly synthesized DNA strand from the original strand sequence. Together, these three discrimination steps enable replication fidelity of less than one mistake for every 109 nucleotides added. The rate of DNA replication in a living cell was first measured as the rate of phage T4 DNA elongation in phage-infected ''E. coli''. During the period of exponential DNA increase at 37 °C, the rate was 749 nucleotides per second. The mutation rate per base pair per replication during phage T4 DNA synthesis is 1.7 per 108.


Replication process

DNA replication, like all biological polymerization processes, proceeds in three enzymatically catalyzed and coordinated steps: initiation, elongation and termination.


Initiation

For a , it must first replicate its DNA. DNA replication is an all-or-none process; once replication begins, it proceeds to completion. Once replication is complete, it does not occur again in the same cell cycle. This is made possible by the division of initiation of the
pre-replication complex A pre-replication complex (pre-RC) is a protein complex A protein complex or multiprotein complex is a group of two or more associated polypeptide chain Peptides (from Greek language Greek (modern , romanized: ''Elliniká'', Ancient Greek ...
.


Pre-replication complex

In late
mitosis In cell biology Cell biology (also cellular biology or cytology) is a branch of biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical proce ...

mitosis
and early
G1 phase The G1 phase, gap 1 phase, or growth 1 phase, is the first of four phases of the cell cycle The cell cycle, or cell-division cycle, is the series of events that take place in a cell Cell most often refers to: * Cell (biology), the function ...
, a large complex of initiator proteins assembles into the pre-replication complex at particular points in the DNA, known as "
origin Origin(s) or The Origin may refer to: Arts, entertainment, and media Comics and manga * Origin (comics), ''Origin'' (comics), a Wolverine comic book mini-series published by Marvel Comics in 2002 * The Origin (Buffy comic), ''The Origin'' (Bu ...
s". In ''
E. coli ''Escherichia coli'' (),Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. also known as ''E. coli'' (), is a Gram-negative Gram-negative bacteria are bacteria Bacteria (; common noun bacteri ...

E. coli
'' the primary initiator protein is
DnaA DnaA is a protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture solved by X-ray crystallography by Max Perutz and Sir John Cowdery Kendrew in 1958, for whi ...
; in
yeast Yeasts are eukaryotic Eukaryotes () are organism In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular ...

yeast
, this is the
origin recognition complex In molecular biology Molecular biology is the branch of biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, m ...
. Sequences used by initiator proteins tend to be "AT-rich" (rich in adenine and thymine bases), because A-T base pairs have two hydrogen bonds (rather than the three formed in a C-G pair) and thus are easier to strand-separate. In eukaryotes, the origin recognition complex catalyzes the assembly of initiator proteins into the pre-replication complex.
Cdc6 Cell division control protein 6 homolog is a protein that in humans is encoded by the ''CDC6'' gene. The protein encoded by this gene is highly similar to Cdc6, Saccharomyces cerevisiae Cdc6, a protein essential for the initiation of DNA replicati ...

Cdc6
and
Cdt1 CDT1 (Chromatin licensing and DNA replication factor 1) is a protein that in humans is encoded by the ''CDT1'' gene. It is a licensing factor that functions to limit DNA from replicating more than once per cell cycle. Role in pre-replication comp ...
then associate with the bound origin recognition complex at the origin in order to form a larger complex necessary to load the Mcm complex onto the DNA. The Mcm complex is the helicase that will unravel the DNA helix at the replication origins and
replication fork In molecular biology, DNA replication is the biological process of producing two identical replicas of DNA from one original DNA molecule. DNA replication occurs in all life, living organisms acting as the most essential part for heredity, biologi ...

replication fork
s in eukaryotes. The Mcm complex is recruited at late G1 phase and loaded by the ORC-Cdc6-Cdt1 complex onto the DNA via ATP-dependent protein remodeling. The loading of the Mcm complex onto the origin DNA marks the completion of pre-replication complex formation. If environmental conditions are right in late G1 phase, the G1 and G1/S
cyclin Cyclin is a family of proteins that controls the progression of a cell through the cell cycle The cell cycle, or cell-division cycle, is the series of events that take place in a cell Cell most often refers to: * Cell (biology), the funct ...
-
CdkCDK may refer to: * CDK Global, a US-based automotive dealer services company * The IATA airport code for George T. Lewis Airport, Cedar Key, Florida, United States. * Chemistry Development Kit, an open source chemical expert system for chemoinforma ...
complexes are activated, which stimulate expression of genes that encode components of the DNA synthetic machinery. G1/S-Cdk activation also promotes the expression and activation of S-Cdk complexes, which may play a role in activating replication origins depending on species and cell type. Control of these Cdks vary depending cell type and stage of development.  This regulation is best understood in
budding yeast ''Saccharomyces cerevisiae'' () is a species of yeast Yeasts are eukaryotic Eukaryotes () are organism In biology, an organism () is any organic, life, living system that functions as an individual entity. All organisms are ...
, where the S cyclins Clb5 and Clb6 are primarily responsible for DNA replication. Clb5,6-Cdk1 complexes directly trigger the activation of replication origins and are therefore required throughout S phase to directly activate each origin. In a similar manner,
Cdc7 Cell division cycle 7-related protein kinase is an enzyme Enzymes () are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrate (chemis ...
is also required through
S phase S phase (Synthesis Phase) is the phase of the cell cycle The cell cycle, or cell-division cycle, is the series of events that take place in a cell that cause it to divide into two daughter cells. These events include the duplication of ...
to activate replication origins. Cdc7 is not active throughout the cell cycle, and its activation is strictly timed to avoid premature initiation of DNA replication. In late G1, Cdc7 activity rises abruptly as a result of association with the regulatory subunit
Dbf4 Protein DBF4 homolog A is a protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture solved by X-ray crystallography by Max Perutz and Sir John Cowdery Kendrew ...
, which binds Cdc7 directly and promotes its protein kinase activity. Cdc7 has been found to be a rate-limiting regulator of origin activity. Together, the G1/S-Cdks and/or S-Cdks and Cdc7 collaborate to directly activate the replication origins, leading to initiation of DNA synthesis.


Preinitiation complex

In early S phase, S-Cdk and Cdc7 activation lead to the assembly of the preinitiation complex, a massive protein complex formed at the origin. Formation of the preinitiation complex displaces Cdc6 and Cdt1 from the origin replication complex, inactivating and disassembling the pre-replication complex. Loading the preinitiation complex onto the origin activates the Mcm helicase, causing unwinding of the DNA helix.  The preinitiation complex also loads α-primase and other DNA polymerases onto the DNA. After α-primase synthesizes the first primers, the primer-template junctions interact with the clamp loader, which loads the sliding clamp onto the DNA to begin DNA synthesis. The components of the preinitiation complex remain associated with replication forks as they move out from the origin.


Elongation

DNA polymerase has 5′–3′ activity. All known DNA replication systems require a free 3′
hydroxyl A hydroxy or hydroxyl group is a functional group with the chemical formula -OH and composed of one oxygen Oxygen is the chemical element Image:Simple Periodic Table Chart-blocks.svg, 400px, Periodic table, The periodic table of the ...

hydroxyl
group before synthesis can be initiated (note: the DNA template is read in 3′ to 5′ direction whereas a new strand is synthesized in the 5′ to 3′ direction—this is often confused). Four distinct mechanisms for DNA synthesis are recognized: #All cellular life forms and many DNA
virus A virus is a submicroscopic infectious agent In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecu ...

virus
es,
phage A bacteriophage (), also known informally as a ''phage'' (), is a virus A virus is a that only inside the living of an . Viruses infect all , from animals and plants to s, including and . Since 's 1892 article describing a non-b ...

phage
s and
plasmid A plasmid is a small, extrachromosomal DNA Extrachromosomal DNA (abbreviated ecDNA) is any DNA that is found off the chromosome A chromosome is a long DNA molecule with part or all of the genetic material of an organism. Most eukaryo ...
s use a
primase DNA primase is an enzyme Enzymes () are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enz ...

primase
to synthesize a short RNA primer with a free 3′ OH group which is subsequently elongated by a DNA polymerase. #The retroelements (including
retrovirus A retrovirus is a type of virus A virus is a submicroscopic infectious agent In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical ...

retrovirus
es) employ a transfer RNA that primes DNA replication by providing a free 3′ OH that is used for elongation by the
reverse transcriptase A reverse transcriptase (RT) is an enzyme Enzymes () are protein Proteins are large s and s that comprise one or more long chains of . Proteins perform a vast array of functions within organisms, including , , , providing and , an ...
. #In the
adenovirus Adenoviruses (members of the family In human society A society is a Social group, group of individuals involved in persistent Social relation, social interaction, or a large social group sharing the same spatial or social territor ...

adenovirus
es and the φ29 family of
bacteriophage A bacteriophage (), also known informally as a ''phage'' (), is a virus A virus is a submicroscopic infectious agent In biology Biology is the natural science that studies life and living organisms, including their anatomy, ...

bacteriophage
s, the 3′ OH group is provided by the side chain of an amino acid of the genome attached protein (the terminal protein) to which nucleotides are added by the DNA polymerase to form a new strand. #In the single stranded DNA viruses—a group that includes the
circovirus ''Circovirus'' is a genus of viruses, in the family ''Circoviridae''. Birds (such as pigeons and ducks) and pigs serve as natural hosts, though dogs have been shown to be infected as well. It is a single stranded DNA virus (ssDNA). There are 49 ...
es, the
geminivirus ''Geminiviridae'' is a family of plant viruses Plant viruses are virus A virus is a submicroscopic infectious agent that Viral replication, replicates only inside the living Cell (biology), cells of an organism. Viruses infect all t ...
es, the
parvovirus Parvoviruses are a family of animal viruses that constitute the family ''Parvoviridae''. They have linear, single-stranded DNA (ssDNA) genomes that typically contain two genes encoding for a replication initiator protein, called NS1, and the pro ...
es and others—and also the many phages and
plasmid A plasmid is a small, extrachromosomal DNA Extrachromosomal DNA (abbreviated ecDNA) is any DNA that is found off the chromosome A chromosome is a long DNA molecule with part or all of the genetic material of an organism. Most eukaryo ...
s that use the rolling circle replication (RCR) mechanism, the RCR endonuclease creates a nick in the genome strand (single stranded viruses) or one of the DNA strands (plasmids). The 5′ end of the nicked strand is transferred to a
tyrosine -Tyrosine or tyrosine (symbol Tyr or Y) or 4-hydroxyphenylalanine is one of the 20 standard amino acid Amino acids are organic compound , CH4; is among the simplest organic compounds. In chemistry, organic compounds are generally any chemi ...

tyrosine
residue on the nuclease and the free 3′ OH group is then used by the DNA polymerase to synthesize the new strand. The first is the best known of these mechanisms and is used by the cellular organisms. In this mechanism, once the two strands are separated,
primase DNA primase is an enzyme Enzymes () are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enz ...

primase
adds RNA primers to the template strands. The leading strand receives one RNA primer while the lagging strand receives several. The leading strand is continuously extended from the primer by a DNA polymerase with high
processivity In molecular biology and biochemistry, processivity is an enzyme's ability to catalyze "consecutive reactions without releasing its enzyme substrate (biology), substrate". For example, processivity is the average number of nucleotides added by a ...
, while the lagging strand is extended discontinuously from each primer forming
Okazaki fragments Okazaki fragments are short sequences of DNA Deoxyribonucleic acid (; DNA) is a molecule A scanning tunneling microscopy image of pentacene molecules, which consist of linear chains of five carbon rings. A molecule is an electric ...
.
RNase Ribonuclease (commonly abbreviated RNase) is a type of nuclease A nuclease (also archaically known as nucleodepolymerase or polynucleotidase) is an enzyme capable of cleaving the phosphodiester bonds between nucleotides of nucleic acids. Nuclea ...
removes the primer RNA fragments, and a low processivity DNA polymerase distinct from the replicative polymerase enters to fill the gaps. When this is complete, a single nick on the leading strand and several nicks on the lagging strand can be found.
Ligase In biochemistry Biochemistry or biological chemistry, is the study of chemical process In a scientific Science () is a systematic enterprise that Scientific method, builds and organizes knowledge in the form of Testability, testable ...

Ligase
works to fill these nicks in, thus completing the newly replicated DNA molecule. The primase used in this process differs significantly between
bacteria Bacteria (; common noun bacteria, singular bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of prokaryotic microorganisms. Typ ...

bacteria
and
archaea Archaea ( ; singular archaeon ) constitute a domain Domain may refer to: Mathematics *Domain of a function, the set of input values for which the (total) function is defined **Domain of definition of a partial function **Natural domain of a pa ...

archaea
/
eukaryote Eukaryotes () are organism In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interact ...

eukaryote
s. Bacteria use a primase belonging to the
DnaG DnaG is a bacterial DNA Deoxyribonucleic acid (; DNA) is a molecule File:Pentacene on Ni(111) STM.jpg, A scanning tunneling microscopy image of pentacene molecules, which consist of linear chains of five carbon rings. A molecule is an ...
protein superfamily which contains a catalytic domain of the TOPRIM fold type. The TOPRIM fold contains an α/β core with four conserved strands in a Rossmann-like topology. This structure is also found in the catalytic domains of
topoisomerase Topoisomerases (or DNA topoisomerases) are enzymes that participate in the overwinding or underwinding of DNA Deoxyribonucleic acid (; DNA) is a molecule File:Pentacene on Ni(111) STM.jpg, A scanning tunneling microscopy image of pent ...
Ia, topoisomerase II, the OLD-family nucleases and DNA repair proteins related to the RecR protein. The primase used by archaea and eukaryotes, in contrast, contains a highly derived version of the
RNA recognition motif RNA recognition motif, RNP-1 is a putative RNA-binding protein domain, domain of about 90 amino acids that are known to bind single-stranded RNAs. It was found in many eukaryotic proteins. The largest group of single strand RNA-binding protein ...
(RRM). This primase is structurally similar to many viral RNA-dependent RNA polymerases, reverse transcriptases, cyclic nucleotide generating cyclases and DNA polymerases of the A/B/Y families that are involved in DNA replication and repair. In eukaryotic replication, the primase forms a complex with Pol α. Multiple DNA polymerases take on different roles in the DNA replication process. In ''
E. coli ''Escherichia coli'' (),Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. also known as ''E. coli'' (), is a Gram-negative Gram-negative bacteria are bacteria Bacteria (; common noun bacteri ...

E. coli
'', is the polymerase enzyme primarily responsible for DNA replication. It assembles into a replication complex at the replication fork that exhibits extremely high processivity, remaining intact for the entire replication cycle. In contrast, DNA Pol I is the enzyme responsible for replacing RNA primers with DNA. DNA Pol I has a 5′ to 3′
exonuclease Exonucleases are enzymes Enzymes () are protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture solved by X-ray crystallography by Max Perutz and Sir ...
activity in addition to its polymerase activity, and uses its exonuclease activity to degrade the RNA primers ahead of it as it extends the DNA strand behind it, in a process called
nick translation Nick translation (or head translation), developed in 1977 by Peter Rigby and Paul Berg, is a tagging technique in molecular biology Molecular biology is the branch of biology Biology is the natural science that studies life and living ...
. Pol I is much less processive than Pol III because its primary function in DNA replication is to create many short DNA regions rather than a few very long regions. In
eukaryote Eukaryotes () are organism In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interact ...

eukaryote
s, the low-processivity enzyme, Pol α, helps to initiate replication because it forms a complex with primase. In eukaryotes, leading strand synthesis is thought to be conducted by Pol ε; however, this view has recently been challenged, suggesting a role for Pol δ. Primer removal is completed Pol δ while repair of DNA during replication is completed by Pol ε. As DNA synthesis continues, the original DNA strands continue to unwind on each side of the bubble, forming a
replication fork In molecular biology, DNA replication is the biological process of producing two identical replicas of DNA from one original DNA molecule. DNA replication occurs in all life, living organisms acting as the most essential part for heredity, biologi ...

replication fork
with two prongs. In bacteria, which have a single origin of replication on their circular chromosome, this process creates a " theta structure" (resembling the Greek letter theta: θ). In contrast, eukaryotes have longer linear chromosomes and initiate replication at multiple origins within these.


Replication fork

The replication fork is a structure that forms within the long helical DNA during DNA replication. It is created by helicases, which break the hydrogen bonds holding the two DNA strands together in the helix. The resulting structure has two branching "prongs", each one made up of a single strand of DNA. These two strands serve as the template for the leading and lagging strands, which will be created as DNA polymerase matches complementary nucleotides to the templates; the templates may be properly referred to as the leading strand template and the lagging strand template. DNA is read by DNA polymerase in the 3′ to 5′ direction, meaning the new strand is synthesized in the 5' to 3' direction. Since the leading and lagging strand templates are oriented in opposite directions at the replication fork, a major issue is how to achieve synthesis of new lagging strand DNA, whose direction of synthesis is opposite to the direction of the growing replication fork.


Leading strand

The leading strand is the strand of new DNA which is synthesized in the same direction as the growing replication fork. This sort of DNA replication is continuous.


Lagging strand

The lagging strand is the strand of new DNA whose direction of synthesis is opposite to the direction of the growing replication fork. Because of its orientation, replication of the lagging strand is more complicated as compared to that of the leading strand. As a consequence, the DNA polymerase on this strand is seen to "lag behind" the other strand. The lagging strand is synthesized in short, separated segments. On the lagging strand ''template'', a
primase DNA primase is an enzyme Enzymes () are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enz ...

primase
"reads" the template DNA and initiates synthesis of a short complementary
RNA Ribonucleic acid (RNA) is a polymer A polymer (; Greek ''wikt:poly-, poly-'', "many" + ''wikt:-mer, -mer'', "part") is a Chemical substance, substance or material consisting of very large molecules, or macromolecules, composed of many Re ...

RNA
primer. A DNA polymerase extends the primed segments, forming
Okazaki fragment Okazaki fragments are short sequences of DNA nucleotides (approximately 150 to 200 base pairs long in eukaryotes) which are synthesized discontinuously and later linked together by the enzyme DNA ligase to create the lagging strand during DNA rep ...
s. The RNA primers are then removed and replaced with DNA, and the fragments of DNA are joined by
DNA ligase DNA ligase is a specific type of enzyme, a ligase In biochemistry Biochemistry or biological chemistry, is the study of es within and relating to living s. A sub-discipline of both and , biochemistry may be divided into three field ...
.


Dynamics at the replication fork

In all cases the helicase is composed of six polypeptides that wrap around only one strand of the DNA being replicated. The two polymerases are bound to the helicase heximer. In eukaryotes the helicase wraps around the leading strand, and in prokaryotes it wraps around the lagging strand. As helicase unwinds DNA at the replication fork, the DNA ahead is forced to rotate. This process results in a build-up of twists in the DNA ahead. This build-up forms a torsional resistance that would eventually halt the progress of the replication fork. Topoisomerases are enzymes that temporarily break the strands of DNA, relieving the tension caused by unwinding the two strands of the DNA helix; topoisomerases (including
DNA gyrase DNA gyrase, or simply gyrase, is an enzyme Enzymes () are s that act as s (biocatalysts). Catalysts accelerate . The molecules upon which enzymes may act are called , and the enzyme converts the substrates into different molecules known as ...
) achieve this by adding negative supercoils to the DNA helix. Bare single-stranded DNA tends to fold back on itself forming
secondary structures Secondary may refer to: * Secondary (chemistry), term used in organic chemistry * Secondary (football), position in American football and Canadian football * Secondary, an obsolete name for the Mesozoic in geosciences * Secondary color, color made ...

secondary structures
; these structures can interfere with the movement of DNA polymerase. To prevent this,
single-strand binding protein Single-strand DNA-binding protein (SSB) is a protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture solved by X-ray crystallography by Max Perutz and Sir John ...
s bind to the DNA until a second strand is synthesized, preventing secondary structure formation. Double-stranded DNA is coiled around
histone In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, Physiology, physiological mechanisms ...
s that play an important role in regulating gene expression so the replicated DNA must be coiled around histones at the same places as the original DNA. To ensure this, histone chaperones disassemble the chromatin before it is replicated and replace the histones in the correct place. Some steps in this reassembly are somewhat speculative. DNA clamp, Clamp proteins form a sliding clamp around DNA, helping the DNA polymerase maintain contact with its template, thereby assisting with processivity. The inner face of the clamp enables DNA to be threaded through it. Once the polymerase reaches the end of the template or detects double-stranded DNA, the sliding clamp undergoes a conformational change that releases the DNA polymerase. Clamp-loading proteins are used to initially load the clamp, recognizing the junction between template and RNA primers.:274-5


DNA replication proteins

At the replication fork, many replication enzymes assemble on the DNA into a complex molecular machine called the replisome. The following is a list of major DNA replication enzymes that participate in the replisome:


Replication machinery

Replication machineries consist of factors involved in DNA replication and appearing on template ssDNAs. Replication machineries include primosotors are replication enzymes; DNA polymerase, DNA helicases, DNA clamps and DNA topoisomerases, and replication proteins; e.g. single-stranded DNA binding proteins (SSB). In the replication machineries these components coordinate. In most of the bacteria, all of the factors involved in DNA replication are located on replication forks and the complexes stay on the forks during DNA replication. These replication machineries are called replisomes or DNA replicase systems. These terms are generic terms for proteins located on replication forks. In eukaryotic and some bacterial cells the replisomes are not formed. Since replication machineries do not move relatively to template DNAs such as factories, they are called a replication factory.James D. Watson et al. (2008), "Molecular Biology of the gene", Pearson Education: 237 In an alternative figure, DNA factories are similar to projectors and DNAs are like as cinematic films passing constantly into the projectors. In the replication factory model, after both DNA helicases for leading strands and lagging strands are loaded on the template DNAs, the helicases run along the DNAs into each other. The helicases remain associated for the remainder of replication process. Peter Meister et al. observed directly replication sites in
budding yeast ''Saccharomyces cerevisiae'' () is a species of yeast Yeasts are eukaryotic Eukaryotes () are organism In biology, an organism () is any organic, life, living system that functions as an individual entity. All organisms are ...
by monitoring green fluorescent protein (GFP)-tagged DNA polymerases α. They detected DNA replication of pairs of the tagged loci spaced apart symmetrically from a replication origin and found that the distance between the pairs decreased markedly by time.Peter Meister, Angela Taddei1, Susan M. Gasser(June 2006), "In and out of the Replication Factory", ''Cell'' 125 (7): 1233–1235 This finding suggests that the mechanism of DNA replication goes with DNA factories. That is, couples of replication factories are loaded on replication origins and the factories associated with each other. Also, template DNAs move into the factories, which bring extrusion of the template ssDNAs and new DNAs. Meister's finding is the first direct evidence of replication factory model. Subsequent research has shown that DNA helicases form dimers in many eukaryotic cells and bacterial replication machineries stay in single intranuclear location during DNA synthesis. The replication factories perform disentanglement of sister chromatids. The disentanglement is essential for distributing the chromatids into daughter cells after DNA replication. Because sister chromatids after DNA replication hold each other by Cohesin rings, there is the only chance for the disentanglement in DNA replication. Fixing of replication machineries as replication factories can improve the success rate of DNA replication. If replication forks move freely in chromosomes, catenation of nuclei is aggravated and impedes mitotic segregation.


Termination

Eukaryotes initiate DNA replication at multiple points in the chromosome, so replication forks meet and terminate at many points in the chromosome. Because eukaryotes have linear chromosomes, DNA replication is unable to reach the very end of the chromosomes. Due to this problem, DNA is lost in each replication cycle from the end of the chromosome. Telomeres are regions of repetitive DNA close to the ends and help prevent loss of genes due to this shortening. Shortening of the telomeres is a normal process in somatic cells. This shortens the telomeres of the daughter DNA chromosome. As a result, cells can only divide a certain number of times before the DNA loss prevents further division. (This is known as the Hayflick limit.) Within the germ cell line, which passes DNA to the next generation, telomerase extends the repetitive sequences of the telomere region to prevent degradation. Telomerase can become mistakenly active in somatic cells, sometimes leading to cancer formation. Increased telomerase activity is one of the hallmarks of cancer. Termination requires that the progress of the DNA replication fork must stop or be blocked. Termination at a specific locus, when it occurs, involves the interaction between two components: (1) a termination site sequence in the DNA, and (2) a protein which binds to this sequence to physically stop DNA replication. In various bacterial species, this is named the DNA replication terminus site-binding protein, or Ter protein. Because bacteria have circular chromosomes, termination of replication occurs when the two replication forks meet each other on the opposite end of the parental chromosome. ''E. coli'' regulates this process through the use of termination sequences that, when bound by the Tus protein, enable only one direction of replication fork to pass through. As a result, the replication forks are constrained to always meet within the termination region of the chromosome.


Regulation


Eukaryotes

Within eukaryotes, DNA replication is controlled within the context of the cell cycle. As the cell grows and divides, it progresses through stages in the cell cycle; DNA replication takes place during the S phase (synthesis phase). The progress of the eukaryotic cell through the cycle is controlled by cell cycle checkpoints. Progression through checkpoints is controlled through complex interactions between various proteins, including
cyclin Cyclin is a family of proteins that controls the progression of a cell through the cell cycle The cell cycle, or cell-division cycle, is the series of events that take place in a cell Cell most often refers to: * Cell (biology), the funct ...
s and cyclin-dependent kinases. Unlike bacteria, eukaryotic DNA replicates in the confines of the nucleus. The G1/S checkpoint (or restriction checkpoint) regulates whether eukaryotic cells enter the process of DNA replication and subsequent division. Cells that do not proceed through this checkpoint remain in the G0 stage and do not replicate their DNA. After passing through the G1/S checkpoint, DNA must be replicated only once in each cell cycle. When the Mcm complex moves away from the origin, the pre-replication complex is dismantled. Because a new Mcm complex cannot be loaded at an origin until the pre-replication subunits are reactivated, one origin of replication can not be used twice in the same cell cycle. Activation of S-Cdks in early S phase promotes the destruction or inhibition of individual pre-replication complex components, preventing immediate reassembly. S and M-Cdks continue to block pre-replication complex assembly even after S phase is complete, ensuring that assembly cannot occur again until all Cdk activity is reduced in late mitosis. In budding yeast, inhibition of assembly is caused by Cdk-dependent phosphorylation of pre-replication complex components. At the onset of S phase, phosphorylation of Cdc6 by Cyclin-dependent kinase 1, Cdk1 causes the binding of Cdc6 to the SCF complex, SCF Ubiquitin--protein ligase, ubiquitin protein ligase, which causes proteolytic destruction of Cdc6. Cdk-dependent phosphorylation of Mcm proteins promotes their export out of the nucleus along with Cdt1 during S phase, preventing the loading of new Mcm complexes at origins during a single cell cycle. Cdk phosphorylation of the origin replication complex also inhibits pre-replication complex assembly. The individual presence of any of these three mechanisms is sufficient to inhibit pre-replication complex assembly. However, mutations of all three proteins in the same cell does trigger reinitiation at many origins of replication within one cell cycle. In animal cells, the protein geminin is a key inhibitor of pre-replication complex assembly. Geminin binds Cdt1, preventing its binding to the origin recognition complex. In G1, levels of geminin are kept low by the APC, which ubiquitinates geminin to target it for degradation. When geminin is destroyed, Cdt1 is released, allowing it to function in pre-replication complex assembly. At the end of G1, the APC is inactivated, allowing geminin to accumulate and bind Cdt1. Replication of chloroplast and mitochondrial genomes occurs independently of the cell cycle, through the process of D-loop replication.


Replication focus

In vertebrate cells, replication sites concentrate into positions called replication foci. Replication sites can be detected by immunostaining daughter strands and replication enzymes and monitoring GFP-tagged replication factors. By these methods it is found that replication foci of varying size and positions appear in S phase of cell division and their number per nucleus is far smaller than the number of genomic replication forks. P. Heun et al.,(2001) tracked GFP-tagged replication foci in budding yeast cells and revealed that replication origins move constantly in G1 and S phase and the Molecular dynamics, dynamics decreased significantly in S phase. Traditionally, replication sites were fixed on spatial structure of chromosomes by nuclear matrix or lamins. The Heun's results denied the traditional concepts, budding yeasts do not have lamins, and support that replication origins self-assemble and form replication foci. By firing of replication origins, controlled spatially and temporally, the formation of replication foci is regulated. D. A. Jackson et al.(1998) revealed that neighboring origins fire simultaneously in mammalian cells. Spatial juxtaposition of replication sites brings clustering of replication forks. The clustering do rescue of stalled replication forks and favors normal progress of replication forks. Progress of replication forks is inhibited by many factors; collision with proteins or with complexes binding strongly on DNA, deficiency of dNTPs, nicks on template DNAs and so on. If replication forks stall and the remaining sequences from the stalled forks are not replicated, the daughter strands have nick obtained un-replicated sites. The un-replicated sites on one parent's strand hold the other strand together but not daughter strands. Therefore, the resulting sister chromatids cannot separate from each other and cannot divide into 2 daughter cells. When neighboring origins fire and a fork from one origin is stalled, fork from other origin access on an opposite direction of the stalled fork and duplicate the un-replicated sites. As other mechanism of the rescue there is application of dormant replication origins that excess origins do not fire in normal DNA replication.


Bacteria

Most bacteria do not go through a well-defined cell cycle but instead continuously copy their DNA; during rapid growth, this can result in the concurrent occurrence of multiple rounds of replication. In ''E. coli'', the best-characterized bacteria, DNA replication is regulated through several mechanisms, including: the hemimethylation and sequestering of the origin sequence, the ratio of Adenosine triphosphate, adenosine triphosphate (ATP) to Adenosine diphosphate, adenosine diphosphate (ADP), and the levels of protein DnaA. All these control the binding of initiator proteins to the origin sequences. Because ''E. coli'' DNA methylation, methylates GATC DNA sequences, DNA synthesis results in hemimethylated sequences. This hemimethylated DNA is recognized by the protein SeqA protein domain, SeqA, which binds and sequesters the origin sequence; in addition, DnaA (required for initiation of replication) binds less well to hemimethylated DNA. As a result, newly replicated origins are prevented from immediately initiating another round of DNA replication. ATP builds up when the cell is in a rich medium, triggering DNA replication once the cell has reached a specific size. ATP competes with ADP to bind to DnaA, and the DnaA-ATP complex is able to initiate replication. A certain number of DnaA proteins are also required for DNA replication — each time the origin is copied, the number of binding sites for DnaA doubles, requiring the synthesis of more DnaA to enable another initiation of replication. In fast-growing bacteria, such as ''E. coli'', chromosome replication takes more time than dividing the cell. The bacteria solve this by initiating a new round of replication before the previous one has been terminated. The new round of replication will form the chromosome of the cell that is born two generations after the dividing cell. This mechanism creates overlapping replication cycles.


Problems with DNA replication

There are many events that contribute to replication stress, including: * Misincorporation of ribonucleotides * Unusual Nucleic acid secondary structure, DNA structures * Conflicts between replication and transcription * Insufficiency of essential replication factors * Chromosomal fragile site, Common fragile sites * Overexpression or constitutive activation of oncogenes * Chromatin inaccessibility


Polymerase chain reaction

Researchers commonly replicate DNA ''in vitro'' using the polymerase chain reaction (PCR). PCR uses a pair of primer (molecular biology), primers to span a target region in template DNA, and then polymerizes partner strands in each direction from these primers using a thermostable
DNA polymerase A DNA polymerase is a member of a family of enzyme Enzymes () are s that act as s (biocatalysts). Catalysts accelerate . The molecules upon which enzymes may act are called , and the enzyme converts the substrates into different molecules ...

DNA polymerase
. Repeating this process through multiple cycles amplifies the targeted DNA region. At the start of each cycle, the mixture of template and primers is heated, separating the newly synthesized molecule and template. Then, as the mixture cools, both of these become templates for annealing of new primers, and the polymerase extends from these. As a result, the number of copies of the target region doubles each round, exponential growth, increasing exponentially.


See also

* Life * Cell (biology) * Cell division * Gene * Gene expression * Epigenetics * Genome * Autopoiesis * Chromosome segregation * Data storage device * Self-replication * Hachimoji DNA


Notes


References

{{DEFAULTSORT:Dna Replication DNA replication Senescence Cellular processes Molecular biology Copying