Cloning - Revision history

Bradley Monk: /* Restriction Endonuclease */

2015-12-11T09:24:53Z

Restriction Endonuclease

← Older revision		Revision as of 02:24, 11 December 2015
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	==Restriction Endonuclease==		==Restriction Endonuclease==
			[[File:BamHI2.png\|thumb\|450px\|Cleavage of a DNA sequence containing the BamHI restriction site. The DNA is cleaved at the palindromic sequence to produce 'sticky ends'.]]
	A [https://en.wikipedia.org/wiki/Restriction_enzyme restriction enzyme] or restriction endonuclease is an enzyme that cuts DNA at or near specific recognition nucleotide sequences known as restriction sites. Restriction enzymes are commonly classified into three types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone strand of the DNA double helix.		A [https://en.wikipedia.org/wiki/Restriction_enzyme restriction enzyme] or restriction endonuclease is an enzyme that cuts DNA at or near specific recognition nucleotide sequences known as restriction sites. Restriction enzymes are commonly classified into three types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone strand of the DNA double helix.

	Over 3000 restriction enzymes have been studied in detail, and more than 600 of these are available commercially. These enzymes are routinely used for DNA modification in laboratories, and are a vital tool in [https://en.wikipedia.org/wiki/Molecular_cloning molecular cloning].		Over 3000 restriction enzymes have been studied in detail, and more than 600 of these are available commercially. These enzymes are routinely used for DNA modification in laboratories, and are a vital tool in [https://en.wikipedia.org/wiki/Molecular_cloning molecular cloning].







	==Example Clone==		==Example Clone==

Bradley Monk: /* Plasmid Vector */

2015-12-11T06:07:29Z

Plasmid Vector

← Older revision		Revision as of 23:07, 10 December 2015
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	A cloning vector almost always contains four DNA segments critically important to its function and experimental utility:		A cloning vector almost always contains four DNA segments critically important to its function and experimental utility:
	# DNA ''replication origin'' is necessary for the vector (and its linked recombinant sequences) to replicate inside the host organism		# DNA '''''replication origin''''' is necessary for the vector (and its linked recombinant sequences) to replicate inside the host organism
	# one or more unique ''restriction endonuclease recognition sites'' to serve as sites where foreign DNA may be introduced		# one or more unique '''''restriction endonuclease recognition sites''''' to serve as sites where foreign DNA may be introduced
	# a ''selectable genetic marker'' gene that can be used to enable the survival of cells that have taken up vector sequences		# a '''''selectable genetic marker''''' gene that can be used to enable the survival of cells that have taken up vector sequences
	# a ''tag'' gene that can be used to screen for cells containing the foreign DNA		# a '''''tag''''' gene that can be used to screen for cells containing the foreign DNA

	==Restriction Endonuclease==		==Restriction Endonuclease==

Bradley Monk: /* Plasmid Vector */

2015-12-11T06:06:17Z

Plasmid Vector

← Older revision		Revision as of 23:06, 10 December 2015
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	~~Whatever combination of host and vector are used, the~~ vector almost always contains four DNA segments ~~that are~~ critically important to its function and experimental utility:		A cloning vector almost always contains four DNA segments critically important to its function and experimental utility:
	# DNA ''replication origin'' is necessary for the vector (and its linked recombinant sequences) to replicate inside the host organism		# DNA ''replication origin'' is necessary for the vector (and its linked recombinant sequences) to replicate inside the host organism
	# one or more unique ''restriction endonuclease recognition sites'' to serve as sites where foreign DNA may be introduced		# one or more unique ''restriction endonuclease recognition sites'' to serve as sites where foreign DNA may be introduced
	# a ''selectable genetic marker'' gene that can be used to enable the survival of cells that have taken up vector sequences		# a ''selectable genetic marker'' gene that can be used to enable the survival of cells that have taken up vector sequences
	# a ''tag'' gene that can be used to screen for cells containing the foreign DNA		# a ''tag'' gene that can be used to screen for cells containing the foreign DNA




	==Restriction Endonuclease==		==Restriction Endonuclease==

Bradley Monk at 06:04, 11 December 2015

2015-12-11T06:04:36Z

← Older revision		Revision as of 23:04, 10 December 2015
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			Whatever combination of host and vector are used, the vector almost always contains four DNA segments that are critically important to its function and experimental utility:
			# DNA ''replication origin'' is necessary for the vector (and its linked recombinant sequences) to replicate inside the host organism
			# one or more unique ''restriction endonuclease recognition sites'' to serve as sites where foreign DNA may be introduced
			# a ''selectable genetic marker'' gene that can be used to enable the survival of cells that have taken up vector sequences
			# a ''tag'' gene that can be used to screen for cells containing the foreign DNA


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	==Example Clone==		==Example Clone==

			{{ExpandBox\|width=90%\|float=left\|opener=View\|Recombinant DNA\|

	{{Style\|font=courier\|color=#000000\|		{{Style\|font=courier\|color=#000000\|
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	}}		}}

			}}


	[[Category:Neurobiology]] [[Category:Synaptic Plasticity]] [[Category:Neuroinformatics]] [[Category:Genetics]] [[Category:Neuroscience Methods]] [[Category:Methods]]		[[Category:Neurobiology]] [[Category:Synaptic Plasticity]] [[Category:Neuroinformatics]] [[Category:Genetics]] [[Category:Neuroscience Methods]] [[Category:Methods]]

Bradley Monk at 05:47, 11 December 2015

2015-12-11T05:47:48Z

Show changes

Bradley Monk: /* Plasmid Vector */

2015-12-11T05:27:02Z

Plasmid Vector

← Older revision		Revision as of 22:27, 10 December 2015
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	A [https://en.wikipedia.org/wiki/Cloning_vector cloning vector] is a small piece of DNA, taken from a virus, a plasmid, or the cell of a higher organism, that can be stably maintained in an organism, and into which a foreign DNA fragment can be inserted for cloning purposes. The vector therefore contains features that allow for the convenient insertion or removal of DNA fragment in or out of the vector, for example by treating the vector and the foreign DNA with a restriction enzyme that creates the same overhang, then ligating the fragments together. After a DNA fragment has been cloned into a cloning vector, it may be further subcloned into another vector designed for more specific use.		A [https://en.wikipedia.org/wiki/Cloning_vector cloning vector] is a small piece of DNA, taken from a virus, a plasmid, or the cell of a higher organism, that can be stably maintained in an organism, and into which a foreign DNA fragment can be inserted for cloning purposes. The vector therefore contains features that allow for the convenient insertion or removal of DNA fragment in or out of the vector, for example by treating the vector and the foreign DNA with a restriction enzyme that creates the same overhang, then ligating the fragments together. After a DNA fragment has been cloned into a cloning vector, it may be further subcloned into another vector designed for more specific use.


			Specialized applications may call for specialized host-vector systems. For example, if the experimentalists wish to harvest a particular protein from the recombinant organism, then an [https://en.wikipedia.org/wiki/Expression_vector expression vector] is chosen that contains appropriate signals for transcription and translation in the desired host organism. Alternatively, if replication of the DNA in different species is desired (for example, transfer of DNA from bacteria to plants), then a multiple host range vector (also termed shuttle vector) may be selected. In practice, however, specialized molecular cloning experiments usually begin with cloning into a bacterial plasmid, followed by subcloning into a specialized vector.

	==Example Clone==		==Example Clone==

Bradley Monk at 05:24, 11 December 2015

2015-12-11T05:24:23Z

← Older revision		Revision as of 22:24, 10 December 2015
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	==Plasmid Vector==		==Plasmid Vector==
	[[Plasmid]]		'''see also: [[Plasmid]]'''


			A [https://en.wikipedia.org/wiki/Cloning_vector cloning vector] is a small piece of DNA, taken from a virus, a plasmid, or the cell of a higher organism, that can be stably maintained in an organism, and into which a foreign DNA fragment can be inserted for cloning purposes. The vector therefore contains features that allow for the convenient insertion or removal of DNA fragment in or out of the vector, for example by treating the vector and the foreign DNA with a restriction enzyme that creates the same overhang, then ligating the fragments together. After a DNA fragment has been cloned into a cloning vector, it may be further subcloned into another vector designed for more specific use.

	==Example Clone==		==Example Clone==

Bradley Monk at 04:17, 11 December 2015

2015-12-11T04:17:13Z

← Older revision		Revision as of 21:17, 10 December 2015
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	[https://en.wikipedia.org/wiki/Molecular_cloning Molecular cloning] is a set of experimental methods in molecular biology that are used to assemble recombinant DNA molecules and to direct their replication within host organisms. The use of the word cloning refers to the fact that the method involves the replication of one molecule to produce a population of cells with identical DNA molecules. Molecular cloning generally uses DNA sequences from two different organisms: the species that is the source of the DNA to be cloned, and the species that will serve as the living host for replication of the recombinant DNA. Molecular cloning methods are central to many contemporary areas of modern biology and medicine.		[https://en.wikipedia.org/wiki/Molecular_cloning Molecular cloning] is a set of experimental methods in molecular biology that are used to assemble recombinant DNA molecules and to direct their replication within host organisms. The use of the word cloning refers to the fact that the method involves the replication of one molecule to produce a population of cells with identical DNA molecules. Molecular cloning generally uses DNA sequences from two different organisms: the species that is the source of the DNA to be cloned, and the species that will serve as the living host for replication of the recombinant DNA. Molecular cloning methods are central to many contemporary areas of modern biology and medicine.

			Molecular cloning is similar to [https://en.wikipedia.org/wiki/Polymerase_chain_reaction polymerase chain reaction (PCR)] in that it permits the replication of DNA sequence. The fundamental difference between the two methods is that molecular cloning involves replication of the DNA in a living microorganism, while PCR replicates DNA in an in vitro solution, free of living cells.

Bradley Monk at 04:11, 11 December 2015

2015-12-11T04:11:44Z

← Older revision		Revision as of 21:11, 10 December 2015
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	==Molecular ~~Cloning==~~		[https://en.wikipedia.org/wiki/Molecular_cloning Molecular cloning] is a set of experimental methods in molecular biology that are used to assemble recombinant DNA molecules and to direct their replication within host organisms. The use of the word cloning refers to the fact that the method involves the replication of one molecule to produce a population of cells with identical DNA molecules. Molecular cloning generally uses DNA sequences from two different organisms: the species that is the source of the DNA to be cloned, and the species that will serve as the living host for replication of the recombinant DNA. Molecular cloning methods are central to many contemporary areas of modern biology and medicine.


			==Restriction Endonuclease==

	==~~restriction endonuclease~~==
	A [https://en.wikipedia.org/wiki/Restriction_enzyme restriction enzyme] or restriction endonuclease is an enzyme that cuts DNA at or near specific recognition nucleotide sequences known as restriction sites. Restriction enzymes are commonly classified into three types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone strand of the DNA double helix.		A [https://en.wikipedia.org/wiki/Restriction_enzyme restriction enzyme] or restriction endonuclease is an enzyme that cuts DNA at or near specific recognition nucleotide sequences known as restriction sites. Restriction enzymes are commonly classified into three types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone strand of the DNA double helix.

	Over 3000 restriction enzymes have been studied in detail, and more than 600 of these are available commercially.~~[7]~~ These enzymes are routinely used for DNA modification in laboratories, and are a vital tool in [https://en.wikipedia.org/wiki/Molecular_cloning molecular cloning].		Over 3000 restriction enzymes have been studied in detail, and more than 600 of these are available commercially. These enzymes are routinely used for DNA modification in laboratories, and are a vital tool in [https://en.wikipedia.org/wiki/Molecular_cloning molecular cloning].

	==Plasmid Vector==		==Plasmid Vector==

Bradley Monk: /* restriction endonuclease */

2015-12-11T04:09:34Z

restriction endonuclease

← Older revision		Revision as of 21:09, 10 December 2015
Line 6:		Line 6:
	==restriction endonuclease==		==restriction endonuclease==
	A [https://en.wikipedia.org/wiki/Restriction_enzyme restriction enzyme] or restriction endonuclease is an enzyme that cuts DNA at or near specific recognition nucleotide sequences known as restriction sites. Restriction enzymes are commonly classified into three types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone strand of the DNA double helix.		A [https://en.wikipedia.org/wiki/Restriction_enzyme restriction enzyme] or restriction endonuclease is an enzyme that cuts DNA at or near specific recognition nucleotide sequences known as restriction sites. Restriction enzymes are commonly classified into three types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone strand of the DNA double helix.

			Over 3000 restriction enzymes have been studied in detail, and more than 600 of these are available commercially.[7] These enzymes are routinely used for DNA modification in laboratories, and are a vital tool in [https://en.wikipedia.org/wiki/Molecular_cloning molecular cloning].

	==Plasmid Vector==		==Plasmid Vector==