File:DNA targeting platforms genome editing.jpg - Revision history

Z8600021: /* Common DNA Targeting Platforms for Genome Editing */

2017-07-28T01:13:43Z

Common DNA Targeting Platforms for Genome Editing

← Older revision		Revision as of 12:13, 28 July 2017
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	{\|		{\|
	! ~~colwidth~~=400px\|Zinc finger nucleases		! width=400px\|Zinc finger nucleases
	! ~~colwidth~~=400px\|Meganucleases		! width=400px\|Meganucleases
	\|-		\|-
	\| valign=top\|Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.		\| valign=top\|Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.

Z8600021: /* Common DNA Targeting Platforms for Genome Editing */

2017-07-28T01:13:19Z

Common DNA Targeting Platforms for Genome Editing

← Older revision		Revision as of 12:13, 28 July 2017
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	{\|		{\|
	! Zinc finger nucleases		! colwidth=400px\|Zinc finger nucleases
	! ~~TALENs~~		! colwidth=400px\|Meganucleases
	\|-		\|-
	\| Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.		\| valign=top\|Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.
	\| ~~The discovery of a simple one~~-~~to-one code dictating~~ the DNA-binding specificity of ~~TALE proteins from~~ the ~~plant pathogen Xanthomonas again raised~~ the ~~exciting possibility for modular design of novel DNA~~-~~binding proteins~~.~~114,115 Highly conserved 33–35 amino acid TALE repeats each bind~~ a ~~single base pair~~ of DNA with specificity dictated by two hypervariable residues. Crystal structures of TALEs bound to DNA revealed that each repeat forms a two-helix structure connected by a loop which presents the hypervariable residue into the major groove as the protein wraps around the DNA in a superhelical structure. These modular TALE repeats can be ~~linked together~~ to ~~build long arrays with custom DNA-binding specificities~~.		\| valign=top\|Meganuclease technology involves re-engineering the DNA-binding specificity of naturally occurring homing endonucleases. The largest class of homing endonucleases is the LAGLIDADG family, which includes the well-characterized and commonly used I-CreI and I-SceI enzymes.140 Through a combination of rational design and selection, these homing endonucleases can be re-engineered to target novel sequences.
	\|-		\|-
	! ~~Meganucleases~~		! TALENs
	! CRISPR/Cas nucleases		! CRISPR/Cas nucleases
	\|-		\|-
	\| ~~Meganuclease technology involves re~~-~~engineering~~ the DNA-binding specificity of ~~naturally occurring homing endonucleases. The largest class of homing endonucleases is~~ the ~~LAGLIDADG family, which includes~~ the ~~well~~-~~characterized and commonly used I-CreI and I-SceI enzymes~~.~~140 Through~~ a ~~combination~~ of ~~rational design and selection, these homing endonucleases~~ can be re-~~engineered to target novel sequences~~.		\| valign=top\|The discovery of a simple one-to-one code dictating the DNA-binding specificity of TALE proteins from the plant pathogen Xanthomonas again raised the exciting possibility for modular design of novel DNA-binding proteins.114,115 Highly conserved 33–35 amino acid TALE repeats each bind a single base pair of DNA with specificity dictated by two hypervariable residues. Crystal structures of TALEs bound to DNA revealed that each repeat forms a two-helix structure connected by a loop which presents the hypervariable residue into the major groove as the protein wraps around the DNA in a superhelical structure. These modular TALE repeats can be linked together to build long arrays with custom DNA-binding specificities.
	\| CRISPR-Cas RNA-guided nucleases are derived from an adaptive immune system that evolved in bacteria to defend against invading plasmids and viruses. Decades of work investigating CRISPR systems in various microbial species has elucidated a mechanism by which short sequences of invading nucleic acids are incorporated into CRISPR loci. They are then transcribed and processed into CRISPR RNAs (crRNAs) which, together with a trans-activating crRNAs (tracrRNAs), complex with CRISPR-associated (Cas) proteins to dictate specificity of DNA cleavage by Cas nucleases through Watson-Crick base pairing between nucleic acids. Building off of two studies showing that the three components required for the type II CRISPR nuclease system are the Cas9 protein, the mature crRNA and the tracrRNA, Doudna, Charpentier and colleagues showed through in vitro DNA cleavage experiments that this system could be reduced to two components by fusion of the crRNA and tracrRNA into a single guide RNA (gRNA). Furthermore, they showed that re-targeting of the Cas9/gRNA complex to new sites could be accomplished by altering the sequence of a short portion of the gRNA. Thereafter, a series of publications demonstrated that the CRISPR/Cas9 system could be engineered for efficient genetic modification in mammalian cells. Collectively these studies have propelled the CRISPR/Cas9 technology into the spotlight of the genome-editing field.		\| valign=top\|CRISPR-Cas RNA-guided nucleases are derived from an adaptive immune system that evolved in bacteria to defend against invading plasmids and viruses. Decades of work investigating CRISPR systems in various microbial species has elucidated a mechanism by which short sequences of invading nucleic acids are incorporated into CRISPR loci. They are then transcribed and processed into CRISPR RNAs (crRNAs) which, together with a trans-activating crRNAs (tracrRNAs), complex with CRISPR-associated (Cas) proteins to dictate specificity of DNA cleavage by Cas nucleases through Watson-Crick base pairing between nucleic acids. Building off of two studies showing that the three components required for the type II CRISPR nuclease system are the Cas9 protein, the mature crRNA and the tracrRNA, Doudna, Charpentier and colleagues showed through in vitro DNA cleavage experiments that this system could be reduced to two components by fusion of the crRNA and tracrRNA into a single guide RNA (gRNA). Furthermore, they showed that re-targeting of the Cas9/gRNA complex to new sites could be accomplished by altering the sequence of a short portion of the gRNA. Thereafter, a series of publications demonstrated that the CRISPR/Cas9 system could be engineered for efficient genetic modification in mammalian cells. Collectively these studies have propelled the CRISPR/Cas9 technology into the spotlight of the genome-editing field.
	\|}		\|}

Z8600021 at 01:09, 28 July 2017

2017-07-28T01:09:50Z

← Older revision		Revision as of 12:09, 28 July 2017
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	==Common DNA Targeting Platforms for Genome Editing==		==Common DNA Targeting Platforms for Genome Editing==

			{\|
	~~'''~~Zinc finger nucleases~~'''~~		! Zinc finger nucleases
			! TALENs
	Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.		\|-
			\| Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.
			\| The discovery of a simple one-to-one code dictating the DNA-binding specificity of TALE proteins from the plant pathogen Xanthomonas again raised the exciting possibility for modular design of novel DNA-binding proteins.114,115 Highly conserved 33–35 amino acid TALE repeats each bind a single base pair of DNA with specificity dictated by two hypervariable residues. Crystal structures of TALEs bound to DNA revealed that each repeat forms a two-helix structure connected by a loop which presents the hypervariable residue into the major groove as the protein wraps around the DNA in a superhelical structure. These modular TALE repeats can be linked together to build long arrays with custom DNA-binding specificities.
	~~'''TALENs'''~~		\|-
			! Meganucleases
	The discovery of a simple one-to-one code dictating the DNA-binding specificity of TALE proteins from the plant pathogen Xanthomonas again raised the exciting possibility for modular design of novel DNA-binding proteins.114,115 Highly conserved 33–35 amino acid TALE repeats each bind a single base pair of DNA with specificity dictated by two hypervariable residues. Crystal structures of TALEs bound to DNA revealed that each repeat forms a two-helix structure connected by a loop which presents the hypervariable residue into the major groove as the protein wraps around the DNA in a superhelical structure. These modular TALE repeats can be linked together to build long arrays with custom DNA-binding specificities.		! CRISPR/Cas nucleases
			\|-
			\| Meganuclease technology involves re-engineering the DNA-binding specificity of naturally occurring homing endonucleases. The largest class of homing endonucleases is the LAGLIDADG family, which includes the well-characterized and commonly used I-CreI and I-SceI enzymes.140 Through a combination of rational design and selection, these homing endonucleases can be re-engineered to target novel sequences.
	~~'''Meganucleases'''~~		\| CRISPR-Cas RNA-guided nucleases are derived from an adaptive immune system that evolved in bacteria to defend against invading plasmids and viruses. Decades of work investigating CRISPR systems in various microbial species has elucidated a mechanism by which short sequences of invading nucleic acids are incorporated into CRISPR loci. They are then transcribed and processed into CRISPR RNAs (crRNAs) which, together with a trans-activating crRNAs (tracrRNAs), complex with CRISPR-associated (Cas) proteins to dictate specificity of DNA cleavage by Cas nucleases through Watson-Crick base pairing between nucleic acids. Building off of two studies showing that the three components required for the type II CRISPR nuclease system are the Cas9 protein, the mature crRNA and the tracrRNA, Doudna, Charpentier and colleagues showed through in vitro DNA cleavage experiments that this system could be reduced to two components by fusion of the crRNA and tracrRNA into a single guide RNA (gRNA). Furthermore, they showed that re-targeting of the Cas9/gRNA complex to new sites could be accomplished by altering the sequence of a short portion of the gRNA. Thereafter, a series of publications demonstrated that the CRISPR/Cas9 system could be engineered for efficient genetic modification in mammalian cells. Collectively these studies have propelled the CRISPR/Cas9 technology into the spotlight of the genome-editing field.
			\|}
	Meganuclease technology involves re-engineering the DNA-binding specificity of naturally occurring homing endonucleases. The largest class of homing endonucleases is the LAGLIDADG family, which includes the well-characterized and commonly used I-CreI and I-SceI enzymes.140 Through a combination of rational design and selection, these homing endonucleases can be re-engineered to target novel sequences.


	~~'''CRISPR/Cas nucleases'''~~

	CRISPR-Cas RNA-guided nucleases are derived from an adaptive immune system that evolved in bacteria to defend against invading plasmids and viruses. Decades of work investigating CRISPR systems in various microbial species has elucidated a mechanism by which short sequences of invading nucleic acids are incorporated into CRISPR loci. They are then transcribed and processed into CRISPR RNAs (crRNAs) which, together with a trans-activating crRNAs (tracrRNAs), complex with CRISPR-associated (Cas) proteins to dictate specificity of DNA cleavage by Cas nucleases through Watson-Crick base pairing between nucleic acids. Building off of two studies showing that the three components required for the type II CRISPR nuclease system are the Cas9 protein, the mature crRNA and the tracrRNA, Doudna, Charpentier and colleagues showed through in vitro DNA cleavage experiments that this system could be reduced to two components by fusion of the crRNA and tracrRNA into a single guide RNA (gRNA). Furthermore, they showed that re-targeting of the Cas9/gRNA complex to new sites could be accomplished by altering the sequence of a short portion of the gRNA. Thereafter, a series of publications demonstrated that the CRISPR/Cas9 system could be engineered for efficient genetic modification in mammalian cells. Collectively these studies have propelled the CRISPR/Cas9 technology into the spotlight of the genome-editing field.


	(text extract above from original article)		(text extract above from original article)
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	===Reference===		===Reference===

	<pubmed>26755333 </pubmed>		<pubmed>26755333</pubmed>

	====Copyright====		====Copyright====

	This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.		This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

	Figure 2		Figure 2

	{{Footer}}		{{Footer}}
			[[Category:Molecular]][[Category:Cartoon]][[Category:Genetics]]

Z8600021 at 01:07, 28 July 2017

2017-07-28T01:07:49Z

← Older revision		Revision as of 12:07, 28 July 2017
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	(text extract above from original article)		(text extract above from original article)


			{{Molecular Links}}

			{{Genetic}}

	===Reference===		===Reference===

Z8600021: ==Common DNA Targeting Platforms for Genome Editing== '''Zinc finger nucleases''' Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encod...

2017-07-28T01:06:29Z

New page

==Common DNA Targeting Platforms for Genome Editing==

'''Zinc finger nucleases'''

Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.

'''TALENs'''

The discovery of a simple one-to-one code dictating the DNA-binding specificity of TALE proteins from the plant pathogen Xanthomonas again raised the exciting possibility for modular design of novel DNA-binding proteins.114,115 Highly conserved 33–35 amino acid TALE repeats each bind a single base pair of DNA with specificity dictated by two hypervariable residues. Crystal structures of TALEs bound to DNA revealed that each repeat forms a two-helix structure connected by a loop which presents the hypervariable residue into the major groove as the protein wraps around the DNA in a superhelical structure. These modular TALE repeats can be linked together to build long arrays with custom DNA-binding specificities.

'''Meganucleases'''

Meganuclease technology involves re-engineering the DNA-binding specificity of naturally occurring homing endonucleases. The largest class of homing endonucleases is the LAGLIDADG family, which includes the well-characterized and commonly used I-CreI and I-SceI enzymes.140 Through a combination of rational design and selection, these homing endonucleases can be re-engineered to target novel sequences.

'''CRISPR/Cas nucleases'''

CRISPR-Cas RNA-guided nucleases are derived from an adaptive immune system that evolved in bacteria to defend against invading plasmids and viruses. Decades of work investigating CRISPR systems in various microbial species has elucidated a mechanism by which short sequences of invading nucleic acids are incorporated into CRISPR loci. They are then transcribed and processed into CRISPR RNAs (crRNAs) which, together with a trans-activating crRNAs (tracrRNAs), complex with CRISPR-associated (Cas) proteins to dictate specificity of DNA cleavage by Cas nucleases through Watson-Crick base pairing between nucleic acids. Building off of two studies showing that the three components required for the type II CRISPR nuclease system are the Cas9 protein, the mature crRNA and the tracrRNA, Doudna, Charpentier and colleagues showed through in vitro DNA cleavage experiments that this system could be reduced to two components by fusion of the crRNA and tracrRNA into a single guide RNA (gRNA). Furthermore, they showed that re-targeting of the Cas9/gRNA complex to new sites could be accomplished by altering the sequence of a short portion of the gRNA. Thereafter, a series of publications demonstrated that the CRISPR/Cas9 system could be engineered for efficient genetic modification in mammalian cells. Collectively these studies have propelled the CRISPR/Cas9 technology into the spotlight of the genome-editing field.

(text extract above from original article)
===Reference===

<pubmed>26755333 </pubmed>

====Copyright====

This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Figure 2
{{Footer}}

← Older revision		Revision as of 12:13, 28 July 2017
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	{\|		{\|
	! ~~colwidth~~=400px\|Zinc finger nucleases		! width=400px\|Zinc finger nucleases
	! ~~colwidth~~=400px\|Meganucleases		! width=400px\|Meganucleases
	\|-		\|-
	\| valign=top\|Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.		\| valign=top\|Zinc finger (ZF) proteins are the most abundant class of transcription factors and the Cys2-His2 zinc finger domain is one of the most common DNA-binding domains encoded in the human genome. The crystal structure of Zif268 has served as the basis for understanding DNA recognition by zinc fingers. In the presence of a zinc atom, the zinc finger domain forms a compact ββα structure with the α-helical portion of each finger making contact with 3 or 4 bp in the major groove of the DNA. Tandem fingers in a zinc finger array wrap around the DNA to bind extended target sequences such that a three-finger protein binds a 9 bp target site.