Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications

Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find...
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Autor*in:	Friedland, Ari E. [verfasserIn] Baral, Reshica Singhal, Pankhuri Loveluck, Katherine Shen, Shen Sanchez, Minerva Marco, Eugenio Gotta, Gregory M. Maeder, Morgan L. Kennedy, Edward M. Kornepati, Anand V. R. Sousa, Alexander Collins, McKensie A. Jayaram, Hari Cullen, Bryan R. Bumcrot, David

Format:	E-Artikel
Sprache:	Englisch

Erschienen:	2015

Schlagwörter:	CRISPR Cas9 Genome engineering Nickases Adeno-associated virus GUIDE-seq

Anmerkung:	© Friedland et al. 2015

Übergeordnetes Werk:	Enthalten in: Genome biology - London : BioMed Central, 2000, 16(2015), 1 vom: 24. Nov.
Übergeordnetes Werk:	volume:16 ; year:2015 ; number:1 ; day:24 ; month:11

Links:	Volltext

DOI / URN:	10.1186/s13059-015-0817-8

Katalog-ID:	SPR030024714

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520			\|a Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors.
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allfields	10.1186/s13059-015-0817-8 doi (DE-627)SPR030024714 (SPR)s13059-015-0817-8-e DE-627 ger DE-627 rakwb eng Friedland, Ari E. verfasserin aut Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Friedland et al. 2015 Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. CRISPR (dpeaa)DE-He213 Cas9 (dpeaa)DE-He213 Genome engineering (dpeaa)DE-He213 Nickases (dpeaa)DE-He213 Adeno-associated virus (dpeaa)DE-He213 GUIDE-seq (dpeaa)DE-He213 Baral, Reshica aut Singhal, Pankhuri aut Loveluck, Katherine aut Shen, Shen aut Sanchez, Minerva aut Marco, Eugenio aut Gotta, Gregory M. aut Maeder, Morgan L. aut Kennedy, Edward M. aut Kornepati, Anand V. R. aut Sousa, Alexander aut Collins, McKensie A. aut Jayaram, Hari aut Cullen, Bryan R. aut Bumcrot, David aut Enthalten in Genome biology London : BioMed Central, 2000 16(2015), 1 vom: 24. Nov. (DE-627)326173617 (DE-600)2040529-7 1474-760X nnns volume:16 year:2015 number:1 day:24 month:11 https://dx.doi.org/10.1186/s13059-015-0817-8 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 16 2015 1 24 11
spelling	10.1186/s13059-015-0817-8 doi (DE-627)SPR030024714 (SPR)s13059-015-0817-8-e DE-627 ger DE-627 rakwb eng Friedland, Ari E. verfasserin aut Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Friedland et al. 2015 Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. CRISPR (dpeaa)DE-He213 Cas9 (dpeaa)DE-He213 Genome engineering (dpeaa)DE-He213 Nickases (dpeaa)DE-He213 Adeno-associated virus (dpeaa)DE-He213 GUIDE-seq (dpeaa)DE-He213 Baral, Reshica aut Singhal, Pankhuri aut Loveluck, Katherine aut Shen, Shen aut Sanchez, Minerva aut Marco, Eugenio aut Gotta, Gregory M. aut Maeder, Morgan L. aut Kennedy, Edward M. aut Kornepati, Anand V. R. aut Sousa, Alexander aut Collins, McKensie A. aut Jayaram, Hari aut Cullen, Bryan R. aut Bumcrot, David aut Enthalten in Genome biology London : BioMed Central, 2000 16(2015), 1 vom: 24. Nov. (DE-627)326173617 (DE-600)2040529-7 1474-760X nnns volume:16 year:2015 number:1 day:24 month:11 https://dx.doi.org/10.1186/s13059-015-0817-8 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 16 2015 1 24 11
allfields_unstemmed	10.1186/s13059-015-0817-8 doi (DE-627)SPR030024714 (SPR)s13059-015-0817-8-e DE-627 ger DE-627 rakwb eng Friedland, Ari E. verfasserin aut Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Friedland et al. 2015 Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. CRISPR (dpeaa)DE-He213 Cas9 (dpeaa)DE-He213 Genome engineering (dpeaa)DE-He213 Nickases (dpeaa)DE-He213 Adeno-associated virus (dpeaa)DE-He213 GUIDE-seq (dpeaa)DE-He213 Baral, Reshica aut Singhal, Pankhuri aut Loveluck, Katherine aut Shen, Shen aut Sanchez, Minerva aut Marco, Eugenio aut Gotta, Gregory M. aut Maeder, Morgan L. aut Kennedy, Edward M. aut Kornepati, Anand V. R. aut Sousa, Alexander aut Collins, McKensie A. aut Jayaram, Hari aut Cullen, Bryan R. aut Bumcrot, David aut Enthalten in Genome biology London : BioMed Central, 2000 16(2015), 1 vom: 24. Nov. (DE-627)326173617 (DE-600)2040529-7 1474-760X nnns volume:16 year:2015 number:1 day:24 month:11 https://dx.doi.org/10.1186/s13059-015-0817-8 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 16 2015 1 24 11
allfieldsGer	10.1186/s13059-015-0817-8 doi (DE-627)SPR030024714 (SPR)s13059-015-0817-8-e DE-627 ger DE-627 rakwb eng Friedland, Ari E. verfasserin aut Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Friedland et al. 2015 Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. CRISPR (dpeaa)DE-He213 Cas9 (dpeaa)DE-He213 Genome engineering (dpeaa)DE-He213 Nickases (dpeaa)DE-He213 Adeno-associated virus (dpeaa)DE-He213 GUIDE-seq (dpeaa)DE-He213 Baral, Reshica aut Singhal, Pankhuri aut Loveluck, Katherine aut Shen, Shen aut Sanchez, Minerva aut Marco, Eugenio aut Gotta, Gregory M. aut Maeder, Morgan L. aut Kennedy, Edward M. aut Kornepati, Anand V. R. aut Sousa, Alexander aut Collins, McKensie A. aut Jayaram, Hari aut Cullen, Bryan R. aut Bumcrot, David aut Enthalten in Genome biology London : BioMed Central, 2000 16(2015), 1 vom: 24. Nov. (DE-627)326173617 (DE-600)2040529-7 1474-760X nnns volume:16 year:2015 number:1 day:24 month:11 https://dx.doi.org/10.1186/s13059-015-0817-8 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 16 2015 1 24 11
allfieldsSound	10.1186/s13059-015-0817-8 doi (DE-627)SPR030024714 (SPR)s13059-015-0817-8-e DE-627 ger DE-627 rakwb eng Friedland, Ari E. verfasserin aut Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © Friedland et al. 2015 Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. CRISPR (dpeaa)DE-He213 Cas9 (dpeaa)DE-He213 Genome engineering (dpeaa)DE-He213 Nickases (dpeaa)DE-He213 Adeno-associated virus (dpeaa)DE-He213 GUIDE-seq (dpeaa)DE-He213 Baral, Reshica aut Singhal, Pankhuri aut Loveluck, Katherine aut Shen, Shen aut Sanchez, Minerva aut Marco, Eugenio aut Gotta, Gregory M. aut Maeder, Morgan L. aut Kennedy, Edward M. aut Kornepati, Anand V. R. aut Sousa, Alexander aut Collins, McKensie A. aut Jayaram, Hari aut Cullen, Bryan R. aut Bumcrot, David aut Enthalten in Genome biology London : BioMed Central, 2000 16(2015), 1 vom: 24. Nov. (DE-627)326173617 (DE-600)2040529-7 1474-760X nnns volume:16 year:2015 number:1 day:24 month:11 https://dx.doi.org/10.1186/s13059-015-0817-8 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 16 2015 1 24 11
language	English
source	Enthalten in Genome biology 16(2015), 1 vom: 24. Nov. volume:16 year:2015 number:1 day:24 month:11
sourceStr	Enthalten in Genome biology 16(2015), 1 vom: 24. Nov. volume:16 year:2015 number:1 day:24 month:11
format_phy_str_mv	Article
institution	findex.gbv.de
topic_facet	CRISPR Cas9 Genome engineering Nickases Adeno-associated virus GUIDE-seq
isfreeaccess_bool	true
container_title	Genome biology
authorswithroles_txt_mv	Friedland, Ari E. @@aut@@ Baral, Reshica @@aut@@ Singhal, Pankhuri @@aut@@ Loveluck, Katherine @@aut@@ Shen, Shen @@aut@@ Sanchez, Minerva @@aut@@ Marco, Eugenio @@aut@@ Gotta, Gregory M. @@aut@@ Maeder, Morgan L. @@aut@@ Kennedy, Edward M. @@aut@@ Kornepati, Anand V. R. @@aut@@ Sousa, Alexander @@aut@@ Collins, McKensie A. @@aut@@ Jayaram, Hari @@aut@@ Cullen, Bryan R. @@aut@@ Bumcrot, David @@aut@@
publishDateDaySort_date	2015-11-24T00:00:00Z
hierarchy_top_id	326173617
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author	Friedland, Ari E.
spellingShingle	Friedland, Ari E. misc CRISPR misc Cas9 misc Genome engineering misc Nickases misc Adeno-associated virus misc GUIDE-seq Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications
authorStr	Friedland, Ari E.
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topic_title	Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications CRISPR (dpeaa)DE-He213 Cas9 (dpeaa)DE-He213 Genome engineering (dpeaa)DE-He213 Nickases (dpeaa)DE-He213 Adeno-associated virus (dpeaa)DE-He213 GUIDE-seq (dpeaa)DE-He213
topic	misc CRISPR misc Cas9 misc Genome engineering misc Nickases misc Adeno-associated virus misc GUIDE-seq
topic_unstemmed	misc CRISPR misc Cas9 misc Genome engineering misc Nickases misc Adeno-associated virus misc GUIDE-seq
topic_browse	misc CRISPR misc Cas9 misc Genome engineering misc Nickases misc Adeno-associated virus misc GUIDE-seq
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title	Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications
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title_full	Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications
author_sort	Friedland, Ari E.
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author_browse	Friedland, Ari E. Baral, Reshica Singhal, Pankhuri Loveluck, Katherine Shen, Shen Sanchez, Minerva Marco, Eugenio Gotta, Gregory M. Maeder, Morgan L. Kennedy, Edward M. Kornepati, Anand V. R. Sousa, Alexander Collins, McKensie A. Jayaram, Hari Cullen, Bryan R. Bumcrot, David
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author-letter	Friedland, Ari E.
doi_str_mv	10.1186/s13059-015-0817-8
title_sort	characterization of staphylococcus aureus cas9: a smaller cas9 for all-in-one adeno-associated virus delivery and paired nickase applications
title_auth	Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications
abstract	Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. © Friedland et al. 2015
abstractGer	Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. © Friedland et al. 2015
abstract_unstemmed	Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. Conclusion Our results reveal an S. aureus Cas9 that is effective for a variety of genome engineering purposes, including paired nickase approaches and all-in-one delivery of Cas9 and multiple gRNA expression cassettes with AAV vectors. © Friedland et al. 2015
collection_details	GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700
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title_short	Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications
url	https://dx.doi.org/10.1186/s13059-015-0817-8
remote_bool	true
author2	Baral, Reshica Singhal, Pankhuri Loveluck, Katherine Shen, Shen Sanchez, Minerva Marco, Eugenio Gotta, Gregory M. Maeder, Morgan L. Kennedy, Edward M. Kornepati, Anand V. R. Sousa, Alexander Collins, McKensie A. Jayaram, Hari Cullen, Bryan R. Bumcrot, David
author2Str	Baral, Reshica Singhal, Pankhuri Loveluck, Katherine Shen, Shen Sanchez, Minerva Marco, Eugenio Gotta, Gregory M. Maeder, Morgan L. Kennedy, Edward M. Kornepati, Anand V. R. Sousa, Alexander Collins, McKensie A. Jayaram, Hari Cullen, Bryan R. Bumcrot, David
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doi_str	10.1186/s13059-015-0817-8
up_date	2024-07-03T13:24:04.330Z
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fullrecord_marcxml	<?xml version="1.0" encoding="UTF-8"?><collection xmlns="http://www.loc.gov/MARC21/slim"><record><leader>01000caa a22002652 4500</leader><controlfield tag="001">SPR030024714</controlfield><controlfield tag="003">DE-627</controlfield><controlfield tag="005">20230519223115.0</controlfield><controlfield tag="007">cr uuu---uuuuu</controlfield><controlfield tag="008">201007s2015 xx \|\|\|\|\|o 00\| \|\|eng c</controlfield><datafield tag="024" ind1="7" ind2=" "><subfield code="a">10.1186/s13059-015-0817-8</subfield><subfield code="2">doi</subfield></datafield><datafield tag="035" ind1=" " ind2=" "><subfield code="a">(DE-627)SPR030024714</subfield></datafield><datafield tag="035" ind1=" " ind2=" "><subfield code="a">(SPR)s13059-015-0817-8-e</subfield></datafield><datafield tag="040" ind1=" " ind2=" "><subfield code="a">DE-627</subfield><subfield code="b">ger</subfield><subfield code="c">DE-627</subfield><subfield code="e">rakwb</subfield></datafield><datafield tag="041" ind1=" " ind2=" "><subfield code="a">eng</subfield></datafield><datafield tag="100" ind1="1" ind2=" "><subfield code="a">Friedland, Ari E.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="245" ind1="1" ind2="0"><subfield code="a">Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications</subfield></datafield><datafield tag="264" ind1=" " ind2="1"><subfield code="c">2015</subfield></datafield><datafield tag="336" ind1=" " ind2=" "><subfield code="a">Text</subfield><subfield code="b">txt</subfield><subfield code="2">rdacontent</subfield></datafield><datafield tag="337" ind1=" " ind2=" "><subfield code="a">Computermedien</subfield><subfield code="b">c</subfield><subfield code="2">rdamedia</subfield></datafield><datafield tag="338" ind1=" " ind2=" "><subfield code="a">Online-Ressource</subfield><subfield code="b">cr</subfield><subfield code="2">rdacarrier</subfield></datafield><datafield tag="500" ind1=" " ind2=" "><subfield code="a">© Friedland et al. 2015</subfield></datafield><datafield tag="520" ind1=" " ind2=" "><subfield code="a">Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against genomic targets with mutually permissive NGGRRT PAMs, the S. pyogenes Cas9 and S. aureus Cas9 yield indels at comparable rates. We additionally show D10A and N580A paired nickase activity with S. aureus Cas9, and we further package it with two gRNAs in a single functional adeno-associated virus (AAV) vector. Finally, we assess comparative S. pyogenes and S. aureus Cas9 specificity using GUIDE-seq. 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Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications

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