A CRISPR Cas9 high-throughput genome editing toolkit for kinetoplastids

Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation of pathogens, including kinetoplastids. CRISPR technology provides the opportunity to develop scalable methods for high-throughput productio...

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Published inRoyal Society open science Vol. 4; no. 5; p. 170095
Main Authors Beneke, Tom, Madden, Ross, Makin, Laura, Valli, Jessica, Sunter, Jack, Gluenz, Eva
Format Journal Article
LanguageEnglish
Published England The Royal Society Publishing 01.05.2017
The Royal Society
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Abstract Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation of pathogens, including kinetoplastids. CRISPR technology provides the opportunity to develop scalable methods for high-throughput production of mutant phenotypes. Here, we report development of a CRISPR-Cas9 toolkit that allows rapid tagging and gene knockout in diverse kinetoplastid species without requiring the user to perform any DNA cloning. We developed a new protocol for single-guide RNA (sgRNA) delivery using PCR-generated DNA templates which are transcribed in vivo by T7 RNA polymerase and an online resource (LeishGEdit.net) for automated primer design. We produced a set of plasmids that allows easy and scalable generation of DNA constructs for transfections in just a few hours. We show how these tools allow knock-in of fluorescent protein tags, modified biotin ligase BirA*, luciferase, HaloTag and small epitope tags, which can be fused to proteins at the N- or C-terminus, for functional studies of proteins and localization screening. These tools enabled generation of null mutants in a single round of transfection in promastigote form Leishmania major, Leishmania mexicana and bloodstream form Trypanosoma brucei; deleted genes were undetectable in non-clonal populations, enabling for the first time rapid and large-scale knockout screens.
AbstractList Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation of pathogens, including kinetoplastids. CRISPR technology provides the opportunity to develop scalable methods for high-throughput production of mutant phenotypes. Here, we report development of a CRISPR-Cas9 toolkit that allows rapid tagging and gene knockout in diverse kinetoplastid species without requiring the user to perform any DNA cloning. We developed a new protocol for single-guide RNA (sgRNA) delivery using PCR-generated DNA templates which are transcribed in vivo by T7 RNA polymerase and an online resource (LeishGEdit.net) for automated primer design. We produced a set of plasmids that allows easy and scalable generation of DNA constructs for transfections in just a few hours. We show how these tools allow knock-in of fluorescent protein tags, modified biotin ligase BirA*, luciferase, HaloTag and small epitope tags, which can be fused to proteins at the N- or C-terminus, for functional studies of proteins and localization screening. These tools enabled generation of null mutants in a single round of transfection in promastigote form Leishmania major , Leishmania mexicana and bloodstream form Trypanosoma brucei ; deleted genes were undetectable in non-clonal populations, enabling for the first time rapid and large-scale knockout screens.
Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation of pathogens, including kinetoplastids. CRISPR technology provides the opportunity to develop scalable methods for high-throughput production of mutant phenotypes. Here, we report development of a CRISPR-Cas9 toolkit that allows rapid tagging and gene knockout in diverse kinetoplastid species without requiring the user to perform any DNA cloning. We developed a new protocol for single-guide RNA (sgRNA) delivery using PCR-generated DNA templates which are transcribed in vivo by T7 RNA polymerase and an online resource (LeishGEdit.net) for automated primer design. We produced a set of plasmids that allows easy and scalable generation of DNA constructs for transfections in just a few hours. We show how these tools allow knock-in of fluorescent protein tags, modified biotin ligase BirA*, luciferase, HaloTag and small epitope tags, which can be fused to proteins at the N- or C-terminus, for functional studies of proteins and localization screening. These tools enabled generation of null mutants in a single round of transfection in promastigote form Leishmania major, Leishmania mexicana and bloodstream form Trypanosoma brucei; deleted genes were undetectable in non-clonal populations, enabling for the first time rapid and large-scale knockout screens.
Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation of pathogens, including kinetoplastids. CRISPR technology provides the opportunity to develop scalable methods for high-throughput production of mutant phenotypes. Here, we report development of a CRISPR-Cas9 toolkit that allows rapid tagging and gene knockout in diverse kinetoplastid species without requiring the user to perform any DNA cloning. We developed a new protocol for single-guide RNA (sgRNA) delivery using PCR-generated DNA templates which are transcribed by T7 RNA polymerase and an online resource (LeishGEdit.net) for automated primer design. We produced a set of plasmids that allows easy and scalable generation of DNA constructs for transfections in just a few hours. We show how these tools allow knock-in of fluorescent protein tags, modified biotin ligase BirA*, luciferase, HaloTag and small epitope tags, which can be fused to proteins at the N- or C-terminus, for functional studies of proteins and localization screening. These tools enabled generation of null mutants in a single round of transfection in promastigote form , and bloodstream form ; deleted genes were undetectable in non-clonal populations, enabling for the first time rapid and large-scale knockout screens.
Author Makin, Laura
Madden, Ross
Beneke, Tom
Gluenz, Eva
Valli, Jessica
Sunter, Jack
AuthorAffiliation Sir William Dunn School of Pathology , University of Oxford , Oxford , UK
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  surname: Gluenz
  fullname: Gluenz, Eva
  email: eva.gluenz@path.ox.ac.uk
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Cites_doi 10.1128/mBio.01012-15
10.1111/j.1432-1033.1993.tb18283.x
10.1371/journal.ppat.1003566
10.1242/jcs.02969
10.1016/j.cell.2014.05.010
10.1038/nature14651
10.1074/jbc.M116.749655
10.1083/jcb.139.1.103
10.1126/science.1258096
10.1038/nature12864
10.1016/S0166-6851(00)00383-2
10.1128/mSphere.00340-16
10.1111/cmi.12456
10.2307/3282883
10.1128/mBio.01414-14
10.1038/nature10771
10.1021/cb3002478
10.1111/jeu.12338
10.1073/pnas.160257897
10.1098/rsob.110037
10.1073/pnas.97.1.406
10.1038/nbt.2925
10.1126/science.1231143
10.1016/0166-6851(96)02598-4
10.1083/jcb.201112098
10.1242/jcs.03078
10.1101/gr.115089.110
10.1038/nmeth.1209
10.1016/j.molbiopara.2014.11.002
10.1371/journal.ppat.1005186
10.1038/nbt1037
10.1371/journal.pone.0134432
10.1101/gr.122945.111
10.1083/jcb.98.1.229
10.1073/pnas.0901698106
10.1016/j.molbiopara.2005.09.002
10.1093/emboj/19.9.1953
10.1016/j.ymeth.2014.02.019
10.1371/journal.ppat.1001161
10.1128/mBio.00861-15
10.1155/2012/165126
10.1083/jcb.132.3.359
10.1021/cb800025k
10.1038/ncomms9964
10.1016/j.molbiopara.2010.09.002
10.1038/nrg980
10.1016/j.cell.2016.08.019
10.1038/nmeth.3063
10.1098/rsob.140197
10.1016/S0166-6851(97)00048-0
10.1038/nmeth.2413
10.1128/mBio.02097-14
10.1128/EC.5.4.696-711.2006
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Keywords CRISPR
T7 RNA polymerase
genome editing
Trypanosoma
Leishmania
Language English
License open-access: Published by the Royal Society under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/4.0/, which permits unrestricted use, provided the original author and source are credited.
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These authors contributed equally to this study.
Electronic supplementary material is available online at https://dx.doi.org/10.6084/m9.figshare.c.3744365.
Present address: School of Life Sciences, University of Dundee, Dundee, UK.
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References 26452044 - PLoS Pathog. 2015 Oct 09;11(10):e1005186
12509749 - Nat Rev Genet. 2003 Jan;4(1):11-9
9233680 - Mol Biochem Parasitol. 1997 Jul;87(1):113-5
19383793 - Proc Natl Acad Sci U S A. 2009 May 5;106(18):7583-8
18533659 - ACS Chem Biol. 2008 Jun 20;3(6):373-82
18454154 - Nat Methods. 2008 Jun;5(6):545-51
27793988 - J Biol Chem. 2016 Dec 2;291(49):25505-25515
25550322 - MBio. 2014 Dec 30;6(1):e02097-14
22278056 - Nature. 2012 Jan 25;482(7384):232-6
10790362 - EMBO J. 2000 May 2;19(9):1953-62
22038252 - Genome Res. 2011 Dec;21(12):2129-42
10908670 - Proc Natl Acad Sci U S A. 2000 Aug 1;97(16):9258-63
23524392 - Nat Methods. 2013 May;10(5):407-9
27315329 - J Eukaryot Microbiol. 2016 Sep;63(5):679-90
22894855 - ACS Chem Biol. 2012 Nov 16;7(11):1848-57
27863903 - Trends Parasitol. 2017 Feb;33(2):80-82
26199333 - MBio. 2015 Jul 21;6(4):e01012
27594426 - Cell. 2016 Sep 8;166(6):1423-1435.e12
27584862 - J Vis Exp. 2016 Aug 12;(114):null
24336212 - Nature. 2014 Jan 30;505(7485):681-5
22412018 - J Cell Biol. 2012 Mar 19;196(6):801-10
23287718 - Science. 2013 Feb 15;339(6121):819-23
25461484 - Mol Biochem Parasitol. 2014 Nov;198(1):11-3
24906146 - Cell. 2014 Jun 5;157(6):1262-78
15558047 - Nat Biotechnol. 2004 Dec;22(12):1567-72
28124028 - mSphere. 2017 Jan 18;2(1):null
16269191 - Mol Biochem Parasitol. 2006 Jan;145(1):117-20
16882690 - J Cell Sci. 2006 Aug 15;119(Pt 16):3443-55
20851719 - Mol Biochem Parasitol. 2011 Jan;175(1):91-4
43092 - Acta Trop. 1979 Sep;36(3):289-92
26199327 - MBio. 2015 Jul 21;6(4):e00861
23990786 - PLoS Pathog. 2013;9(8):e1003566
24987097 - MBio. 2014 Jul 01;5(4):e01414-14
6707088 - J Cell Biol. 1984 Jan;98(1):229-36
26667778 - Nat Commun. 2015 Dec 15;6:8964
27226299 - Proc Natl Acad Sci U S A. 2016 Jun 28;113(26):7225-30
25430774 - Science. 2014 Nov 28;346(6213):1258096
9314532 - J Cell Biol. 1997 Oct 6;139(1):103-14
25567099 - Open Biol. 2015 Jan;5(1):140197
16607017 - Eukaryot Cell. 2006 Apr;5(4):696-711
24880488 - Nat Biotechnol. 2014 Aug;32(8):819-21
25108687 - Nat Methods. 2014 Sep;11(9):915-8
2614608 - J Parasitol. 1989 Dec;75(6):985-9
22523640 - J Parasitol Res. 2012;2012:165126
24576617 - Methods. 2014 Sep;69(2):128-36
11254965 - Mol Biochem Parasitol. 2001 Mar;113(1):171-3
21060810 - PLoS Pathog. 2010 Oct 28;6(10):e1001161
7693464 - Eur J Biochem. 1993 Oct 15;217(2):603-15
16720646 - J Cell Sci. 2006 Jun 15;119(Pt 12):2405-13
8381972 - Proc Natl Acad Sci U S A. 1993 Feb 15;90(4):1599-603
21363968 - Genome Res. 2011 Jun;21(6):915-24
10618431 - Proc Natl Acad Sci U S A. 2000 Jan 4;97(1):406-11
26176919 - Nature. 2015 Jul 23;523(7561):477-80
22645659 - Open Biol. 2012 Feb;2(2):110037
26266938 - PLoS One. 2015 Aug 12;10(8):e0134432
25939677 - Cell Microbiol. 2015 Oct;17(10):1405-12
8813669 - Mol Biochem Parasitol. 1996 May;77(2):235-9
8636214 - J Cell Biol. 1996 Feb;132(3):359-70
e_1_3_6_30_2
e_1_3_6_51_2
e_1_3_6_32_2
e_1_3_6_53_2
e_1_3_6_19_2
Peng D (e_1_3_6_22_2) 2015; 6
e_1_3_6_13_2
e_1_3_6_38_2
e_1_3_6_59_2
e_1_3_6_11_2
e_1_3_6_17_2
e_1_3_6_34_2
e_1_3_6_55_2
e_1_3_6_36_2
e_1_3_6_57_2
e_1_3_6_40_2
e_1_3_6_21_2
e_1_3_6_42_2
e_1_3_6_4_2
e_1_3_6_2_2
e_1_3_6_8_2
e_1_3_6_6_2
Dyer P (e_1_3_6_14_2) 2016; 114
e_1_3_6_27_2
e_1_3_6_48_2
e_1_3_6_29_2
e_1_3_6_23_2
e_1_3_6_44_2
e_1_3_6_25_2
e_1_3_6_46_2
e_1_3_6_52_2
e_1_3_6_31_2
e_1_3_6_54_2
e_1_3_6_10_2
e_1_3_6_50_2
Brun R (e_1_3_6_56_2) 1979; 36
Dean S (e_1_3_6_15_2) 2016; 1862
e_1_3_6_37_2
e_1_3_6_12_2
e_1_3_6_39_2
e_1_3_6_18_2
e_1_3_6_33_2
e_1_3_6_16_2
e_1_3_6_35_2
e_1_3_6_58_2
e_1_3_6_41_2
e_1_3_6_20_2
e_1_3_6_43_2
e_1_3_6_5_2
e_1_3_6_3_2
e_1_3_6_9_2
e_1_3_6_7_2
e_1_3_6_26_2
e_1_3_6_49_2
e_1_3_6_28_2
e_1_3_6_45_2
e_1_3_6_24_2
e_1_3_6_47_2
References_xml – ident: e_1_3_6_23_2
  doi: 10.1128/mBio.01012-15
– ident: e_1_3_6_49_2
  doi: 10.1111/j.1432-1033.1993.tb18283.x
– volume: 1862
  start-page: 210
  year: 2016
  ident: e_1_3_6_15_2
  article-title: TrypTag.org: a trypanosome genome-wide protein localisation resource
  publication-title: Trends Parasitol.
  contributor:
    fullname: Dean S
– ident: e_1_3_6_53_2
  doi: 10.1371/journal.ppat.1003566
– ident: e_1_3_6_43_2
  doi: 10.1242/jcs.02969
– ident: e_1_3_6_54_2
  doi: 10.1016/j.cell.2014.05.010
– ident: e_1_3_6_20_2
  doi: 10.1038/nature14651
– ident: e_1_3_6_26_2
  doi: 10.1074/jbc.M116.749655
– ident: e_1_3_6_51_2
  doi: 10.1083/jcb.139.1.103
– ident: e_1_3_6_16_2
  doi: 10.1126/science.1258096
– ident: e_1_3_6_4_2
  doi: 10.1038/nature12864
– ident: e_1_3_6_11_2
  doi: 10.1016/S0166-6851(00)00383-2
– ident: e_1_3_6_27_2
  doi: 10.1128/mSphere.00340-16
– ident: e_1_3_6_25_2
  doi: 10.1111/cmi.12456
– ident: e_1_3_6_55_2
  doi: 10.2307/3282883
– volume: 114
  start-page: e54342
  year: 2016
  ident: e_1_3_6_14_2
  article-title: High-throughput gene tagging in Trypanosoma brucei
  publication-title: J. Vis. Exp.
  contributor:
    fullname: Dyer P
– ident: e_1_3_6_19_2
  doi: 10.1128/mBio.01414-14
– ident: e_1_3_6_6_2
  doi: 10.1038/nature10771
– ident: e_1_3_6_37_2
  doi: 10.1021/cb3002478
– ident: e_1_3_6_8_2
– ident: e_1_3_6_5_2
– ident: e_1_3_6_17_2
  doi: 10.1111/jeu.12338
– ident: e_1_3_6_47_2
  doi: 10.1073/pnas.160257897
– ident: e_1_3_6_50_2
  doi: 10.1098/rsob.110037
– ident: e_1_3_6_45_2
  doi: 10.1073/pnas.97.1.406
– volume: 36
  start-page: 289
  year: 1979
  ident: e_1_3_6_56_2
  article-title: Cultivation and in vitro cloning or procyclic culture forms of Trypanosoma brucei in a semi-defined medium. Short communication
  publication-title: Acta Trop.
  contributor:
    fullname: Brun R
– ident: e_1_3_6_18_2
  doi: 10.1038/nbt.2925
– ident: e_1_3_6_28_2
  doi: 10.1126/science.1231143
– ident: e_1_3_6_35_2
  doi: 10.1016/0166-6851(96)02598-4
– ident: e_1_3_6_38_2
  doi: 10.1083/jcb.201112098
– ident: e_1_3_6_42_2
  doi: 10.1242/jcs.03078
– ident: e_1_3_6_3_2
  doi: 10.1101/gr.115089.110
– ident: e_1_3_6_52_2
  doi: 10.1038/nmeth.1209
– ident: e_1_3_6_30_2
  doi: 10.1016/j.molbiopara.2014.11.002
– ident: e_1_3_6_39_2
  doi: 10.1371/journal.ppat.1005186
– ident: e_1_3_6_34_2
  doi: 10.1038/nbt1037
– ident: e_1_3_6_59_2
  doi: 10.1371/journal.pone.0134432
– ident: e_1_3_6_9_2
  doi: 10.1101/gr.122945.111
– ident: e_1_3_6_40_2
  doi: 10.1083/jcb.98.1.229
– ident: e_1_3_6_57_2
  doi: 10.1073/pnas.0901698106
– ident: e_1_3_6_12_2
  doi: 10.1016/j.molbiopara.2005.09.002
– ident: e_1_3_6_48_2
  doi: 10.1093/emboj/19.9.1953
– ident: e_1_3_6_29_2
  doi: 10.1016/j.ymeth.2014.02.019
– ident: e_1_3_6_7_2
  doi: 10.1371/journal.ppat.1001161
– ident: e_1_3_6_24_2
  doi: 10.1128/mBio.00861-15
– ident: e_1_3_6_46_2
  doi: 10.1155/2012/165126
– ident: e_1_3_6_41_2
  doi: 10.1083/jcb.132.3.359
– ident: e_1_3_6_36_2
  doi: 10.1021/cb800025k
– ident: e_1_3_6_58_2
  doi: 10.1038/ncomms9964
– ident: e_1_3_6_31_2
  doi: 10.1016/j.molbiopara.2010.09.002
– ident: e_1_3_6_2_2
  doi: 10.1038/nrg980
– ident: e_1_3_6_21_2
  doi: 10.1016/j.cell.2016.08.019
– ident: e_1_3_6_32_2
  doi: 10.1038/nmeth.3063
– ident: e_1_3_6_13_2
  doi: 10.1098/rsob.140197
– ident: e_1_3_6_10_2
  doi: 10.1016/S0166-6851(97)00048-0
– ident: e_1_3_6_33_2
  doi: 10.1038/nmeth.2413
– volume: 6
  year: 2015
  ident: e_1_3_6_22_2
  article-title: CRISPR-Cas9-mediated single-gene and gene family disruption in Trypanosoma cruzi
  publication-title: mBio
  doi: 10.1128/mBio.02097-14
  contributor:
    fullname: Peng D
– ident: e_1_3_6_44_2
  doi: 10.1128/EC.5.4.696-711.2006
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Snippet Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation...
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SubjectTerms Cellular And Molecular Biology
Crispr
Genome Editing
leishmania
T7 Rna Polymerase
trypanosoma
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Title A CRISPR Cas9 high-throughput genome editing toolkit for kinetoplastids
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