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CRISPR header icon CRISPR/Cas Plasmids: Empty gRNA Expression Vectors


With the CRISPR/Cas system, you have to express both a Cas9 protein and a target-specific gRNA in the same cell at the same time. Single plasmids containing both the gRNA and Cas9 act as an all-in-one vector, but their function (cut, nick, activate, interfere, etc.) is limited to that of the Cas9 present on the plasmid. gRNA plasmids that do not co-express Cas9 require a separate plasmid that does so; however, these independent gRNA plasmids can be paired with a wide variety of Cas9 plasmids and therefore are not limited to a single Cas9 function. Alternatively, independent gRNA plasmids can be used in cells or organisms in which Cas9 has been integrated into the genome.

Single gRNA Empty Vectors

Many gRNA empty vectors have been deposited at Addgene. To find the gRNA vector for your system, you can search or sort this table based on:

  • Your particular expression system, eg. Mammalian, Lentiviral, AAV, Bacteria, Drosophila, Yeast, Plant, Xenopus, Worm, or Other.
  • Whether the plasmid contains Cas9 and if so, which function of Cas9. We have plasmids with Cas9 that can cut, activate, interfere, or nick.
  • Selection, such as Puromycin or EGFP
  • Cloning enzyme used for insertion of your gRNA sequence, such as BsbI or BsaI
  • Cas9 (or other CRISPR) system that the vector was designed to be used with, such as S. pyogenes, S. aureus, N. meningitidis, etc. gRNA scaffolds are specific for the species of Cas9 it is to be used with. crRNAs used with Cpf1 or other CRISPR systems are also listed here and denoted as such.
Plasmid Expression System Cloning Enzyme Co-expressed Cas9 Cas9 System Selection PI
pCRISPR42875 Bacteria BsaI none S. pyogenes Chloramphenicol Marraffini
pCas942876 Bacteria BsaI yes, cut S. pyogenes Chloramphenicol Marraffini
pgRNA-bacteria44251 Bacteria SpeI + HindIII none S. pyogenes Qi
pDD162 (Peft-3::Cas9 + Empty sgRNA)47549 C. elegans yes, cut S. pyogenes Goldstein
DR27442250 C. elegans BsaI none S. pyogenes Joung
pCFD3-dU6:3gRNA49410 Drosophila BbsI none S. pyogenes Virmilion Bullock and Port
pAc-sgRNA-Cas949330 Drosophila BspQI yes, cut S. pyogenes Puro Liu
pCFD4-U6:1_U6:3tandemgRNAs49411 Drosophila BbsI none S. pyogenes Virmilion Bullock and Port
pU6-BbsI-chiRNA45946 Drosophila BbsI none S. pyogenes O'Connor-Giles, Harrison, Wildonger
gRNA_Cloning Vector41824 Mammalian AfIII none S. pyogenes Bleocin Church
MLM363643860 Mammalian BsmBI none S. pyogenes Joung
pSPgRNA47108 Mammalian BbsI none S. pyogenes Gersbach
pUC57-sgRNA expression vector51132 Mammalian BsaI none S. pyogenes Huang
pAC154-dual-dCas9VP160-sgExpression48240 Mammalian BbsI yes, activate S. pyogenes Jaenisch
pSQT131353370 Mammalian BsmBI none S. pyogenes Joung
PX461 (3rd Gen)48140 Mammalian BbsI yes, nick S. pyogenes EGFP Zhang
PX460 (3rd Gen)48873 Mammalian BbsI yes, nick S. pyogenes Zhang
PX458 (3rd Gen)48138 Mammalian BbsI yes, cut S. pyogenes EGFP Zhang
PX335 (2nd Gen)42335 Mammalian BbsI yes, nick S. pyogenes Zhang
PX334 (1st Gen)42333 Mammalian BbsI yes, nick S. pyogenes Puro Zhang
PX330 (2nd Gen)42230 Mammalian BbsI yes, cut S. pyogenes Zhang
PX260 (1st Gen)42229 Mammalian BbsI yes, cut S. pyogenes Puro Zhang
sgRNA(MS2) cloning backbone61424 Mammalian BbsI none S. pyogenes Zhang
pSpCas9n(BB)-2A-Puro (PX462) V2.062987 Mammalian BbsI yes, nick S. pyogenes Puro Zhang
pSpCas9(BB)-2A-Puro (PX459) V2.062988 Mammalian BbsI yes, cut S. pyogenes Puro Zhang
phH1-gRNA53186 Mammalian BbsI none S. pyogenes Gersbach
pmU6-gRNA53187 Mammalian BbsI none S. pyogenes Gersbach
phU6-gRNA53188 Mammalian BbsI none S. pyogenes Gersbach
ph7SK-gRNA53189 Mammalian BbsI none S. pyogenes Gersbach
PX55260958 Mammalian/AAV SapI none S. pyogenes Zhang
pgRNA-humanized44248 Mammalian/Lentiviral BstXI + XhoI none S. pyogenes Puro, mCherrry Qi
pLX-sgRNA50662 Mammalian/Lentiviral See paper none S. pyogenes Blast Sabatini and Lander
lentiCRISPR v252961 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes Puro Zhang
pKLV-U6gRNA(BbsI)-PGKpuro2ABFP50946 Mammalian/Lentiviral BbsI none S. pyogenes Puro, TagBFP Yusa
lentiGuide-Puro52963 Mammalian/Lentiviral BsmBI none S. pyogenes Puro Zhang
lenti sgRNA(MS2)_zeo backbone61427 Mammalian/Lentiviral BsmBI none S. pyogenes Zhang
pL-CRISPR.EFS.GFP57818 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes EGFP Ebert
pLKO.1-puro U6 sgRNA BfuAI stuffer50920 Mammalian/Lentiviral BfuAI none S. pyogenes Puro Wolfe
pL-CRISPR.EFS.tRFP57819 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes tagRFP Ebert
pFGC-pcoCas952256 Plant AscI, PacI, SbfI yes, cut S. pyogenes Sheen
pT7-gRNA46759 Zebrafish BsmBI none S. pyogenes Chen and Wente
DR27442250 Zebrafish BsaI none S. pyogenes Joung
pCRISPathBrick65006 Bacteria BsaI yes, interfere S. pyogenes Koffas
BPK764 65767 Bacteria BsaI yes, cut S. pyogenes Joung
MSP71265768 Bacteria BsaI yes, interfere S. pyogenes Joung
sgRNA with U6 promoter48962 Worm HindIII none S. pyogenes de Bono
sgRNA with rpr-1 promoter48961 Worm EcoRI none S. pyogenes de Bono
pMB6047941 Worm BsaI none S. pyogenes Boxem
pIK198 65629 Worm Gibson none S. pyogenes Katic
pHKMC1: Empty sgRNA for Cloning67720 Worm NotI/BamHI none S. pyogenes Colaiacovo
pRB1752527 Fly blunt end cloning none S. pyogenes Foerstemann
U6-BbsI-crRNA51026 Fly BbsI none S. pyogenes O'Connor-Giles, Harrison, Wildonger
pDCC559984 Fly BbsI yes, nick S. pyogenes Duchek
pAC5-dual-dCas9VP48-sgTetO48237 Mammalian BbsI yes, activate S. pyogenes Jaenisch
pU6_gRNA_handle_U6t49016 Mammalian SacI none S. pyogenes EBFP Lu
pH1v160244 Mammalian Gibson none S. pyogenes Zack
pHL-H1-ccdB-mEF1a-RiH60601 Mammalian BamHI/EcoRI none S. pyogenes Hygro Hotta
pUC-H1-gRNA61089 Mammalian BsaI none S. pyogenes Huang
PX85362885 Mammalian BbsI yes, cut, N-terminal S. pyogenes Zhang
PX85462886 Mammalian BbsI yes, cut, C-terminal S. pyogenes Zhang
pGuide 64711 Mammalian BbsI none S. pyogenes Musunuru
pU6-(BbsI)_CBh-Cas9-T2A-mCherry-H1-(BamHI)64217 Mammalian BbsI  yes, cut S. pyogenes mCherry Kuhn
pU6-(BbsI)_CBh-Cas9-T2A-mcherry-P2A-Ad4E1B64221 Mammalian see plasmid page yes, cut S. pyogenes mCherry Kuhn
pU6-(BbsI)_CBh-Cas9-T2A-BFP64323 Mammalian BbsI  yes, cut S. pyogenes BFP Kuhn
pU6-(BbsI)_CBh-Cas9-T2A-mCherry64324 Mammalian BbsI  yes, cut S. pyogenes mCherry Kuhn
pEX-A-U6-gRNA65626 Mammalian none S. pyogenes Bultmann
BPK152065777 Mammalian BsmBI none S. pyogenes Joung
AAV:ITR-U6-sgRNA (Backbone) PCB-FlPO-WPRE-syntetisk pA-UTR68347 Mammalian/AAV none S. pyogenes Elverlov-Jakobsen
pU6T7G71463 Mammalian/in vitro transcription BsaI none S. pyogenes Mark
pU6T771462 Mammalian/in vitro transcription BsaI none S. pyogenes Mark
pLenti-sgRNA-Lib53121 Mammalian/Lentiviral BsmBI none S. pyogenes ccdB Wei
pLKO5.sgRNA.EFS.PAC57825 Mammalian/Lentiviral BsmBI none S. pyogenes Puro Ebert
pL-CRISPR.EFS.PAC57828 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes Puro Ebert
pL-CRISPR.SFFV.PAC57829 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes Puro Ebert
pLenti SpBsmBI sgRNA Hygro62205 Mammalian/Lentiviral BsmBI none S. pyogenes Hygro Maehr
pLenti SpBsmBI sgRNA Puro62207 Mammalian/Lentiviral BsmBI none S. pyogenes Puro Maehr
pKLV-U6gRNA-EF(BbsI)-PGKpuro2ABFP62348 Mammalian/Lentiviral BbsI none S. pyogenes Puro Ochiai
pSB70064046 Mammalian/Lentiviral none S. pyogenes Cerulean Church
LRG (Lenti_sgRNA_EFS_GFP)65656 Mammalian/Lentiviral LIC none S. pyogenes GFP Vakoc
U6>sgRNA(F+E)59986 Other/Ciona intestinalis BsaI none S. pyogenes Christiaen
p(U6a-BsaI-gRNA)65955 Other/Tribolium castaneum BsaI none S. pyogenes Averof
p(U6b-BsaI-gRNA)65956 Other/Tribolium castaneum BsaI none S. pyogenes Averof
pBUN6A1150579 Plant BsaI yes, activate S. pyogenes Bar Chen
pBUN6I1150580 Plant BsaI yes, interfere S. pyogenes Bar Chen
pBUN41150581 Plant BsaI yes, cut S. pyogenes Bar Chen
pBUN50150582 Plant BsaI yes, nick S. pyogenes Bar Chen
pHSN6A0150586 Plant BsaI yes, activate S. pyogenes Hyg Chen
pHSN6I0150587 Plant BsaI yes, interfere S. pyogenes Hyg Chen
pHSN50150589 Plant BsaI yes, nick S. pyogenes Hyg Chen
pCBC-DT3T450592 Plant BsaI none S. pyogenes Chen
pCBC-MT1T250593 Plant BsaI none S. pyogenes Chen
pCBC-MT2T350594 Plant BsaI none S. pyogenes Chen
pCBC-MT3T450595 Plant BsaI none S. pyogenes Chen
pBUN42162204 Plant BsaI yes, cut S. pyogenes Bar Chen
pU3-gRNA53063 Plant AarI none S. pyogenes Gao
pZmU3-gRNA53061 Plant none S. pyogenes Gao
pU6-gRNA53062 Plant BbsI none S. pyogenes Gao
pBlu/gRNA59188 Plant BbsI none S. pyogenes Stupar
pRGE3263159 Plant BsaI yes, cut S. pyogenes Hygro Yang
pBUE41162200 Plant yes, cut S. pyogenes Basta Chen
pKSE40162202 Plant yes, cut S. pyogenes Kan Chen
pHUE41162203 Plant yes, cut S. pyogenes Hyg Chen
pHAtC78098 Plant AarI yes, cut S. pyogenes Hygro Kim
pBAtC78097 Plant AarI yes, cut S. pyogenes Basta Kim
pMZ28352224 Yeast CspCI none S. pyogenes URA4 Zaratiegui
pJZC58862315 Yeast none S. pyogenes URA3 Lim, Qi
pRPR1(1xTetO)_gRNA_handle_RPR1t62966 Yeast HindIII none S. pyogenes Puro Lu
pML10467638 Yeast BclI-SwaI  yes, cut S. pyogenes URA3 Wyrick
pML10767639 Yeast BclI-SwaI  yes, cut S. pyogenes LEU2 Wyrick
pT04067640 Yeast BclI-SwaI  none S. pyogenes URA3 Wyrick
pCRISPomyces-261737 Bacteria BbsI yes, cut S. pyogenes Apramycin Zhao
pRB101759936 Worm BsaI none S. pyogenes Fire
pGL3-U6-sgRNA-PGK-Puro51133 Mammalian none S. pyogenes Puro Huang
pAC152-dual-dCas9VP64-sgExpression48238 Mammalian BbsI yes, activate S. pyogenes Jaenisch
pU6-(BbsI)_CBh-Cas9-T2A-mcherry-P2A-Ad4E4orf664222 Mammalian see plasmid page yes, cut S. pyogenes mCherry Kuhn
AAV:ITR-U6-sgRNA(backbone)-pCBh-Cre-WPRE-hGHpA-ITR60229 Mammalian/AAV SapI none S. pyogenes Zhang
AAV:ITR-U6-sgRNA(backbone)-hSyn-Cre-2A-EGFP-KASH-WPRE-shortPA-ITR60231 Mammalian/AAV SapI none S. pyogenes Zhang
AAV:ITR-U6-sgRNA(backbone)-pEFS-Rluc-2A-Cre-WPRE-hGHpA-ITR60226 Mammalian/AAV SapI none S. pyogenes Zhang
pLKO5.sgRNA.EFS.GFP57822 Mammalian/Lentiviral BsmBI none S. pyogenes EGFP Ebert
pSECC60820 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes Jacks
pU6-sgRNA EF1Alpha-puro-T2A-BFP60955 Mammalian/Lentiviral none S. pyogenes Puro Weissman
pLSC-562889 Mammalian/Lentiviral BsmBI yes, cut (split) S. pyogenes Zhang
pICSL01009::AtU6p46968 Plant BsaI none S. pyogenes Kamoun
pUC119-gRNA52255 Plant PCR template none S. pyogenes Sheen
pRGEB3151295 Plant BsaI none S. pyogenes Yang
pRPR1_gRNA_handle_RPR1t 49014 Yeast HindIII none S. pyogenes LEU2 Lu
pDestTol2CG2-U6:gRNA63156 Zebrafish BseRI none S. pyogenes Zon
pDestTol2pA2-U6:gRNA63157 Zebrafish BseRI none S. pyogenes Zon
pCRISPomyces-161736 Bacteria BbsI yes, cut S. pyogenes Apramycin Zhao
pMB7047943 Worm BsaI none S. pyogenes Boxem
PU6::klp-12_sgRNA46170 Worm PCR (see paper) none S. pyogenes Calarco
SP6-sgRNA-scaffold47912 Worm AflII none S. pyogenes Sternberg
pJW131161254 Worm none, fusion PCR none S. pyogenes Ward
pCFD1-dU6:1gRNA49408 Fly BbsI none S. pyogenes Virmilion Bullock and Port
pCFD2-dU6:2gRNA49409 Fly BbsI none S. pyogenes Virmilion Bullock and Port
pDCC659985 Fly BbsI yes, cut S. pyogenes Duchek
pAC155-pCR8-sgExpression49045 Mammalian BbsI none S. pyogenes Jaenisch
pAC153-dual-dCas9VP96-sgExpression48239 Mammalian BbsI yes, activate S. pyogenes Jaenisch
pAC2-dual-dCas9VP48-sgExpression48236 Mammalian BbsI yes, activate S. pyogenes Jaenisch
piCRg Entry58904 Mammalian BbsI yes, cut S. pyogenes Huangfu
pX33364073 Mammalian BbsI and BsaI yes, cut S. pyogenes Ventura
PX85162883 Mammalian BbsI yes, cut, N-terminal S. pyogenes Zhang
PX85262884 Mammalian BbsI yes, cut, C-terminal S. pyogenes Zhang
PX85562887 Mammalian BbsI yes, activate, N-terminal S. pyogenes Zhang
PX85662888 Mammalian BbsI yes, activate, C-terminal S. pyogenes Zhang
pU6-(BbsI)_CBh-Cas9-T2A-BFP-P2A-Ad4E1B64218 Mammalian see plasmid page yes, cut S. pyogenes BFP Kuhn
pLKO.1-puro U6 sgRNA BfuAI large stuffer52628 Mammalian/Lentiviral BfuAI none S. pyogenes Puro Wolfe
pLKO5.sgRNA.EFS.tRFP65757824 Mammalian/Lentiviral BsmBI none S. pyogenes tagRFP657 Ebert
pL-CRISPR.SFFV.GFP57827 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes EGFP Ebert
pLKO5.sgRNA.EFS.tRFP57823 Mammalian/Lentiviral BsmBI none S. pyogenes tagRFP Ebert
pL-CRISPR.SFFV.tRFP57826 Mammalian/Lentiviral BsmBI yes, cut S. pyogenes tagRFP Ebert
pSAG1::CAS9-U6::sgUPRT54467 Other/Toxoplasma gondii none, Q5 mutagenesis yes, cut S. pyogenes Sibley
pU6-Universal52694 Other/Toxoplasma gondii BsaI yes, cut S. pyogenes Lourido
pRGE3150929 Plant BsaI none S. pyogenes Yang
pUC gRNA Shuttle47024 Plant inFusion none S. pyogenes Parrott
pCBC-DT1T150590 Plant BsaI none S. pyogenes Chen
pCBC-DT2T350591 Plant BsaI none S. pyogenes Chen
pHSN40150588 Plant BsaI yes, cut S. pyogenes Hyg Chen
pRGEB3263142 Plant BsaI yes, cut S. pyogenes Hygro Yang
pHSE40162201 Plant yes, cut S. pyogenes Hyg Chen
pUC57-Simple-gRNA backbone51306 Xenopus BsaI none S. pyogenes Chen
pMZ37459896 Yeast CspCI yes, cut S. pyogenes URA4 Zaratiegui
pdCas946569 Bacteria BsaI yes, interfere S. pyogenes Marraffini
pWJ6646570 Bacteria BsaI yes, activate S. pyogenes Marraffini
pWJ6846571 Bacteria BsaI yes, activate S. pyogenes Marraffini
pSimpleII-U6-tracr-U6-BsmBI-NLS-NmCas9-HA-NLS(s)47868 Mammalian U6 yes, cut N. meningitidis Thomson
pSmart-Nm-sgRNA-BbsI49157 Mammalian U6 none N. meningitidis Thomson
pXCas9H840A60900 Mammalian U6 yes, nick S. pyogenes Conklin
pXFokI-dCas960901 Mammalian U6 yes, FokI S. pyogenes Conklin
pcrRNA.ind61284 Bacteria araB none S. pyogenes Beisel
pcrRNA.con61285 Bacteria J23119 none S. pyogenes Beisel
pZDonor-Nm-Cas9-gRNA-hU661366 Mammalian U6 none N. meningitidis Gersbach
pSPneogRNAH63556 Other/Leishmania none S. pyogenes Matlashewski
pSPneoHHgRNAH63557 Other/Leishmania none S. pyogenes Matlashewski
pLH-spsgRNA264114 Mammalian/Lentiviral U6 none S. pyogenes Pederson
pLH-nmsgRNA1.164115 Mammalian/Lentiviral U6 none N. meningitidis Pederson
pLH-stsgRNA1.164116 Mammalian/Lentiviral U6 none N. meningitidis Pederson
pLH-stsgRNA2.164117 Mammalian/Lentiviral U6 none N. meningitidis Pederson
pLH-stsgRNA3.164118 Mammalian/Lentiviral U6 none N. meningitidis Pederson
pU6-(BbsI)_CBh-Cas9-T2A-BFP-P2A-Ad4E4orf664220 Mammalian U6 yes, cut S. pyogenes Kuehn
pMCS-rybozyme-IRES-CAS964668 Mammalian yes, cut S. pyogenes W Fujii
MSP167365769 Bacteria T7 yes, cut N. meningitidis Joung
Cas9 sgRNA vector68463 Mammalian U6 none S. pyogenes Zhang
Cas9/pTREX-n68708 Other/Trypanosoma cruzi yes, cut S. pyogenes Neo Docampo
tdTomato/pTREX-b68709 Other/Trypanosoma cruzi none S. pyogenes Blast Docampo
pDonor_hU669312 Mammalian hU6 none S. pyogenes Ventura
pDonor_mU669350 Mammalian mU6 none S. pyogenes Ventura
pDonor_sU669351 Mammalian sU6 none S. pyogenes Ventura
pCRISPRyl70007 Other/Yarrowia lipolytica SCR1'-tRNA yes, cut S. pyogenes LEU2 Wheeldon
FgH1tUTG70183 Mammalian/Lentiviral H1-Tet none S. pyogenes Herold
lentiGuide-Crimson70683 Mammalian/Lentiviral hU6 none S. pyogenes  Crimson Bauer
BPK266070709 Mammalian U6 none S. aureus Joung
pLV hU6-sgRNA hUbC-dCas9-KRAB-T2a-Puro71236 Mammalian/Lentiviral hU6 yes, interfere S. pyogenes Puro Gersbach
pHEE40171286 Plant U6-26p yes, cut S. pyogenes Hygro Chen
pHEE401E71287 Plant U6-26p yes, cut S. pyogenes Hygro Chen
pLenti-sgRNA71409 Mammalian/Lentiviral hU6 none S. pyogenes Puro Sabatini
sgBbsI (p2Tol-U6-2xBbsI-sgRNA-HygR)71485 Mammalian none S. pyogenes Hygro Sherwood
pMpGE_En0171534 Other/Marchantia polymorpha MpU6 none S. pyogenes Hara-Nishimura
pMpGE_En0371535 Other/Marchantia polymorpha MpU6 none S. pyogenes Noji
pGRB71539 Bacteria none S. pyogenes Chen
pdCas9-DNMT3A-EGFP71666 Mammalian U6 yes, methylation S. pyogenes EGFP Zoldos
pdCas9-DNMT3A-PuroR71667 Mammalian U6 yes, methylation S. pyogenes Puro Zoldos
pX330-U6-Chimeric_BB-CBh-hSpCas9-hGem(1/110)71707 Mammalian hU6 yes, cut S. pyogenes Draetta
pCAS1yl73226 Other/Yarrowia lipolytica yes, cut S. pyogenes LEU2 Yang
pLCKO73311 Mammalian/Lentiviral hU6 none S. pyogenes Puro Moffat
lenti sgRNA(MS2)_puro backbone73795 Mammalian/Lentiviral U6 none S. pyogenes Puro Zhang
pRS416-dCas9-Mxi1 + TetR + pRPR1(TetO)-NotI-gRNA73796 Yeast pRPR1(TetO) yes, interfere S. pyogenes  URA3 Davis
lenti sgRNA(MS2)_puro optimized backbone73797 Mammalian/Lentiviral U6 none S. pyogenes  Puro Zhang
pAWp2873850 Mammalian U6 none S. pyogenes Lu
pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA274009 Zebrafish U6 yes, cut S. pyogenes EGFP Del Bene
pUAS:Cas9T2ACre;U6:sgRNA1;U6:sgRNA274010 Zebrafish U6 yes, cut S. pyogenes Cre Del Bene
pTE439674041 Mammalian hU6 yes, cut As Cpf1 Neo Welker
pTE439874042 Mammalian hU6 yes, cut Lb Cpf1 Neo Welker
AIO-GFP74119 Mammalian U6x2 yes, nick S. pyogenes EGFP Jackson
AIO-mCherry74120 Mammalian U6x2 yes, nick S. pyogenes mCherry Jackson
pMZ37674213 Yeast rrk1 yes, cut S. pyogenes  KanMX Zaratiegui
pMZ37774214 Yeast rrk1 yes, cut S. pyogenes  LEU2 Zaratiegui
pMZ37974215 Yeast rrk1 yes, cut S. pyogenes  NatMX Zaratiegui
All_in_one_CRISPR/Cas9_LacZ74293 Mammalian S. pyogenes  Neo, mCherry Postovit
pdCas9-DNMT3A-PuroR_v274407 Mammalian U6 yes, methylation S. pyogenes Puro Zoldos
pBLO1811_Cas9_noNLS_human74490 Mammalian U6 yes, cut S. pyogenes mCherry Savage
pBLO1808_arC9_2xNLS_human74491 Mammalian U6 yes, cut S. pyogenes mCherry Savage
pSG4K574492 Bacteria J23119 none S. pyogenes  Wang
pBLO1811_arC9_noNLS_human74494 Mammalian U6 yes, cut S. pyogenes mCherry Savage
AIO-Puro74630 Mammalian U6x2 yes, nick S. pyogenes Puro Jackson
pBluescriptSKII+ U6-sgRNA(F+E) empty74707 Mammalian hU6  none S. pyogenes Yeo
pKS diaCas9_sgRNA74923 Other/Phaeodactylum tricornutum U6 yes, cut S. pyogenes Winge
pJJR5075026 Worm R07E5.16 (U6) none S. pyogenes Boxem
lentiSAMv275112 Mammalian/Lentiviral U6 yes, activate S. pyogenes Blast Zhang
PXL75349 Mammalian U6 yes, activate S. pyogenes Luikart
pRubiG-T2A-Cas975348 Mammalian/Retroviral from pXL yes, activate S. pyogenes EGFP Luikart
pRubiC-T2A-Cas9 75347 Mammalian/Retroviral from pXL yes, activate S. pyogenes mCherry Luikart
pDECKO_mCherry78534 Mammalian hU6 none S. pyogenes Puro, mCherry Guigo
BPK307978741 Mammalian U6 none As Cpf1 Joung
BPK308278742 Mammalian U6 none Lb Cpf1 Joung
pLentiCRISPR-E78852 Mammalian/Lentiviral U6 yes, cut (enhanced) S. pyogenes Puro Abbosh
pCFD3-MS2tail-Scaffold78906 Fly dU6:3 none S. pyogenes Virmilion Perrimon
pHDE-35S-Cas9-mCherry-UBQ78932 Plant U6-26 yes, cut S. pyogenes Hygro, mCherry Zhao
pU6-As-crRNA78956 Mammalian U6 none As Cpf1 Kim
pU6-Lb-crRNA78957 Mammalian U6 none Lb Cpf1 Kim
pU6-Fn-crRNA78958 Mammalian U6 none Fn Cpf1 Kim
pU6-Mb-crRNA78959 Mammalian U6 none Mb Cpf1 Kim
pCAG-SpCas9-GFP-U6-gRNA79144 Mammalian hU6 yes, cut S. pyogenes EGFP Zou
pCAG-eCas9-GFP-U6-gRNA79145 Mammalian hU6 yes, cut (enhanced) S. pyogenes EGFP Zou
pRS416gT-GalL-Cas979903 Yeast RPR1(TetO)  yes, cut S. pyogenes URA3 Davis
eSpCas9(1.1) 71814 Mammalian U6 yes, cut (enhanced) S. pyogenes Zhang
gRNA Cloning Vector Bbs I ver. 285586 Mammalian BbsI None S. pyogenes H Fujii
pX60161591 Mammalian/AAV BsaI yes, cut S. aureus Zhang
pX60261593 Mammalian/AAV BsaI yes, cut S. aureus Zhang
pSaGuide64710 Mammalian BbsI none S. aureus Musunuru
BPK230165778 Mammalian BsmBI none, need plasmid 65775 N. meningitidis Joung
VVT165779 Mammalian BsmBI none, need plasmid 65776 S. aureus Joung
BPK210165770 Bacteria BsmBI yes, cut S. aureus Joung

Multiplex gRNA Vectors

Several systems have been developed to generate multiplex gRNA vectors. These systems can be used to express multiple gRNAs from a single plasmid with or without expression of Cas9.

Plasmids Expression System Description PI
Multiplex CRISPR/Cas9 Assembly System Kit Mammalian A system for constructing all-in-one expression vectors containing multiple guide RNA expression cassettes and a Cas9 nuclease/nickase expression cassette using the Golden Gate cloning method. A separate accessory pack is available for dCas9 and FokI-dCas9. Yamamoto
Multiplex CRISPR/Cas9-based genome engineering from a single lentiviral vector Mammalian Lentiviral expression of Cas9, dCas9, or dCas9-VP64 along with 1-4 sgRNAs expressed from independent promoters. sgRNAs are cloned into separate vectors and then assembled into one construct using Golden Gate assembly. Gersbach
MuLE System Mammalian MultiSite Gateway recombination-based cloning of complex polycistronic lentiviruses. Can express Cas9 with up to 3 gRNAs. Frew
pSQT1313 Mammalian Single plasmid for the expression of multipe gRNAs. Csy4 cleavable cassettes; requires expression of Csy4 (eg Plasmid 53369). Joung
pX333 Mammalian Single all-in-one plasmid for tandem expression of two sgRNAs from independent U6 promoters and Cas9 from CBh promoter. Ventura
A CRISPR/Cas9 toolkit for multiplex genome editing in plants Plant Plasmids for six gRNA module vectors, including three designed for dicots and three designed for monocots. Using these gRNA module vectors, multiple gRNA expression cassettes can be assembled using Golden Gate cloning or the Gibson Assembly. Chen
Boosting CRISPR/Cas9 multiplex editing capability with the endogenous tRNA-processing system Plant Utilizes the tRNA processing system to reduce size and allow for expression of more sgRNAs in a single vector. sgRNAs are expressed as polycistronic glycine tRNA-gRNA genes (PTGs) and then PTG cassettes are assembled into a Cas9 expressing vector. Yang
A Robust CRISPR/Cas9 System for Convenient, High-Efficiency Multiplex Genome Editing in Monocot and Dicot Plants Plant PCR-based procedure to rapidly generate multiple sgRNA expression cassettes, which can be assembled into the binary CRISPR/Cas9 vectors in one round of cloning by Golden Gate ligation or Gibson Assembly Liu
A CRISPR/Cas9 toolbox for multiplexed plant genome editing and transcriptional regulation Plant Three types of plasmids available for use in this Golden Gate and Gateway based assembly method: plasmids for assembly of single gRNA vectors with various promoters, plasmids for assembling multiple gRNAs into one vector, and plasmids that provide Cas9 variants. All components are then assembled into a single T-DNA binary vector of choice through Gateway recombination. Qi
Multiplex Conditional Mutagenesis Using Transgenic Expression of Cas9 and sgRNAs Zebrafish Plasmids for assembly of gRNA cassettes using Golden Gate ligation into Tol2-based pGGDestTol2LC vector. Uses five distinct zebrafish U6 promoters for sgRNA expression. Chen
pCFD4-U6:1_U6:3tandemgRNAs Drosophila Single plasmid for the expression of two gRNAs from Drosophila U6:1 and U6:3 promoters. Bullock
pCFD5 Drosophila Utilizes endogenous tRNA processing machinery to excise multiple functional gRNAs from a single precursor transcript Bullock
pCRISPathBrick Bacteria Allows rapid modular assembly of natural type II-A CRISPR arrays capable of simultaneously repressing multiple target genes in Escherichia coli. Koffas


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