LOCUS       sequence-231527-        6510 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  expression of AB1-fused-dcas9 plasmids.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6510)
  AUTHORS   Wang H, Xu X, Nguyen CM, Liu Y, Gao Y, Lin X, Daley T, Kipniss NH, 
            La Russa M, Qi LS
  TITLE     CRISPR-Mediated Programmable 3D Genome Positioning and Nuclear 
            Organization.
  JOURNAL   Cell. 2018 Nov 15;175(5):1405-1417.e14. doi: 
            10.1016/j.cell.2018.09.013. Epub 2018 Oct 11.
  PUBMED    30318144
REFERENCE   2  (bases 1 to 6510)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2018.09.013"; journalName: "Cell"; date: 
            "2018-11-15- 15"; volume: "175"; issue: "5"; pages: "1405-1417.e14"
COMMENT     Sequence Label: pHR-PGK-ABI-dCas9-P2A-mcherry
FEATURES             Location/Qualifiers
     source          1..6510
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        5..504
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(40..63)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     437..456
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             join(1495..1521,1522..1524,1525..4011,4012..4014,
                     4015..5598)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             1495..5598
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5602..5628
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             5608..5634
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             5647..5667
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5674..5694
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5743..5799
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5800..6510
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(5945..5963)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     6213..6232
                     /label=mCherry-F
                     /note="mCherry, forward primer"
ORIGIN
        1 taccgggtag gggaggcgct tttcccaagg cagtctggag catgcgcttt agcagccccg
       61 ctgggcactt ggcgctacac aagtggcctc tggcctcgca cacattccac atccaccggt
      121 aggcgccaac cggctccgtt ctttggtggc cccttcgcgc caccttctac tcctccccta
      181 gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca ggacgtgaca aatggaagta
      241 gcacgtctca ctagtctcgt gcagatggac agcaccgctg agcaatggaa gcgggtaggc
      301 ctttggggca gcggccaata gcagctttgc tccttcgctt tctgggctca gaggctggga
      361 aggggtgggt ccgggggcgg gctcaggggc gggctcaggg gcggggcggg cgcccgaagg
      421 tcctccggag gcccggcatt ctgcacgctt caaaagcgca cgtctgccgc gctgttctcc
      481 tcttcctcat ctccgggcct ttcgacctgc agcccaagct tacgcgtctg caggatatca
      541 agcttgcggt accgcgggcc cggggccacc atgacgcgtg tgcctttgta tggttttact
      601 tcgatttgtg gaagaagacc tgagatggaa gctgctgttt cgactatacc aagattcctt
      661 caatcttcct ctggttcgat gttagatggt cggtttgatc ctcaatccgc cgctcatttc
      721 ttcggtgttt acgacggcca tggcggttct caggtagcga actattgtag agagaggatg
      781 catttggctt tggcggagga gatagctaag gagaaaccga tgctctgcga tggtgatacg
      841 tggctggaga agtggaagaa agctcttttc aactcgttcc tgagagttga ctcggagatt
      901 gagtcagttg cgccggagac ggttgggtca acgtcggtgg ttgccgttgt tttcccgtct
      961 cacatcttcg tcgctaactg cggtgactct agagccgttc tttgccgcgg caaaactgca
     1021 cttccattat ccgttgacca taaaccggat agagaagatg aagctgcgag gattgaagcc
     1081 gcaggaggga aagtgattca gtggaatgga gctcgtgttt tcggtgttct cgccatgtcg
     1141 agatccattg gcgatagata cttgaaacca tccatcattc ctgatccgga agtgacggct
     1201 gtgaagagag taaaagaaga tgattgtctg attttggcga gtgacggggt ttgggatgta
     1261 atgacggatg aagaagcgtg tgagatggca aggaagcgga ttctcttgtg gcacaagaaa
     1321 aacgcggtgg ctggggatgc atcgttgctc gcggatgagc ggagaaagga agggaaagat
     1381 cctgcggcga tgtccgcggc tgagtatttg tcaaagctgg cgatacagag aggaagcaaa
     1441 gacaacataa gtgtggtggt ggttgatttg aagcatatgg gtggaggctc cgggatggac
     1501 aagaagtaca gcatcggcct ggccatcggc accaactctg tgggctgggc cgtgatcacc
     1561 gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc
     1621 atcaagaaga acctgatcgg cgccctgctg ttcgacagcg gagaaacagc cgaggccacc
     1681 cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg
     1741 caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa
     1801 gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc
     1861 gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg
     1921 gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc
     1981 aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac
     2041 aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac
     2101 gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg
     2161 gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg caacctgatt
     2221 gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc
     2281 aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc
     2341 ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg
     2401 agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc
     2461 aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag
     2521 ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac
     2581 atcgatggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag
     2641 atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag
     2701 cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt
     2761 ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag
     2821 atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc
     2881 gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg
     2941 gacaagggcg ccagcgccca gagcttcatc gagcggatga ccaacttcga taagaacctg
     3001 cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtacaac
     3061 gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc
     3121 gagcagaaaa aagccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag
     3181 cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc
     3241 gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag
     3301 gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc
     3361 ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg
     3421 ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg
     3481 agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc
     3541 ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg
     3601 acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag
     3661 cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag
     3721 gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa
     3781 atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag
     3841 cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa
     3901 aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg
     3961 tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga cgctatcgtg
     4021 cctcagagct ttctgaagga cgactccatc gataacaaag tgctgactcg gagcgacaag
     4081 aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac
     4141 tggcgccagc tgctgaatgc caagctgatt acccagagga agttcgacaa tctgaccaag
     4201 gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg
     4261 gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag
     4321 tacgacgaga acgacaaact gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg
     4381 gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac
     4441 cacgcccacg acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct
     4501 aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc
     4561 gccaagagcg agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc
     4621 atgaactttt tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg
     4681 atcgagacaa acggcgaaac aggcgagatc gtgtgggata agggccggga ctttgccacc
     4741 gtgcggaaag tgctgtctat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca
     4801 ggcggcttca gcaaagagtc tatcctgccc aagaggaaca gcgacaagct gatcgccaga
     4861 aagaaggact gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct
     4921 gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag
     4981 ctgctgggga tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg
     5041 gaagccaagg gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc
     5101 ctgttcgagc tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag
     5161 ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat
     5221 gagaagctga agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac
     5281 aaacactacc tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg
     5341 gccgacgcta atctggacaa ggtgctgagc gcctacaaca agcacagaga caagcctatc
     5401 agagagcagg ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc
     5461 gccttcaagt actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg
     5521 ctggacgcca ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg
     5581 tctcagctgg gaggcgacgc ctatccctat gacgtgcccg attatgccag cctgggcagc
     5641 ggctccccca agaaaaaacg caaggtggaa gatcctaaga aaaagcggaa agtggacggc
     5701 attggtagtg ggagcaacgg cagcagcgga tccggaagcg gagctactaa cttcagcctg
     5761 ctgaagcagg ctggagacgt ggaggagaac cctggaccta tggtgagcaa gggcgaggag
     5821 gataacatgg ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg
     5881 aacggccacg agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag
     5941 accgccaagc tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc
     6001 cctcagttca tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac
     6061 ttgaagctgt ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc
     6121 ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg
     6181 aagctgcgcg gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc
     6241 tgggaggcct cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag
     6301 cagaggctga agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag
     6361 gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc
     6421 tcccacaacg aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc
     6481 accggcggca tggacgagct gtacaagtaa
//
