LOCUS       sequence-194482-        9033 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Encodes dCas9(C) (without VP64) fused to VEGFR1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 9033)
  AUTHORS   Baeumler TA, Ahmed AA, Fulga TA
  TITLE     Engineering Synthetic Signaling Pathways with Programmable 
            dCas9-Based Chimeric Receptors.
  JOURNAL   Cell Rep. 2017 Sep 12;20(11):2639-2653. doi: 
            10.1016/j.celrep.2017.08.044.
  PUBMED    28903044
REFERENCE   2  (bases 1 to 9033)
  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.celrep.2017.08"; journalName: "Cell Rep"; date: 
            "2017-09-12- 12"; volume: "20"; issue: "11"; pages: "2639-2653"
COMMENT     Sequence Label: pVEGFR1_TEV(C)_TCS(Q'G)_NLS-HA-dCas9(C)
FEATURES             Location/Qualifiers
     source          1..9033
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     regulatory      228..237
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2979..2999
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3000..3020
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3039..3065
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(3093..4073,4074..4076,4077..5591)
                     /codon_start=1
                     /product="C-terminal portion of catalytically inactive 
                     Streptococcus pyogenes dCas9 carrying the N863A mutation 
                     (Zetsche et al., 2015)"
                     /label=dCas9(C)
                     /translation="KPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEI
                     SGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTY
                     AHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIH
                     DDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPE
                     NIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYL
                     QNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPSEEV
                     VKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQI
                     LDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVV
                     GTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITL
                     ANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESIL
                     PKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIME
                     RSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALP
                     SKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLD
                     KVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATL
                     IHQSITGLYETRIDLSQLGGD"
     CDS             5595..5615
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5661..5678)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5667..5874
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5883..6023
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5883..6012
                     /label=AAV2 ITR
     rep_origin      6098..6553
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6185..6204)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6395..6416
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6570..6589)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6689..6711
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6749..6767)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6835..6939
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6940..7008,7009..7800)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7158..7177)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7971..8559
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8460..8479
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        8631..8916
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
ORIGIN
        1 cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc ccggctctga
       61 ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc gggctgtaat
      121 tagctgagca agaggtaagg gtttaaggga tggttggttg gtggggtatt aatgtttaat
      181 tacctggagc acctgcctga aatcactttt tttcaggttg gaccggtgcc accatggtca
      241 gctactggga caccggggtc ctgctgtgcg cgctgctcag ctgtctgctt ctcacaggat
      301 ctagttcagg ttcaaaatta aaagatcctg aactgagttt aaaaggcacc cagcacatca
      361 tgcaagcagg ccagacactg catctccaat gcagggggga agcagcccat aaatggtctt
      421 tgcctgaaat ggtgagtaag gaaagcgaaa ggctgagcat aactaaatct gcctgtggaa
      481 gaaatggcaa acaattctgc agtactttaa ccttgaacac agctcaagca aaccacactg
      541 gcttctacag ctgcaaatat ctagctgtac ctacttcaaa gaagaaggaa acagaatctg
      601 caatctatat atttattagt gatacaggta gacctttcgt agagatgtac agtgaaatcc
      661 ccgaaattat acacatgact gaaggaaggg agctcgtcat tccctgccgg gttacgtcac
      721 ctaacatcac tgttacttta aaaaagtttc cacttgacac tttgatccct gatggaaaac
      781 gcataatctg ggacagtaga aagggcttca tcatatcaaa tgcaacgtac aaagaaatag
      841 ggcttctgac ctgtgaagca acagtcaatg ggcatttgta taagacaaac tatctcacac
      901 atcgacaaac caatacaatc atagatgtcc aaataagcac accacgccca gtcaaattac
      961 ttagaggcca tactcttgtt ctcaattgta ctgctaccac tcccttgaac acgagagttc
     1021 aaatgacctg gagttaccct gatgaaaaaa ataagagagc ttccgtaagg cgacgaattg
     1081 accaaagcaa ttcccatgcc aacatattct acagtgttct tactattgac aaaatgcaga
     1141 acaaagacaa aggactttat acttgtcgtg taaggagtgg accatcattc aaatctgtta
     1201 acacctcagt gcatatatat gataaagcat tcatcactgt gaaacatcga aaacagcagg
     1261 tgcttgaaac cgtagctggc aagcggtctt accggctctc tatgaaagtg aaggcatttc
     1321 cctcgccgga agttgtatgg ttaaaagatg ggttacctgc gactgagaaa tctgctcgct
     1381 atttgactcg tggctactcg ttaattatca aggacgtaac tgaagaggat gcagggaatt
     1441 atacaatctt gctgagcata aaacagtcaa atgtgtttaa aaacctcact gccactctaa
     1501 ttgtcaatgt gaaaccccag atttacgaaa aggctgtgtc atcgtttcca gacccggctc
     1561 tctacccact gggcagcaga caaatcctga cttgtaccgc atatggtatc cctcaaccta
     1621 caatcaagtg gttctggcac ccctgtaacc ataatcattc cgaagcaagg tgtgactttt
     1681 gttccaataa tgaagagtcc tttatcctgg atgctgacag caacatggga aacagaattg
     1741 agagcatcac tcagcgcatg gcaataatag aaggaaagaa taagatggct agcaccttgg
     1801 ttgtggctga ctctagaatt tctggaatct acatttgcat agcttccaat aaagttggga
     1861 ctgtgggaag aaacataagc ttttatatca cagatgtgcc aaatgggttt catgttaact
     1921 tggaaaaaat gccgacggaa ggagaggacc tgaaactgtc ttgcacagtt aacaagttct
     1981 tatacagaga cgttacttgg attttactgc ggacagttaa taacagaaca atgcactaca
     2041 gtattagcaa gcaaaaaatg gccatcacta aggagcactc catcactctt aatcttacca
     2101 tcatgaatgt ttccctgcaa gattcaggca cctatgcctg cagagccagg aatgtataca
     2161 caggggaaga aatcctccag aagaaagaaa ttacaatcag agatcaggaa gcaccatacc
     2221 tcctgcgaaa cctcagtgat cacacagtgg ccatcagcag ttccaccact ttagactgtc
     2281 atgctaatgg tgtccccgag cctcagatca cttggtttaa aaacaaccac aaaatacaac
     2341 aagagcctgg aattatttta ggaccaggaa gcagcacgct gtttattgaa agagtcacag
     2401 aagaggatga aggtgtctat cactgcaaag ccaccaacca gaagggctct gtggaaagtt
     2461 cagcatacct cactgttcaa ggaacctcgg acaagcctaa tctggagctg atcactctaa
     2521 catgcacctg tgtggctgcg actctcttct ggctcctatt aaccctcttt atcggcggtg
     2581 gctcgggtgg tggctcgaag agcatgtcta gcatggtgtc agacactagt tgcacattcc
     2641 cttcatctga tggcatattc tggaagcatt ggattcaaac caaggatggg cagtgtggca
     2701 gtccattagt atcaactaga gatgggttca ttgttggtat acactcagca tcgaatttca
     2761 ccaacacaaa caattatttc acaagcgtgc cgaaaaactt catggaattg ttgacaaatc
     2821 aggaggcgca gcagtgggtt agtggttggc gattaaatgc tgactcagta ttgtgggggg
     2881 gccataaagt tttcatggtg aaacctgaag agccttttca gccagttaag gaagcgactc
     2941 aactcatgaa tcgtcgtcgc cgtcccggcg gtggctcgga aaacctgtat tttcaggggc
     3001 ccaagaagaa gaggaaggtg gggggaggca gcactagtta cccatacgac gtaccagatt
     3061 acgctggagg ttctggatcc ggcggtggct cgaagcccgc cttcctgagc ggcgagcaga
     3121 aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga
     3181 aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag
     3241 atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg
     3301 acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac
     3361 tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg
     3421 acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga
     3481 agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt
     3541 ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta
     3601 aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg
     3661 ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg
     3721 acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca
     3781 gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg
     3841 aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc
     3901 agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg
     3961 accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc gtgcctcaga
     4021 gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac aaggcccggg
     4081 gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcggc
     4141 agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc aaggccgaga
     4201 gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc
     4261 ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg
     4321 agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg
     4381 atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc
     4441 acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg
     4501 aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga
     4561 gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact
     4621 ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga
     4681 caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc accgtgcgga
     4741 aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct
     4801 tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc agaaagaagg
     4861 actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg
     4921 tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg
     4981 ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca
     5041 agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg
     5101 agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg
     5161 aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc
     5221 tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaagcact
     5281 acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg
     5341 ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc atcagagagc
     5401 aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca
     5461 agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg
     5521 ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc
     5581 tgggaggcga cagccccaag aagaagagaa aggtggaggc cagcggaggc agctaagaat
     5641 tcctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     5701 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     5761 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     5821 ggggcaggac agcaaggggg aggattggga agagaatagc aggcatgctg gggagcggcc
     5881 gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga
     5941 ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga
     6001 gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg
     6061 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     6121 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg
     6181 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     6241 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     6301 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     6361 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     6421 acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc
     6481 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     6541 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     6601 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     6661 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     6721 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     6781 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     6841 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     6901 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     6961 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     7021 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     7081 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     7141 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     7201 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     7261 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     7321 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     7381 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     7441 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     7501 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     7561 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     7621 tttattgctg ataaatctgg agccggtgag cgtggaagcc gcggtatcat tgcagcactg
     7681 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     7741 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     7801 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     7861 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     7921 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     7981 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     8041 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     8101 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     8161 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     8221 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     8281 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     8341 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     8401 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     8461 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     8521 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     8581 ttacggttcc tggccttttg ctggcctttt gctcacatgc tagaggtacc cgttacataa
     8641 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
     8701 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
     8761 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
     8821 ggtaaatggc ccgcctggca ttgtgcccag tacatgacct tatgggactt tcctacttgg
     8881 cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc cacgttctgc
     8941 ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt tattttttaa
     9001 ttattttgtg cagcgatggg ggcggggggg ggg
//
