LOCUS       sequence-194465-        9355 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Encodes dCas9(N) fused to VEGFR2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 9355)
  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 9355)
  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: 
            pVEGFR2_TEV(N)_NES_TCS(Q'L)_HA-dCas9(N)_P2A_Puro-WPRE
FEATURES             Location/Qualifiers
     source          1..9355
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             3054..3080
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(3096..3119,3120..3122,3123..4697)
                     /codon_start=1
                     /product="N-terminal portion of catalytically inactive 
                     Streptococcus pyogenes dCas9 carrying the D10A mutation 
                     (Zetsche et al., 2015)"
                     /label=dCas9(N)
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMR"
     CDS             4728..4784
                     /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"
     primer_bind     complement(4791..4810)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5287..5307
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(5424..5444)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5842..5853)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5984..6001)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5990..6197
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6206..6346
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6206..6335
                     /label=AAV2 ITR
     rep_origin      6421..6876
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6508..6527)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6718..6739
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6893..6912)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7012..7034
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7072..7090)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7158..7262
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7263..7331,7332..8123)
                     /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(7481..7500)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8294..8882
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8783..8802
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        8953..9238
                     /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 accatgcaga
      241 gcaaggtgct gctggccgtc gccctgtggc tctgcgtgga gacccgggcc gcctctgtgg
      301 gtttgcctag tgtttctctt gatctgccca ggctcagcat acaaaaagac atacttacaa
      361 ttaaggctaa tacaactctt caaattactt gcaggggaca gagggacttg gactggcttt
      421 ggcccaataa tcagagtggc agtgagcaaa gggtggaggt gactgagtgc agcgatggcc
      481 tcttctgtaa gacactcaca attccaaaag tgatcggaaa tgacactgga gcctacaagt
      541 gcttctaccg ggaaactgac ttggcctcgg tcatttatgt ctatgttcaa gattacagat
      601 ctccatttat tgcttctgtt agtgaccaac atggagtcgt gtacattact gagaacaaaa
      661 acaaaactgt ggtgattcca tgtctcgggt ccatttcaaa tctcaacgtg tcactttgtg
      721 caagataccc agaaaagaga tttgttcctg atggtaacag aatttcctgg gacagcaaga
      781 agggctttac tattcccagc tacatgatca gctatgctgg catggtcttc tgtgaagcaa
      841 aaattaatga tgaaagttac cagtctatta tgtacatagt tgtcgttgta gggtatagga
      901 tttatgatgt ggttctgagt ccgtctcatg gaattgaact atctgttgga gaaaagcttg
      961 tcttaaattg tacagcaaga actgaactaa atgtggggat tgacttcaac tgggaatacc
     1021 cttcttcgaa gcatcagcat aagaaacttg taaaccgaga cctaaaaacc cagtctggga
     1081 gtgagatgaa gaaatttttg agcaccttaa ctatagatgg tataacccgg agtgaccaag
     1141 gattgtacac ctgtgcagca tccagtgggc tgatgaccaa gaagaacagc acatttgtca
     1201 gggtccatga aaaacctttt gttgcttttg gaagtggcat ggaatctctg gtggaagcca
     1261 cggtggggga gcgtgtcaga atccctgcga agtaccttgg ttacccaccc ccagaaataa
     1321 aatggtataa aaatggaata ccccttgagt ccaatcacac aattaaagcg gggcatgtac
     1381 tgacgattat ggaagtgagt gaaagagaca caggaaatta cactgtcatc cttaccaatc
     1441 ccatttcaaa ggagaagcag agccatgtgg tctctctggt tgtgtatgtc ccaccccaga
     1501 ttggtgagaa atctctaatc tctcctgtgg attcctacca gtacggcacc actcaaacgc
     1561 tgacatgtac ggtctatgcc attcctcccc cgcatcacat ccactggtat tggcagttgg
     1621 aggaagagtg cgccaacgag cccagccaag ctgtctcagt gacaaaccca tacccttgtg
     1681 aagaatggag aagtgtggag gacttccagg gaggaaataa aattgaagtt aataaaaatc
     1741 aatttgctct aattgaagga aaaaacaaaa ctgtaagtac ccttgttatc caagcggcaa
     1801 atgtgtcagc tttgtacaaa tgtgaagcgg tcaacaaagt cgggagagga gagagggtga
     1861 tctccttcca cgtgaccagg ggtcctgaaa ttactttgca acctgacatg cagcccactg
     1921 agcaggagag cgtgtctttg tggtgcactg cagacagatc tacgtttgag aacctcacat
     1981 ggtacaagct tggcccacag cctctgccaa tccatgtggg agagttgccc acacctgttt
     2041 gcaagaactt ggatactctt tggaaattga atgccaccat gttctctaat agcacaaatg
     2101 acattttgat catggagctt aagaatgcat ccttgcagga ccaaggagac tatgtctgcc
     2161 ttgctcaaga caggaagacc aagaaaagac attgcgtggt caggcagctc acagtcctag
     2221 agcgtgtggc acccacgatc acaggaaacc tggagaatca gacgacaagt attggggaaa
     2281 gcatcgaagt ctcatgcacg gcatctggga atccccctcc acagatcatg tggtttaaag
     2341 ataatgagac ccttgtagaa gactcaggca ttgtattgaa ggatgggaac cggaacctca
     2401 ctatccgcag agtgaggaag gaggacgaag gcctctacac ctgccaggca tgcagtgttc
     2461 ttggctgtgc aaaagtggag gcatttttca taatagaagg tgcccaggaa aagacgaact
     2521 tggaaatcat tattctagta ggcacggcgg tgattgccat gttcttctgg ctacttcttg
     2581 tcatcatcgg cggtggctcg ggtggtggct cgggagaaag cttgtttaag gggccgcgtg
     2641 attacaaccc gatatcgagc accatttgtc atttgacgaa tgaatctgat gggcacacaa
     2701 catcgttgta tggtattgga tttggtccct tcatcattac aaacaagcac ttgtttagaa
     2761 gaaataatgg aacactgttg gtccaatcac tacatggtgt attcaaggtc aagaacacca
     2821 cgactttgca acaacacctc attgatggga gggacatgat aattattcgc atgcctaagg
     2881 atttcccacc atttcctcaa aagctgaaat ttagagagcc acaaagggaa gagcgcatat
     2941 gtcttgtgac aaccaacttc caaactggcg gtggctcgct agatttagct agcttaatcc
     3001 tcggcaagct tggcgaaaac ctgtattttc agctaggcgg aggcagcact agttacccat
     3061 acgacgtacc agattacgct ggaggttctg gatccgacaa gaagtacagc atcggcctgg
     3121 ccatcggcac caactctgtg ggctgggccg tgatcaccga cgagtacaag gtgcccagca
     3181 agaaattcaa ggtgctgggc aacaccgacc ggcacagcat caagaagaac ctgatcggag
     3241 ccctgctgtt cgacagcggc gaaacagccg aggccacccg gctgaagaga accgccagaa
     3301 gaagatacac cagacggaag aaccggatct gctatctgca agagatcttc agcaacgaga
     3361 tggccaaggt ggacgacagc ttcttccaca gactggaaga gtccttcctg gtggaagagg
     3421 ataagaagca cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg
     3481 agaagtaccc caccatctac cacctgagaa agaaactggt ggacagcacc gacaaggccg
     3541 acctgcggct gatctatctg gccctggccc acatgatcaa gttccggggc cacttcctga
     3601 tcgagggcga cctgaacccc gacaacagcg acgtggacaa gctgttcatc cagctggtgc
     3661 agacctacaa ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg
     3721 ccatcctgtc tgccagactg agcaagagca gacggctgga aaatctgatc gcccagctgc
     3781 ccggcgagaa gaagaatggc ctgttcggca acctgattgc cctgagcctg ggcctgaccc
     3841 ccaacttcaa gagcaacttc gacctggccg aggatgccaa actgcagctg agcaaggaca
     3901 cctacgacga cgacctggac aacctgctgg cccagatcgg cgaccagtac gccgacctgt
     3961 ttctggccgc caagaacctg tccgacgcca tcctgctgag cgacatcctg agagtgaaca
     4021 ccgagatcac caaggccccc ctgagcgcct ctatgatcaa gagatacgac gagcaccacc
     4081 aggacctgac cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag tacaaagaga
     4141 ttttcttcga ccagagcaag aacggctacg ccggctacat tgacggcgga gccagccagg
     4201 aagagttcta caagttcatc aagcccatcc tggaaaagat ggacggcacc gaggaactgc
     4261 tcgtgaagct gaacagagag gacctgctgc ggaagcagcg gaccttcgac aacggcagca
     4321 tcccccacca gatccacctg ggagagctgc acgccattct gcggcggcag gaagattttt
     4381 acccattcct gaaggacaac cgggaaaaga tcgagaagat cctgaccttc cgcatcccct
     4441 actacgtggg ccctctggcc aggggaaaca gcagattcgc ctggatgacc agaaagagcg
     4501 aggaaaccat caccccctgg aacttcgagg aagtggtgga caagggcgct tccgcccaga
     4561 gcttcatcga gcggatgacc aacttcgata agaacctgcc caacgagaag gtgctgccca
     4621 agcacagcct gctgtacgag tacttcaccg tgtataacga gctgaccaaa gtgaaatacg
     4681 tgaccgaggg aatgagaggt ggcggtggct cgggtaccgg atctggagcc accaattttt
     4741 ctctgctgaa gcaggctggt gatgttgaag aaaaccctgg acctgaattc atgaccgagt
     4801 acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg
     4861 ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg
     4921 agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg
     4981 tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag
     5041 cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg
     5101 ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt
     5161 tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg
     5221 tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg
     5281 cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg
     5341 tgcccgaagg accgcgcacc tggtgcatga cccgcaaggg gtaactaagt aaggatcgat
     5401 ccaagatatc gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     5461 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     5521 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     5581 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     5641 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     5701 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     5761 gggaagctga cgtcctttcc atggctgctc gcctgtgttg ccacctggat tctgcgcggg
     5821 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     5881 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     5941 ctttgggccg cctccccgcc aattcctaga gctcgctgat cagcctcgac tgtgccttct
     6001 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     6061 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     6121 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagagaat
     6181 agcaggcatg ctggggagcg gccgcaggaa cccctagtga tggagttggc cactccctct
     6241 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     6301 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     6361 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     6421 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     6481 ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     6541 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     6601 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     6661 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     6721 gactcttgtt ccaaactgga acaacactca actctatctc gggctattct tttgatttat
     6781 aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta
     6841 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     6901 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     6961 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     7021 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     7081 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     7141 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     7201 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     7261 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     7321 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     7381 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     7441 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     7501 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     7561 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     7621 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     7681 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     7741 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     7801 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     7861 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     7921 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtggaa
     7981 gccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     8041 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     8101 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     8161 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     8221 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     8281 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     8341 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     8401 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     8461 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     8521 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     8581 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     8641 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     8701 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     8761 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     8821 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     8881 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     8941 tctagaggta cccgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     9001 cccccgccca ttgacgtcaa tagtaacgcc aatagggact ttccattgac gtcaatgggt
     9061 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
     9121 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattgtgccc agtacatgac
     9181 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
     9241 cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat
     9301 tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg ggggg
//
