LOCUS       sequence-155831-        5847 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid that expresses Cas9C in mammalian cells; co-expresses 
            turboGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5847)
  AUTHORS   Chew WL, Tabebordbar M, Cheng JK, Mali P, Wu EY, Ng AH, Zhu K, 
            Wagers AJ, Church GM
  TITLE     A multifunctional AAV-CRISPR-Cas9 and its host response.
  JOURNAL   Nat Methods. 2016 Sep 5. doi: 10.1038/nmeth.3993.
  PUBMED    27595405
REFERENCE   2  (bases 1 to 5847)
  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"; journalName: "Nat 
            Methods. 2016 Sep 5. doi: 10.1038/nmeth.3993."
COMMENT     Sequence Label: pSMVP-Cas9C-P2A-turboGFP
FEATURES             Location/Qualifiers
     source          1..5847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(44..68)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(75..95)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     enhancer        325..704
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        705..908
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     859..879
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          1000..1132
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1162..1171
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1166..1175
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1772..1783
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             3301..3321
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3340..3396
                     /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             3397..4062
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina 
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /label=CopGFP
                     /translation="MPAMKIECRITGTLNGVEFELVGGGEGTPEQGRMTNKMKSTKGAL
                     TFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYR
                     YEAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDG
                     GYYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA
                     "
     CDS             3397..4062
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina 
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /label=CopGFP
                     /translation="MPAMKIECRITGTLNGVEFELVGGGEGTPEQGRMTNKMKSTKGAL
                     TFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYR
                     YEAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDG
                     GYYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA
                     "
     primer_bind     complement(4076..4095)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    4105..4186
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     4130..4149
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(4384..4972)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4464..4483)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5053..5847)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(5053..5847)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5164..5183)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5774..5793
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 aatattattg aagcatttat cagggttcgt ctcgtccctc tagacccagc tttcttgtac
       61 aaagttggca ttatggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
      121 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accagtcccg gtctcctccc
      181 atgcatgtca atattggcca ttagccatat tattcattgg ttatatagca taaatcaata
      241 ttggctattg gccattgcat acgttgtatc tatatcataa tatgtacatt tatattggct
      301 catgtccaat atgaccgcca tgttggcatt gattattgac tagttattaa tagtaatcaa
      361 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      421 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      481 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      541 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtccgccc cctattgacg
      601 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta cgggactttc
      661 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
      721 agtacaccaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
      781 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
      841 ataaccccgc cccgttgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
      901 gcagaggtcg tttagtgaac cgtcagatca ctagtagctt tattgcggta gtttatcaca
      961 gttaaattgc taacgcagtc agtgctcgac tgatcacagg taagtatcaa ggttacaaga
     1021 caggtttaag gaggccaata gaaactgggc ttgtcgagac agagaagatt cttgcgtttc
     1081 tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac aggggtaccg
     1141 aagccgctag cgctaccggt cgccaccatg gcggcggcgt gcccggaact gcgtcagctg
     1201 gcgcagagcg atgtgtattg ggatccgatt gtgagcattg aaccggatgg cgtggaagaa
     1261 gtgtttgatc tgaccgtgcc gggcccgcat aactttgtgg cgaacgatat tattgcgcat
     1321 aactctggcc agggggacag tcttcacgag cacatcgcta atcttgcagg tagcccagct
     1381 atcaaaaagg gaatactgca gaccgttaag gtcgtggatg aactcgtcaa agtaatggga
     1441 aggcataagc ccgagaatat cgttatcgag atggcccgag agaaccaaac tacccagaag
     1501 ggacagaaga acagtaggga aaggatgaag aggattgaag agggtataaa agaactgggg
     1561 tcccaaatcc ttaaggaaca cccagttgaa aacacccagc ttcagaatga gaagctctac
     1621 ctgtactacc tgcagaacgg cagggacatg tacgtggatc aggaactgga catcaatcgg
     1681 ctctccgact acgacgtgga tcatatcgtg ccccagtctt ttctcaaaga tgattctatt
     1741 gataataaag tgttgacaag atccgataaa aatagaggga agagtgataa cgtcccctca
     1801 gaagaagttg tcaagaaaat gaaaaattat tggcggcagc tgctgaacgc caaactgatc
     1861 acacaacgga agttcgataa tctgactaag gctgaacgag gtggcctgtc tgagttggat
     1921 aaagccggct tcatcaaaag gcagcttgtt gagacacgcc agatcaccaa gcacgtggcc
     1981 caaattctcg attcacgcat gaacaccaag tacgatgaaa atgacaaact gattcgagag
     2041 gtgaaagtta ttactctgaa gtctaagctg gtctcagatt tcagaaagga ctttcagttt
     2101 tataaggtga gagagatcaa caattaccac catgcgcatg atgcctacct gaatgcagtg
     2161 gtaggcactg cacttatcaa aaaatatccc aagcttgaat ctgaatttgt ttacggagac
     2221 tataaagtgt acgatgttag gaaaatgatc gcaaagtctg agcaggaaat aggcaaggcc
     2281 accgctaagt acttctttta cagcaatatt atgaattttt tcaagaccga gattacactg
     2341 gccaatggag agattcggaa gcgaccactt atcgaaacaa acggagaaac aggagaaatc
     2401 gtgtgggaca agggtaggga tttcgcgaca gtccggaagg tcctgtccat gccgcaggtg
     2461 aacatcgtta aaaagaccga agtacagacc ggaggcttct ccaaggaaag tatcctcccg
     2521 aaaaggaaca gcgacaagct gatcgcacgc aaaaaagatt gggaccccaa gaaatacggc
     2581 ggattcgatt ctcctacagt cgcttacagt gtactggttg tggccaaagt ggagaaaggg
     2641 aagtctaaaa aactcaaaag cgtcaaggaa ctgctgggca tcacaatcat ggagcgatca
     2701 agcttcgaaa aaaaccccat cgactttctc gaggcgaaag gatataaaga ggtcaaaaaa
     2761 gacctcatca ttaagcttcc caagtactct ctctttgagc ttgaaaacgg ccggaaacga
     2821 atgctcgcta gtgcgggcga gctgcagaaa ggtaacgagc tggcactgcc ctctaaatac
     2881 gttaatttct tgtatctggc cagccactat gaaaagctca aagggtctcc cgaagataat
     2941 gagcagaagc agctgttcgt ggaacaacac aaacactacc ttgatgagat catcgagcaa
     3001 ataagcgaat tctccaaaag agtgatcctc gccgacgcta acctcgataa ggtgctttct
     3061 gcttacaata agcacaggga taagcccatc agggagcagg cagaaaacat tatccacttg
     3121 tttactctga ccaacttggg cgcgcctgca gccttcaagt acttcgacac caccatagac
     3181 agaaagcggt acacctctac aaaggaggtc ctggacgcca cactgattca tcagtcaatt
     3241 acggggctct atgaaacaag aatcgacctc tctcagctcg gtggagacag cagggctgac
     3301 cccaagaaga agaggaaggt gtctcgagct ggatccggag ccacgaactt ctctctgtta
     3361 aagcaagcag gggacgtgga agaaaacccc ggtcctatgc ccgccatgaa gatcgagtgc
     3421 cgcatcaccg gcaccctgaa cggcgtggag ttcgagctgg tgggcggcgg agagggcacc
     3481 cccgagcagg gccgcatgac caacaagatg aagagcacca aaggcgccct gaccttcagc
     3541 ccctacctgc tgagccacgt gatgggctac ggcttctacc acttcggcac ctaccccagc
     3601 ggctacgaga accccttcct gcacgccatc aacaacggcg gctacaccaa cacccgcatc
     3661 gagaagtacg aggacggcgg cgtgctgcac gtgagcttca gctaccgcta cgaggccggc
     3721 cgcgtgatcg gcgacttcaa ggtggtgggc accggcttcc ccgaggacag cgtgatcttc
     3781 accgacaaga tcatccgcag caacgccacc gtggagcacc tgcaccccat gggcgataac
     3841 gtgctggtgg gcagcttcgc ccgcaccttc agcctgcgcg acggcggcta ctacagcttc
     3901 gtggtggaca gccacatgca cttcaagagc gccatccacc ccagcatcct gcagaacggg
     3961 ggccccatgt tcgccttccg ccgcgtggag gagctgcaca gcaacaccga gctgggcatc
     4021 gtggagtacc agcacgcctt caagaccccc atcgccttcg ccagatctcg agctcgatga
     4081 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     4141 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     4201 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa
     4261 cctctacaaa tgtggtactt aagaggggga gaccaaaggg cgagacgtta aggcctcacg
     4321 tgacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4381 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4441 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4501 gcgctctcct gttccgaccc tgccgcttac gggatacctg tccgcctttc tcccttcggg
     4561 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4621 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4681 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4741 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4801 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4861 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4921 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4981 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     5041 ggtcatgccg tctcagaaga actcgtcaag aaggcgatag aaggcgatgc gctgcgaatc
     5101 gggagcggcg ataccgtaaa gcacgaggaa gcggtcagcc cattcgccgc caagctcttc
     5161 agcaatatca cgggtagcca acgctatgtc ctgatagcgg tccgccacac ccagccggcc
     5221 acagtcgatg aatccagaaa agcggccatt ttccaccatg atattcggca agcaggcatc
     5281 gccatgggtc acgacgagat cctcgccgtc gggcatgctc gccttgagcc tggcgaacag
     5341 ttcggctggc gcgagcccct gatgctcttc gtccagatca tcctgatcga caagaccggc
     5401 ttccatccga gtacgtgctc gctcgatgcg atgtttcgct tggtggtcga atgggcaggt
     5461 agccggatca agcgtatgca gccgccgcat tgcatcagcc atgatggata ctttctcggc
     5521 aggagcaagg tgagatgaca ggagatcctg ccccggcact tcgcccaata gcagccagtc
     5581 ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa ggaacgcccg tcgtggccag
     5641 ccacgatagc cgcgctgcct cgtcttgcag ttcattcagg gcaccggaca ggtcggtctt
     5701 gacaaaaaga accgggcgcc cctgcgctga cagccggaac acggcggcat cagagcagcc
     5761 gattgtctgt tgtgcccagt catagccgaa tagcctctcc acccaagcgg ccggagaacc
     5821 tgcgtgcaat ccatcttgtt caatcat
//
