LOCUS       sequence-443304-        5903 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Co-expression of dead-mutant SNAP-tags to outer surface in mammalian
            cells coupled with an EGFP transfection marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5903)
  TITLE     Frank Lab Plasmids 2024
REFERENCE   2  (bases 1 to 5903)
  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"
COMMENT     Sequence Label: hSyn-pDisplay-SNAP[C145A]-T2A-EGFP
FEATURES             Location/Qualifiers
     source          1..5903
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     448..467
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   784..913
                     /label=AAV2 ITR
     polyA_signal    1075..1196
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1112..1131)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1166..1185
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1225..1241
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1226..1242
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1245..1833
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1351..1362
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1760..1780
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(1846..2559,2560..2562))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             complement(join(1849..2559,2560..2562))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1891..1912)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2240..2259
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2499..2520
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(2563..2616)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(2635..2781)
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     CDS             complement(2785..2814)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(3388..3414)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     sig_peptide     complement(3418..3477)
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     regulatory      3474..3483
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(3515..3962)
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     repeat_region   4044..4173
                     /label=AAV2 ITR
     rep_origin      4245..4700
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4332..4351)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4542..4563
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4726..4830
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4831..4899,4900..5691)
                     /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(5049..5068)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(5862..5903,1..547)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
       61 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
      121 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
      181 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
      241 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
      301 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
      361 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
      421 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
      481 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
      541 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
      601 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
      661 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
      721 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
      781 cagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc
      841 tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc
      901 actaggggtt ccttgtagtt aatgattaac ccgccatgct acttatctac gtagccatgc
      961 tctaggaaga tccttatcga ttttaccaca tttgtagagg ttttacttgc tttaaaaaac
     1021 ctcccacatc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     1081 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     1141 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     1201 gtctgctcga agcggccgcc cgggtcgagg tcgacggtat cgatgcgggg aggcggccca
     1261 aagggagatc cgactcgtct gagggcgaag gcgaagacgc ggaagaggcc gcagagccgg
     1321 cagcaggccg cgggaaggaa ggtccgctgg attgagggcc gaagggacgt agcagaagga
     1381 cgtcccgcgc agaatccagg tggcaacaca ggcgagcagc caaggaaagg acgatgattt
     1441 ccccgacaac accacggaat tgtcagtgcc caacagccga gcctctgtcc agcagcgggc
     1501 aaggcaggcg gcgatgagtt ccgccgtggc aatagggagg gggaaagcga aagtcccgga
     1561 aaggagctga caggtggtgg caatgcccca accagtgggg gttgcgtcag caaacacagt
     1621 gcacaccacg ccacgttgcc tgacaacggg ccacaactcc tcataaagag acagcaacca
     1681 ggatttatac aaggaggaga aaatgaaagc catacgggaa gcaatagcat gatacaaagg
     1741 cattaaagca gcgtatccac atagcgtaaa aggagcaaca tagttaagaa taccagtcaa
     1801 tctttcacaa attttgtaat ccagaggttg attatcgata agcttctact tgtacagctc
     1861 gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca tgtgatcgcg
     1921 cttctcgttg gggtctttgc tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag
     1981 cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga gctgcacgct
     2041 gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct tctgcttgtc
     2101 ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt
     2161 gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc
     2221 gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc
     2281 ctggacgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg
     2341 gtagcggctg aagcactgca cgccgtaggt cagggtggtc acgagggtgg gccagggcac
     2401 gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc
     2461 ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag
     2521 gatgggcacc accccggtga acagctcctc gcccttgctc actgggccag gattctcctc
     2581 gacgtcaccg catgttagca gacttcctct gccctctcca ctgccgcggc caccacgtgg
     2641 cttcttctgc caaagcatga tgaggatgat aagggagatg atggtgagca ccaccagggc
     2701 caggatggct gagatcacca ccaccttaaa gggcaaggag tgtggcacca cgatgacctc
     2761 ctgcgtgtcc tggcccacag cattcagatc ctcttctgag atgagttttt gttcgtcgac
     2821 gccgcccagg cctggtttac ccagacggtg accttcgtgc gccagcagcc attctttcac
     2881 agccaggccg ccttcgtagc cacctacgtc caggtcgccc tgaaccacac ggtgtgccgg
     2941 gatcagaatt ggcaccggat taccagacag agcggttttc accgccgcag ttgccgctgg
     3001 gttgccagcc agcgccgcca gatgggaata ggagatcacc tcgccgaatt tcactacttt
     3061 cagcagtttc cacagtacct gacgggtaaa ggattcctgc tgaaataccg ggtgatgcag
     3121 ggccggtaca gggaactctt caatcgcctc tggctggtga aaatatgcgt tcagccatgc
     3181 ggtagcctgc atcagcggct ccgggccacc cagaaccgca gccggagccg gtacttcgac
     3241 cgcatccgca gcgctcgtac ctttgcccag gaaaatgatg cggtgcaggc cctgctcgca
     3301 accgctcagt tccagtttac ccagcgggct gtccagggta gtgcgtttca tttcgcagtc
     3361 tttatctgat ctggccggct gggccccagc ataatctgga acatcatatg gatagtcacc
     3421 agtggaacct ggaacccaga gcagcagtac ccatagcagg agtgtgtctg tctccatggt
     3481 ggcgggtgga tccttgctag cagcttgaat tcgactgcgc tctcaggcac gacacgactc
     3541 ctccgctgcc caccgcagac tgaggcagcg ctgagtcgcc ggcgccgcag cgcagatggt
     3601 cgcgcccgtg cccccctatc tcgcgcctcg cgtggtgcgg tccggctggg ccggcggcgg
     3661 cgcggacgcg accaaggtgg ccgggaaggg gagtttgcgg gggaccggcg agtgacgtca
     3721 gcgcgccttc agtgctgagg cggcggtggc gcgcgccgcc aggcgggggc gaaggcactg
     3781 tccgcggtgc tgaagctggc agtgcgcacg cgcctcgccg catcctgttt cccctccccc
     3841 tctctgatag gggatgcgca atttggggaa tgggggttgg gtgcttgtcc agtgggtcgg
     3901 ggtcggtcgt caggtaggca cccccacccc gcctcatcct ggtcctaaaa cccacttgca
     3961 ctcatacgca gggccctctg cagagatctt cctagagcat ggctacgtag ataagtagca
     4021 tggcgggtta atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct
     4081 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     4141 ccgggcggcc tcagtgagcg agcgagcgcg cagctggcgt aatagcgaag aggcccgcac
     4201 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg
     4261 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     4321 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     4381 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     4441 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     4501 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     4561 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     4621 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     4681 aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     4741 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     4801 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     4861 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     4921 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     4981 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     5041 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     5101 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     5161 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     5221 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     5281 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     5341 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     5401 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     5461 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     5521 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     5581 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     5641 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     5701 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     5761 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     5821 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     5881 cgcgtaatct gctgcttgca aac
//
