LOCUS       sequence-476057-        7426 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mGFP and Synaptophysin-mRuby expression from Flp-expressing neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7426)
  AUTHORS   Beier KT, Steinberg EE, DeLoach KE, Xie S, Miyamichi K, Schwarz L, 
            Gao XJ, Kremer EJ, Malenka RC, Luo L
  TITLE     Circuit Architecture of VTA Dopamine Neurons Revealed by Systematic 
            Input-Output Mapping.
  JOURNAL   Cell. 2015 Jul 30;162(3):622-34. doi: 10.1016/j.cell.2015.07.015.
  PUBMED    26232228
REFERENCE   2  (bases 1 to 7426)
  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.2015.07"; journalName: "Cell"; date: "2015-07-30- 
            30"; volume: "162"; issue: "3"; pages: "622-34"
COMMENT     Sequence Label: pAAV hSyn FLExFRT mGFP-2A-Synaptophysin-mRuby
FEATURES             Location/Qualifiers
     source          1..7426
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(225..236)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    374..850
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(573..592)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   890..1030
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   890..1019
                     /label=AAV2 ITR
     rep_origin      1105..1560
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1192..1211)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1402..1423
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1577..1596)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1696..1718
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1756..1774)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1842..1946
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1947..2015,2016..2807)
                     /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(2165..2184)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2978..3566
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3467..3486
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   3628..3757
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        3802..4249
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    4327..4374
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             complement(4493..5173)
                     /codon_start=1
                     /product="monomeric variant of the red fluorescent protein 
                     eqFP611 (Kredel et al., 2009)"
                     /label=mRuby
                     /translation="MNSLIKENMRMKVVLEGSVNGHQFKCTGEGEGNPYMGTQTMRIKV
                     IEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGGVITV
                     MQDTSLEDGCLVYHAQVRGVNFPSNGAVMQKKTKGWEPNTEMMYPADGGLRGYTHMALK
                     VDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAVAKFA
                     GLGGG"
     CDS             complement(6125..6178)
                     /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(join(6179..6889,6890..6892,6893..6895))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(6179..6889,6890..6892,6893..6895))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6221..6242)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     6570..6589
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6829..6850
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     protein_bind    complement(6967..7014)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    join(7180..7426,1..342)
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7233..7253)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
ORIGIN
        1 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
       61 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
      121 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctatg
      181 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
      241 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
      301 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgagcgctgc
      361 tcgagagatc tacgggtggc atccctgtga cccctcccca gtgcctctcc tggccctgga
      421 agttgccact ccagtgccca ccagccttgt cctaataaaa ttaagttgca tcattttgtc
      481 tgactaggtg tccttctata atattatggg gtggaggggg gtggtatgga gcaaggggca
      541 agttgggaag acaacctgta gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt
      601 ggcacaatct tggctcactg caatctccgc ctcctgggtt caagcgattc tcctgcctca
      661 gcctcccgag ttgttgggat tccaggcatg catgaccagg ctcagctaat ttttgttttt
      721 ttggtagaga cggggtttca ccatattggc caggctggtc tccaactcct aatctcaggt
      781 gatctaccca ccttggcctc ccaaattgct gggattacag gcgtgaacca ctgctccctt
      841 ccctgtcctt ctgattttgt aggtaaccac gtgcggaccg agcggccgca ggaaccccta
      901 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
      961 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
     1021 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     1081 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     1141 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg
     1201 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     1261 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     1321 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     1381 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     1441 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     1501 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     1561 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     1621 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     1681 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     1741 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     1801 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     1861 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     1921 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     1981 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     2041 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     2101 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     2161 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     2221 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     2281 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     2341 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     2401 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     2461 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     2521 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     2581 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     2641 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     2701 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     2761 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     2821 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     2881 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     2941 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3001 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3061 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3121 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3181 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3241 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3301 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     3361 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     3421 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     3481 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     3541 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     3601 ccttttgctg gccttttgct cacatgtcct gcaggcagct gcgcgctcgc tcgctcactg
     3661 aggccgcccg ggcgtcgggc gacctttggt cgcccggcct cagtgagcga gcgagcgcgc
     3721 agagagggag tggccaactc catcactagg ggttcctgcg gccgcacgcg tgtgtctaga
     3781 ctgcagaggg ccctgcgtat gagtgcaagt gggttttagg accaggatga ggcggggtgg
     3841 gggtgcctac ctgacgaccg accccgaccc actggacaag cacccaaccc ccattcccca
     3901 aattgcgcat cccctatcag agagggggag gggaaacagg atgcggcgag gcgcgtgcgc
     3961 actgccagct tcagcaccgc ggacagtgcc ttcgcccccg cctggcggcg cgcgccaccg
     4021 ccgcctcagc actgaaggcg cgctgacgtc actcgccggt cccccgcaaa ctccccttcc
     4081 cggccacctt ggtcgcgtcc gcgccgccgc cggcccagcc ggaccgcacc acgcgaggcg
     4141 cgagataggg gggcacgggc gcgaccatct gcgctgcggc gccggcgact cagcgctgcc
     4201 tcagtctgcg gtgggcagcg gaggagtcgt gtcgtgcctg agagcgcagt cgagaaggta
     4261 ccggatcctg tgctgtctca tcattttggc aaagaattga tttgataccg cgggcccggg
     4321 ggtaccgaag ttcctattcc gaagttccta ttctctagaa agtataggaa cttcatcaaa
     4381 ataggaagac caatgcttca ccatcgaccc gaattgccaa gcatcaccat cgacagaagt
     4441 tcctattccg aagttcctat tcttcaaaag gtataggaac ttcgtcgact taccctccgc
     4501 ccaggccggc gaacttggcc acggcgtgct cccgctgcac cacgaacatc tcgttgtcgg
     4561 actcttccag cctttccagc cggtggtcca cggcgtggat gccgggcatc ttgatgttgc
     4621 ccacggtttt cttggatcgg taggtggtca cgaagctgca gctcaggtgc cctccgccgt
     4681 ccaccttcag ggccatgtgg gtgtagcctc tcagtccgcc gtcagcgggg tacatcatct
     4741 cggtgttggg ctcccagccc ttggttttct tctgcatcac ggcgccgttg ctgggaaaat
     4801 tcacgcccct gacctgggca tggtacacca ggcagccatc ttccaggctg gtgtcctgca
     4861 tcacggtgat cacgccgccg tcctcgtatc tggtcactct ctcccaggtg aagccctcgg
     4921 ggaagctctg cttgaagaaa tcggggatgc ccttggggta cttgatgaag gtccggctgc
     4981 cgtacatgaa ggatgtggcc aggatgtcga aggcgaaggg cagaggtccg ccctcgatca
     5041 ctttgatccg catggtctgg gtgcccatgt aggggttgcc ctcgccctcg ccggtgcact
     5101 tgaactggtg gccgttcacg ctgccttcca gcaccacctt catccgcatg ttttctttga
     5161 tcaggctgtt catcgtacgg gtggcgaccg gtggatccga catctgattg gagaaggagg
     5221 tgggcgcacc ctgttggccg tagccttgct gcccatagtc gccctgaggc ccgtagccac
     5281 cgccaccgcc gctggctggc tgcccgtaat cgggttgata accaccctga ggcccgtagg
     5341 agtcctgggg cccatagcct ccggggccct gcccatagcc cgcatcgccg taggcatcgc
     5401 caggagctgg ttgcttttct ggggcgcctg gaggtgcgcg catgaatggg gcggcccagc
     5461 ctgtctcctt gaacacgaac cataggttgc caacccagag caccaggttc aggaagccaa
     5521 acaccaccga ggtgttgagt cctgaagtca cagggtccct cagttccttg catgtgtttc
     5581 ctgtctggcg gcacataggc atctccttga taatgttctc tgggtcagtg gccatcttca
     5641 catcggacag gcctttggcc caggcggatg agctaactag ccacatgaaa gcgaacactg
     5701 ctgtggccag gaagtccatc attggccctt tgttgttctc tcggtacttg ttctgcagga
     5761 agatgtaggt ggccagggcc cccatggagt agaggaaggc aaacacagcc acggtgacaa
     5821 agaattcagc cgaggaggag tagtcaccaa ctaggaagat cttggtagtg ccccctttaa
     5881 cgcaggaggg tgcatcaaag tacacttggt gcagcctgaa tgggtactca aattcgactt
     5941 cgatgttgag ggcactctcc gtcttgttgg cacactccac gctcagccga agctctccgg
     6001 tgtagctgcc gcacgtagca aaggcgaaga tggcaaagac ccactgcagc accttcacga
     6061 agccaagggg ctccttgacc acccggaact gacccccagc caccagctga ttcaccacgt
     6121 ccatggggcc ggggttctcc tccacgtcgc cgcaggtcag caggctgccc ctgccctcct
     6181 tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca
     6241 tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga ctgggtgctc aggtagtggt
     6301 tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga
     6361 gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct
     6421 tctgcttgtc ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc
     6481 ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg
     6541 tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga
     6601 tggtgcgctc ctggacgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca
     6661 tgtggtcggg gtagcggctg aagcactgca cgccgtaggt cagggtggtc acgagggtgg
     6721 gccagggcac gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg
     6781 tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca
     6841 gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc accatgatct
     6901 tttggtcctc atcattcttt tcaacctgtt tggttcttct catacagcac agcatggtgg
     6961 cgcgccgaag ttcctatact ttctagagaa taggaacttc ggaataggaa cttccattgg
     7021 gattcttcct attttgatcc aagcatcacc atcgaccctc tagtccagat ctcaccatcg
     7081 acccgaagtt cctatacctt ttgaagaata ggaacttcgg aataggaact tcaagcttat
     7141 atgcggccga tcgatgaatt cgatatcaag cttatcgata atcaacctct ggattacaaa
     7201 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     7261 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     7321 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     7381 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggg
//
