LOCUS       sequence-318008-        7836 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EGFP Fluorescent reporter for overexpressed synaptophysin in 
            mammalian cell.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7836)
  AUTHORS   Harwell CC, Parker PR, Gee SM, Okada A, McConnell SK, Kreitzer AC, 
            Kriegstein AR
  TITLE     Sonic hedgehog expression in corticofugal projection neurons directs
            cortical microcircuit formation.
  JOURNAL   Neuron. 2012 Mar 22;73(6):1116-26. doi: 
            10.1016/j.neuron.2012.02.009. Epub 2012 Mar 21.
  PUBMED    22445340
REFERENCE   2  (bases 1 to 7836)
  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.neuron.2012.02.009"; journalName: "Neuron"; date: 
            "2012-03-22- 22"; volume: "73"; issue: "6"; pages: "1116-26"
COMMENT     Sequence Label: CAG Double flox Synaptophysin-EGFP WPRE
FEATURES             Location/Qualifiers
     source          1..7836
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(24..57)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(108..141)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(152..865,866..868))
                     /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(155..865,866..868))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(197..218)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     546..565
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     805..826
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1834..1843
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1881..1914
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    1965..1998
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2030..2618
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2083..2103)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2501..2512)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2650..3126
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2849..2868)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3166..3306
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3166..3295
                     /label=AAV2 ITR
     rep_origin      3381..3836
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3468..3487)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3678..3699
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3853..3872)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3972..3994
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4032..4050)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4118..4222
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4223..4291,4292..5083)
                     /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(4441..4460)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5254..5842
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5743..5762
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        6172..6475
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          6753..7769
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     7693..7715
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     7777..7796
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
ORIGIN
        1 gatcctctag agtcgactcc ggaataactt cgtataggat actttatacg aagttatgca
       61 gaatggtagc tggattgtag ctgctattag caatatgaaa cctcttaata acttcgtata
      121 gcatacatta tacgaagtta tggcgcgccc tttacttgta cagctcgtcc atgccgagag
      181 tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc tcgttggggt
      241 ctttgctcag ggcggactgg gtgctcaggt agtggttgtc gggcagcagc acggggccgt
      301 cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgctgccg tcctcgatgt
      361 tgtggcggat cttgaagttc accttgatgc cgttcttctg cttgtcggcc atgatataga
      421 cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg tcctccttga
      481 agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac ttcacctcgg
      541 cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg acgtagcctt
      601 cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag cggctgaagc
      661 actgcacgcc gtaggtcagg gtggtcacga gggtgggcca gggcacgggc agcttgccgg
      721 tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg ccctcgccgg
      781 acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc
      841 cggtgaacag ctcctcgccc ttgctcacgt ggatcccact gctataataa caataagttt
      901 ttggttctac ccctttcatc tgattggaga aggaggtggg cgcaccctgt tggccatagc
      961 cttgctgccc atagtcgccc tgaggcccgt agccaccgcc accgctggct ggctgcccgt
     1021 aatcgggttg ataaccaccc tgaggcccgt aggagtcctg gggcccatag cctccggggc
     1081 cctgcccgta gcccgcatcg ccgtaggcat cgccaggtgc tggttgcttt tccggggcgc
     1141 ctggaggtgc gcgcatgaat ggggctgccc agcctgtctc cttgaacacg aaccataagt
     1201 tgccaaccca gagcaccagg ttcaggaagc caaacaccac tgaggtgttg agtcctgaag
     1261 tcacagggtc cctcagttcc ttgcatgtgt tccctgtctg gcggcacatg ggcatctcct
     1321 tgataatgtt ctctgggtcc gtggccatct tcacatcgga caggcctttg gcccaggctg
     1381 atgaactaac tagccacatg aaagcgaaca cggctgtagc cagaaagtcc atcataggcc
     1441 ctttgttgtt ctctcggtac ttgttctgca ggaagatgta ggtggccagg gcccccatgg
     1501 agtagaggaa ggcaaacaca gccacggtga caaagaattc agccgacgag gagtagtccc
     1561 caaccaggaa gatcttggta gtgccccctt tgacgcagga gggtgcatca aagtacactt
     1621 ggtgcagcct gaaggggtac tcgaattcaa cttcgatgtt gagggcactc tccgtcttgt
     1681 tggcacactc cacgctcagc cgaagctccc cggtgtagct gccacacgta gcaaaggcga
     1741 agatggcaaa gacccactgc agcaccttca cgaagccaag gggctccttg accacccgga
     1801 actgaccccc agccaccagc tgattcacca cgtccatggt ggcaagggcg aattccagca
     1861 cactggcggc cgttactagc ataacttcgt ataaagtatc ctatacgaag ttatttgcct
     1921 taacccagaa attatcactg ttattcttta gaatggtgca aagaataact tcgtataatg
     1981 tatgctatac gaagttatga attcgatatc ccaaatcaag cttatcgata atcaacctct
     2041 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct
     2101 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
     2161 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
     2221 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
     2281 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
     2341 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     2401 caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctatgttgc
     2461 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
     2521 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
     2581 tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgag cgctgctcga
     2641 gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt
     2701 gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac
     2761 taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt
     2821 gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca
     2881 caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcagcct
     2941 cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg
     3001 tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc
     3061 tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct
     3121 gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga
     3181 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg
     3241 tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc
     3301 tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     3361 tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     3421 ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt
     3481 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     3541 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg
     3601 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3661 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca actctatctc
     3721 gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt taaaaaatga
     3781 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg
     3841 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc
     3901 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     3961 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     4021 gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     4081 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     4141 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     4201 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     4261 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     4321 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     4381 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     4441 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     4501 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     4561 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     4621 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     4681 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     4741 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     4801 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     4861 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     4921 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     4981 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     5041 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     5101 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     5161 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     5221 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5281 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5341 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5401 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5461 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     5521 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     5581 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     5641 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5701 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5761 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5821 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5881 ttgctggcct tttgctcaca tgtcctgcag gcagctgcgc gctcgctcgc tcactgaggc
     5941 cgggcgacct ttggtctgag gccgggcgac ctttggtcgc ccggcctcag tgagcgagcg
     6001 agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctgcggcc gcacgcgtaa
     6061 cattatggcc ttaggtcact tcatctccat ggggttcttc ttctgatttt ctagttatta
     6121 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
     6181 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
     6241 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
     6301 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
     6361 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
     6421 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtcga
     6481 ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac ccccaatttt
     6541 gtatttattt attttttaat tattttgtgc agcgatgggg gcgggggggg ggggggggcg
     6601 cgccgggggg gggggggggg gggggggggg gggggggggg ggggcggaga ggtgcggcgg
     6661 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc
     6721 ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg
     6781 tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc
     6841 cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagcgc ttggtttaat
     6901 gacggcttgt ttcttttctg tggctgcgtg aaagccttga ggggctccgg gagggccctt
     6961 tgtgcggggg gagcggctcg gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt
     7021 gcggctccgc gctgcccggc ggctgtgagc gctgcgggcg cggcgcgggg ctttgtgcgc
     7081 tccgcagtgt gcgcgagggg agcgcggccg ggggcggtgc cccgcggtgc ggggggggct
     7141 gcgaggggaa caaaggctgc gtgcggggtg tgtgcgtggg ggggtgagca gggggtgtgg
     7201 gcgcgtcggt cgggctgcaa ccccccctgc acccccctcc ccgagttgct gagcacggcc
     7261 cggcttcggg tgcggggctc cgtacggggc gtggcgcggg gctcgccgtg ccgggcgggg
     7321 ggtggcggca ggtgggggtg ccgggcgggg cggggccgcc tcgggccggg gagggctcgg
     7381 gggaggggcg cggcggcccc cggagcgccg gcggctgtcg aggcgcggcg agccgcagcc
     7441 attgcctttt atggtaatcg tgcgagaggg cgcagggact tcctttgtcc caaatctgtg
     7501 cggagccgaa atctgggagg cgccgccgca ccccctctag cgggcgcggg gcgaagcggt
     7561 gcggcgccgg caggaaggaa atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc
     7621 cccttctccc tctccagcct cggggctgtc cgcgggggga cggctgcctt cgggggggac
     7681 ggggcagggc ggggttcggc ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc
     7741 atgttcatgc cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt
     7801 ctcatcattt tggcaaagaa ttcggtaccg agctcg
//
