LOCUS       sequence-162265-        5533 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Double floxed EGFP under the control of human synapsin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5533)
  AUTHORS   Bryan Roth
  TITLE     Roth lab DREADDs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5533)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-hSyn-DIO-EGFP
FEATURES             Location/Qualifiers
     source          1..5533
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        175..622
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(660..693)
                     /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(744..777)
                     /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(786..1499,1500..1502,1503..1505))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(789..1499,1500..1502,1503..1505))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(831..852)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1180..1199
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1439..1460
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1502..1511
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1518..1551
                     /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    1602..1635
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    1660..2248
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1713..1733)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2131..2142)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2280..2756
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2479..2498)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   2796..2936
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2796..2925
                     /label=AAV2 ITR
     rep_origin      3011..3466
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3098..3117)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3308..3329
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3483..3502)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3602..3624
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3662..3680)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3748..3852
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3853..3921,3922..4713)
                     /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(4071..4090)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4884..5472
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5373..5392
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtgtgtct agactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc agtcgagaag gtaccggatc ctctagagtc gactccggaa
      661 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
      721 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
      781 gcgccctact tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc
      841 agcaggacca tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga ctgggtgctc
      901 aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag
      961 tggtcggcga gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg
     1021 atgccgttct tctgcttgtc ggccatgata tagacgttgt ggctgttgta gttgtactcc
     1081 agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg
     1141 ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc
     1201 ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca tggcggactt gaagaagtcg
     1261 tgctgcttca tgtggtcggg gtagcggctg aagcactgca cgccgtaggt cagggtggtc
     1321 acgagggtgg gccagggcac gggcagcttg ccggtggtgc agatgaactt cagggtcagc
     1381 ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg
     1441 tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc
     1501 accatggtgg cgctagcata acttcgtata aagtatccta tacgaagtta tttgccttaa
     1561 cccagaaatt atcactgtta ttctttagaa tggtgcaaag aataacttcg tataatgtat
     1621 gctatacgaa gttatgaatt cgatatcaag cttatcgata atcaacctct ggattacaaa
     1681 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     1741 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     1801 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     1861 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     1921 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     1981 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     2041 gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctatgttgc cacctggatt
     2101 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     2161 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     2221 cggatctccc tttgggccgc ctccccgcat cgataccgag cgctgctcga gagatctacg
     2281 ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt gccactccag
     2341 tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac taggtgtcct
     2401 tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt gggaagacaa
     2461 cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca caatcttggc
     2521 tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcagcct cccgagttgt
     2581 tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg tagagacggg
     2641 gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc tacccacctt
     2701 ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct gtccttctga
     2761 ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga tggagttggc
     2821 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     2881 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg
     2941 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag
     3001 caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3061 agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc
     3121 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3181 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     3241 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3301 tttaatagtg gactcttgtt ccaaactgga acaacactca actctatctc gggctattct
     3361 tttgatttat aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa
     3421 caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc
     3481 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     3541 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     3601 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     3661 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     3721 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     3781 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     3841 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     3901 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     3961 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     4021 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     4081 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     4141 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     4201 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     4261 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     4321 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     4381 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     4441 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     4501 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     4561 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     4621 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     4681 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     4741 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     4801 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     4861 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     4921 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     4981 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     5041 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     5101 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     5161 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     5221 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     5281 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     5341 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     5401 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     5461 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     5521 tttgctcaca tgt
//
