LOCUS       sequence-317821-        6647 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express FLPo and GFP under the control of human synapsin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6647)
  AUTHORS   An JJ, Kinney CE, Tan JW, Liao GY, Kremer EJ, Xu B
  TITLE     TrkB-expressing paraventricular hypothalamic neurons suppress 
            appetite through multiple neurocircuits.
  JOURNAL   Nat Commun. 2020 Apr 7;11(1):1729. doi: 10.1038/s41467-020-15537-w.
  PUBMED    32265438
REFERENCE   2  (bases 1 to 6647)
  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 
            Commun."; date: "2020-04-7"; volume: "11(1):1729. doi"; pages: " 
            10.1038/s41467-020-15537-w"
COMMENT     Sequence Label: pAAV-hsyn-FLPo-T2A-GFP
FEATURES             Location/Qualifiers
     source          1..6647
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   2..131
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    187..235
                     /note="synthetic polyadenylation signal"
     misc_feature    249..340
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     289..308
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        376..823
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      854..863
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(860..865,866..886,887..2155)
                     /codon_start=1
                     /product="nuclear-targeted site-specific recombinase"
                     /label=FLPo
                     /note="FLPo is a mouse codon-optimized fusion between the 
                     SV40 nuclear localization signal and a thermostable version
                     of the Saccharomyces cerevisiae site-specific recombinase 
                     FLP.  Recombination occurs at FRT sequences."
                     /translation="MAPKKKRKVMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAA
                     ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK
                     LIPAWEFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESI
                     WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV
                     IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY
                     QLLKDNLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS
                     DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL
                     KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI"
     CDS             2222..2275
                     /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             join(2276..2278,2279..2992)
                     /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             join(2276..2278,2279..2989)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2318..2339)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2579..2598)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2926..2947
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3005..3593
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3058..3078)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3476..3487)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3626..3643)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3632..3839
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3848..3977
                     /label=AAV2 ITR
     rep_origin      4076..4531
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4163..4182)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4373..4394
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4598..4617)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4717..4739
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4777..4795)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4863..4967
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4968..5036,5037..5828)
                     /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(5186..5205)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5999..6587
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6488..6507
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt
       61 tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac
      121 taggggttcc tgcggccgca cgcgttctag aaatattaag gtacgggagg tacttggagc
      181 ggccgcaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg
      241 atagtactaa catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag
      301 gctgtcccca gtgcaagtgc aggtgccaga acatttctct attctagact cgagctgcag
      361 agggccctgc gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc
      421 ctacctgacg accgaccccg acccactgga caagcaccca acccccattc cccaaattgc
      481 gcatccccta tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc
      541 agcttcagca ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct
      601 cagcactgaa ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca
      661 ccttggtcgc gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat
      721 aggggggcac gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc
      781 tgcggtgggc agcggaggag tcgtgtcgtg cctgagagcg cagtcgaatt caagctgcta
      841 gcaaggatcc gccgccacca tggctcctaa gaagaagagg aaggtgatga gccagttcga
      901 catcctgtgc aagacccccc ccaaggtgct ggtgcggcag ttcgtggaga gattcgagag
      961 gcccagcggc gagaagatcg ccagctgtgc cgccgagctg acctacctgt gctggatgat
     1021 cacccacaac ggcaccgcca tcaagagggc caccttcatg agctacaaca ccatcatcag
     1081 caacagcctg agcttcgaca tcgtgaacaa gagcctgcag ttcaagtaca agacccagaa
     1141 ggccaccatc ctggaggcca gcctgaagaa gctgatcccc gcctgggagt tcaccatcat
     1201 cccttacaac ggccagaagc accagagcga catcaccgac atcgtgtcca gcctgcagct
     1261 gcagttcgag agcagcgagg aggccgacaa gggcaacagc cacagcaaga agatgctgaa
     1321 ggccctgctg tccgagggcg agagcatctg ggagatcacc gagaagatcc tgaacagctt
     1381 cgagtacacc agcaggttca ccaagaccaa gaccctgtac cagttcctgt tcctggccac
     1441 attcatcaac tgcggcaggt tcagcgacat caagaacgtg gaccccaaga gcttcaagct
     1501 ggtgcagaac aagtacctgg gcgtgatcat tcagtgcctg gtgaccgaga ccaagacaag
     1561 cgtgtccagg cacatctact ttttcagcgc cagaggcagg atcgaccccc tggtgtacct
     1621 ggacgagttc ctgaggaaca gcgagcccgt gctgaagaga gtgaacagga ccggcaacag
     1681 cagcagcaac aagcaggagt accagctgct gaaggacaac ctggtgcgca gctacaacaa
     1741 ggccctgaag aagaacgccc cctaccccat cttcgctatc aagaacggcc ctaagagcca
     1801 catcggcagg cacctgatga ccagctttct gagcatgaag ggcctgaccg agctgacaaa
     1861 cgtggtgggc aactggagcg acaagagggc ctccgccgtg gccaggacca cctacaccca
     1921 ccagatcacc gccatccccg accactactt cgccctggtg tccaggtact acgcctacga
     1981 ccccatcagc aaggagatga tcgccctgaa ggacgagacc aaccccatcg aggagtggca
     2041 gcacatcgag cagctgaagg gcagcgccga gggcagcatc agataccccg cctggaacgg
     2101 catcatcagc caggaggtgc tggactacct gagcagctac atcaacaggc ggatcaagct
     2161 taggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaaggaat tcggcagtgg
     2221 agagggcaga ggaagtctgc taacatgcgg tgacgtcgag gagaatcctg gcccagtgag
     2281 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     2341 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     2401 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     2461 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
     2521 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     2581 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     2641 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     2701 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     2761 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     2821 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     2881 cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     2941 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt aaaagcttat
     3001 cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     3061 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     3121 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     3181 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     3241 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     3301 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     3361 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     3421 gctcgcctgt gttgccacct ggattctgcg cgggacgccc ttctgctacg tcccttcggc
     3481 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     3541 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata
     3601 ccgtcgacta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3661 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3721 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3781 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggag
     3841 agatctgagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     3901 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     3961 agcgagcgag cgcgcagaga gggccccccc ccccccccct gcagcctggc gtaatagcga
     4021 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg
     4081 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     4141 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     4201 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     4261 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     4321 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     4381 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     4441 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     4501 aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc
     4561 tgtgcggtat ttcacaccgc atatggtgca ctctcagtac aatctgctct gatgccgcat
     4621 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     4681 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     4741 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat
     4801 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     4861 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     4921 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     4981 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5041 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     5101 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     5161 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     5221 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     5281 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     5341 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     5401 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     5461 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     5521 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     5581 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     5641 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     5701 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     5761 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     5821 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     5881 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     5941 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6001 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6061 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     6121 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     6181 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     6241 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     6301 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     6361 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     6421 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     6481 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6541 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6601 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatg
//
