LOCUS       sequence-415596-        7763 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Neuropeptide degradation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7763)
  AUTHORS   Kim D-I, Park S, Ye M, Chen JY, Jhang J, Hunker AC, Zweifel LS, 
            Palmiter RD, Han S
  TITLE     Novel genetically encoded tools for imaging or silencing 
            neuropeptide release from presynaptic terminals in vivo
  JOURNAL   bioRxiv 2023.01.19.524797
REFERENCE   2  (bases 1 to 7763)
  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: "bioRxiv 
            2023.01.19.524797"
COMMENT     Sequence Label: pAAV-hSyn1-FLEX-NEPLDCV-P2A-mRUBY3-WPRE
FEATURES             Location/Qualifiers
     source          1..7763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          533..665
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     648..667
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    678..711
                     /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    774..807
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(830..1537)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="VSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTMR
                     IKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGGV
                     VTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTDI
                     ALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAVA
                     KYSNLGGGMDELYK"
     CDS             complement(1538..1594)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    complement(3802..3835)
                     /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(3906..3939)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3981..4569
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4034..4054)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4452..4463)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4647..4768
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4658..4677)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4712..4731
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4930..5059
                     /label=AAV2 ITR
     rep_origin      complement(5296..5884)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5376..5395)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6055..6846,6847..6915))
                     /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     6678..6697
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6916..7020)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7046..7502)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7183..7204)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7395..7414
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   7574..7703
                     /label=AAV2 ITR
ORIGIN
        1 gagctccagt gtgctggaat tcgccctttt cagtatttaa attgacgacc gaccccgacc
       61 cactggacaa gcacccaacc cccattcccc aaattgcgca tcccctatca gagaggggga
      121 ggggaaacag gatgcggcga ggcgcgtgcg cactgccagc ttcagcaccg cggacagtgc
      181 cttcgccccc gcctggcggc gcgcgccacc gccgcctcag cactgaaggc gcgctgacgt
      241 cactcgccgg tcccccgcaa actccccttc ccggccacct tggtcgcgtc cgcgccgccg
      301 ccggcccagc cggaccgcac cacgcgaggc gcgagatagg ggggcacggg cgcgaccatc
      361 tgcgctgcgg cgccggcgac tcagcgctgc ctcagtctgc ggtgggcagc ggaggagtcg
      421 tgtcgtgcct gagagcgcag ctgtgctcct gggcaccgcg cagtccgccc ccgcggctcc
      481 tggccagacc acccctagga ccccctgccc caagtcgcag ccgttggatc aggtaagtat
      541 caaggttaca agacaggttt aaggagacca atagaaactg ggcttgtcga gacagagaag
      601 actcttgcgt ttctgatagg cacctattgg tcttactgac atccactttg cctttctctc
      661 cacaggtgat gatccccata acttcgtata aagtatccta tacgaagtta tatcaaaata
      721 ggaagaccaa tgcttcacca tcgacccgaa ttgccaagca tcaccatcga cacataactt
      781 cgtataatgt atgctatacg aagttatact aggagctcgg cgcgccttac ttgtacagct
      841 cgtccatgcc accaccaagg ttgctgtatt tggcaactgc cacttctctt tgcactacgt
      901 aggtttcatt gtcactctcc tcgatccttt ccaggcggtg atcaacggca tggacaccgg
      961 gcatcttgat gttcccgacg gtcttttttg acctgtaagt tgtcacgaag ttgcagtgca
     1021 gatggccacc accatcaact ttcagtgcga tgtcagtgta tcctctcaga ccaccatctg
     1081 ctggatacat catctctgta ttaggctccc aacccttggt cttcttctgc atcacgggac
     1141 cattggaggg aaagtttacc cctctgacct tgacgttgta gacgagctcg ccatcctcaa
     1201 ggctggtgtc ctgcgtgacg gtgacgactc caccatcttc gtatctcgta actctttccc
     1261 aagtaaaacc ctcaggaaag gactgtttaa agaaatcagg gatgtcggcc gggtacttga
     1321 taaaggtacg gctgccatac atgaacgacg tggcaagaat gtcaaaggca aatggcaggg
     1381 gtcctccctc gatgactttg atcctcatgg tttgcactcc ctcgtacggt ctgccttctc
     1441 cttcacctgt gcatttgaat tggtggccgt tgaccgaacc ttccatgacc accttcatac
     1501 gcatattttc cttgatcagc tcttcgccct tagacacagg accggggttt tcttccacgt
     1561 ctcctgcttg ctttaacaga gagaagttcg tggctccgga tccccaaacc cggcacttct
     1621 tttctggatt catgtatgaa ttcttgcggc agtgaaaggc ttctgaaaac tctgcagagt
     1681 tctgcaaagt cccaataatc ctgaaattgc ctggactgtg cacatctgtt ttaatggagt
     1741 taaccgcata ctctggccta taggttccac accacacctg tgcaaagttc aagaaaaata
     1801 gttgtttgtg atttaggtca agtccaggaa gtaatttttc ttcgccattc tttttaatat
     1861 aattctgata ggctctgtat gcttgaccaa gacctccatt atcagcaatg ttttctccca
     1921 gtgtattaat tccattaagg tgctgtccac ctgccaggtc ccaggaaaag tttccatact
     1981 gatacaccat gcactgggat tgctccttaa agttacttgc agactgttga gtccaccagt
     2041 caacgaggtc tccatctttg ttaaagtttc tgccattgtc atcgaagcca tgggtgattt
     2101 cgtgtcctat gaccatgccg atgcccccat agttcaatga gttggactgc tgggcactaa
     2161 agaagggggg ctgcagaatg ccggctggga agactatctg atttcttcct gaagagtaaa
     2221 atgcattgac tacagctgct ccacttatcc actcatcttt gtccaccttt tctcggagct
     2281 tcttcagttg tttactttgg ctgaatttca aattttgaat tatgttctcg aagtattcat
     2341 cttctttgta gttcaactcg aggtactcat tattcagttt gttatcattt gaaacaatgt
     2401 catcaggata gccgatcctt tctttaattg ctaaggcctt ttcttcagct ctcttttttg
     2461 tctcggcatc catccaagtg aggtcatcta aagtctgaat aaaaacttct cggatctgtg
     2521 caatcaaatc ctcgaccaca tgtttactct ctccagcaaa tgctgcttcc acataaagcc
     2581 tccccacagc attttccata ttcccattga catagtttgc acaacgtctc caagttgctg
     2641 tttctgaggt tgtaccataa agggccttgc ggaaagcatt tctggactcc ttgtaggttc
     2701 ggctgaggct gcttacaaga tccattatga atctccagga cattaaattt tgaagatctc
     2761 tggcagaata tttggtaaga atgggcttaa gtttggttaa atattctgga gcataaacaa
     2821 ccacatcttc ctcatttgta atactaatat tcacagttga catgatttca tttgtgaaat
     2881 tcaaccagct gaatggcttc ccattgatct ctagtgaaaa gttattttgg atctgggcca
     2941 atgtcatctt gttatacaga agcattggat catttcgatc ttcaggttta gccgtagcat
     3001 tggcaatttc tttttccaat tccataactt tattcatttc caaagcaagc tggttttcat
     3061 cgatgggcaa tctttcttcc tgacgaatca atctggccac agaaatcata aaatccacat
     3121 atgctgtaca agcctcttta tagattccag tgcattcata gtaatctcta gaagggaggc
     3181 caagtcgagg ttggtcaata tgaattacat gattcacaga attcttatca tcagtgccaa
     3241 caaacaaatt aataaggact tttttcccat atttagaatt cagttgtgca atagcttttt
     3301 cagctgtcca agaagcacca tatttttgct cccagttttc tgttgctact ggccacccat
     3361 atatgtctgg taacagtttg agtagaggtt ctccacctct gctatcaata gcagattcat
     3421 ttatacaaga cctgtacaat gcttttgctt tctgcactgc tactatatct tcagttttgg
     3481 gttcttgaag gacatctttc aaaacgactt ctagttcatc tcttaaaatg tcaaagttgc
     3541 cgtaacggga gctggtctcg ggaatgacat tacgtttcaa ccagcctccg caagcatatt
     3601 tgaaaaagtc tgtacaaggc tcagtggtgg catccatgtt ttggatcagt cgagcagctg
     3661 attttatgca gtctgatgac ttgcaaatgg gcttgccacg cacttccatg gaggcctgaa
     3721 gcagcaaggc cagcaacagg gcccccgagc ggctgcagca cgatctcggc atggtggggg
     3781 ctagcggtac cgaattctag tataacttcg tataggatac tttatacgaa gttatcattg
     3841 ggattcttcc tattttgatc caagcatcac catcgaccct ctagtccaga tctcaccatc
     3901 gacccataac ttcgtatagc atacattata cgaagttatg tccctcgaag aggttcgaat
     3961 tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg
     4021 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     4081 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     4141 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     4201 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     4261 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     4321 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     4381 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     4441 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     4501 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     4561 cctccccgca tcgataccgt cgacccgggc ggccgctcga gcatgcatct agagtcgacc
     4621 cgggcggccg cttcgagcag acatgataag atacattgat gagtttggac aaaccacaac
     4681 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     4741 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     4801 ggttcagggg gagatgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtaa
     4861 aatcgataag gatcttccta gagcatggct acgtagataa gtagcatggc gggttaatca
     4921 ttaactacaa ggaaccccta gtgatggagt tggccactcc ctctctgcgc gctcgctcgc
     4981 tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag
     5041 tgagcgagcg agcgcgcagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     5101 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     5161 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     5221 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5281 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5341 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5401 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5461 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5521 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5581 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5641 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5701 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5761 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5821 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5881 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5941 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     6001 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     6061 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     6121 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     6181 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     6241 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6301 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6361 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6421 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6481 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6541 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6601 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6661 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6721 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6781 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6841 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6901 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6961 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     7021 cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg
     7081 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     7141 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     7201 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     7261 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     7321 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     7381 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     7441 gcggtcacgc tgcgcgtaan ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     7501 gtcccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     7561 gctattacgc cagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg
     7621 tcgggcgacc tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc
     7681 caactccatc actaggggtt ccttgtagtt aatgattaac ccgccatgct acttatctac
     7741 gtagccatgc tctaggaaga tcc
//
