LOCUS       sequence-454869-        7126 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Gluc-NHP.RMA and EGFP under the hSyn promoter. 
            Gluc-NHP.RMA for monitoring neuronal transduction..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7126)
  AUTHORS   Lee S, Romac M, Watanabe S, Chernov M, Li H, Raisley E, Szablowski 
            HO, Costa VD
  TITLE     Synthetic Serum Markers Enable Noninvasive Monitoring of Gene 
            Expression in Primate Brains
  JOURNAL   bioRxiv 2025.06.01.657212; doi: 10.1101/2025.06.01.657212
REFERENCE   2  (bases 1 to 7126)
  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 
            2025.06.01.657212; doi: 10.1101/2025.06.01.657212"
COMMENT     Sequence Label: pAAV-hSyn-Gluc-NHP.RMA-IRES-EGFP
FEATURES             Location/Qualifiers
     source          1..7126
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   5..134
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        179..626
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      667..676
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             673..1227
                     /codon_start=1
                     /product="enhanced variant of secreted Gaussia luciferase"
                     /label=Gaussia-Dura Luc
                     /note="human codon-optimized"
                     /translation="MGVKVLFALICIAVAEAKPTENNEDFNIVAVASNFATTDLDADRG
                     KLPGEKLPLEVLKELEANARKAGCTRGCLICLSHIKCTPKMKKFIPGRCHTYEGDKESA
                     QGGIGEAIDDIPEIPGFKDLEPIEQFIAQVDLCVDCTTGCLKGLANVQCSDLLKKWLPQ
                     RCATFASKIQGQVDKIKGAGDD"
     CDS             673..1227
                     /codon_start=1
                     /product="secreted Gaussia luciferase"
                     /label=hGLuc
                     /note="human codon-optimized"
                     /translation="MGVKVLFALICIAVAEAKPTENNEDFNIVAVASNFATTDLDADRG
                     KLPGEKLPLEVLKELEANARKAGCTRGCLICLSHIKCTPKMKKFIPGRCHTYEGDKESA
                     QGGIGEAIDDIPEIPGFKDLEPIEQFIAQVDLCVDCTTGCLKGLANVQCSDLLKKWLPQ
                     RCATFASKIQGQVDKIKGAGDD"
     misc_feature    join(1893..2467,2468..2470,2471..2479)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2060..2077)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2287..2306
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2480..2482,2483..2485,2486..3196)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2480..2482,2483..2485,2486..3196)
                     /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(2525..2546)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2786..2805)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3133..3154
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3206..3226
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    3257..3845
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3310..3330)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3728..3739)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3877..4353
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4076..4095)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4393..4533
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4393..4522
                     /label=AAV2 ITR
     rep_origin      4608..5063
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4695..4714)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4905..4926
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5080..5099)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5199..5221
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5259..5277)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5345..5449
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5450..5518,5519..6310)
                     /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(5668..5687)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6481..7069
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6970..6989
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgtcctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc gtcgggcgac
       61 ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg ccaactccat
      121 cactaggggt tcctgcggcc gcacgcgtgt gtctagactg cagagggccc tgcgtatgag
      181 tgcaagtggg ttttaggacc aggatgaggc ggggtggggg tgcctacctg acgaccgacc
      241 ccgacccact ggacaagcac ccaaccccca ttccccaaat tgcgcatccc ctatcagaga
      301 gggggagggg aaacaggatg cggcgaggcg cgtgcgcact gccagcttca gcaccgcgga
      361 cagtgccttc gcccccgcct ggcggcgcgc gccaccgccg cctcagcact gaaggcgcgc
      421 tgacgtcact cgccggtccc ccgcaaactc cccttcccgg ccaccttggt cgcgtccgcg
      481 ccgccgccgg cccagccgga ccgcaccacg cgaggcgcga gatagggggg cacgggcgcg
      541 accatctgcg ctgcggcgcc ggcgactcag cgctgcctca gtctgcggtg ggcagcggag
      601 gagtcgtgtc gtgcctgaga gcgcagtcga gaaggtaccg gatcctctag agtcgagctc
      661 aagcttgcca ccatgggagt caaagttctg tttgccctga tctgcatcgc tgtggccgag
      721 gccaagccca ccgagaacaa cgaagacttc aacatcgtgg ccgtggccag caacttcgcg
      781 accacggatc tcgatgctga ccgcgggaag ttgcccggcg agaagctgcc gctggaggtg
      841 ctcaaagagt tggaagccaa tgcccggaaa gctggctgca ccaggggctg tctgatctgc
      901 ctgtcccaca tcaagtgcac gcccaagatg aagaagttca tcccaggacg ctgccacacc
      961 tacgaaggcg acaaagagtc cgcacagggc ggcataggcg aggcgatcga cgacattcct
     1021 gagattcctg ggttcaagga cttggagccc atagagcagt tcatcgcaca ggtcgatctg
     1081 tgtgtggact gcacaactgg ctgcctcaaa gggcttgcca acgtgcagtg ttctgacctg
     1141 ctcaagaagt ggctgccgca acgctgcgcg acctttgcca gcaagatcca gggccaggtg
     1201 gacaagatca agggggccgg tgatgacggc tccctcctgg ggggaccgtc agtcttcctc
     1261 ttccccccaa aacccaagga caccctcatg atctcccgga cccctgaggt cacatgcgtg
     1321 gtggtagacg tgagccagga agaccccgat gtcaagttca actggtacgt aaacggcgcg
     1381 gaggtgcatc atgcccagac gaagccacgg gagacgcagt acaacagcac atatcgtgtg
     1441 gtcagcgtcc tcaccgtcac gcaccaggac tggctgaacg gcaaggagta cacgtgcaag
     1501 gtctccaaca aagccctccc ggcccccatc cagaaaacca tctccaaaga caaagggcag
     1561 ccccgagagc ctcaggtgta caccctgccc ccgtcccggg aggagctgac caagaaccag
     1621 gtcagcctga cctgcctggt caaaggcttc taccccagcg acatcgtcgt ggagtgggag
     1681 agcagcgggc agccggagaa cacctacaag accacgccgc ccgtgctgga ctccgacggc
     1741 tcctacttcc tctacagcaa gctcaccgtg gacaagagca ggtggcagca ggggaacgtc
     1801 ttctcatgct ccgtgatgca tgaggctctg cacaaccact acacccagaa gagcctctcc
     1861 gtgtctccgg gtaaatgatt ctaggggatc cgcccctctc cctccccccc ccctaacgtt
     1921 actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt attttccacc
     1981 atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt cttgacgagc
     2041 attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag
     2101 gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg
     2161 cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg tgtataagat
     2221 acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt tgtggaaaga
     2281 gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca gaaggtaccc
     2341 cattgtatgg gatctgatct ggggcctcgg tacacatgct ttacatgtgt ttagtcgagg
     2401 ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg aaaaacacga
     2461 tgataatatg gccacaacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     2521 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     2581 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
     2641 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
     2701 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     2761 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     2821 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     2881 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     2941 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     3001 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     3061 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
     3121 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     3181 ggacgagctg tacaagggtg gttctcccaa gaagaagagg aaagtctaag tttaattcga
     3241 tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat
     3301 tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca
     3361 tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc
     3421 tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc
     3481 tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt
     3541 cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg
     3601 gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc
     3661 ctttccttgg ctgctcgcct atgttgccac ctggattctg cgcgggacgt ccttctgcta
     3721 cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg
     3781 gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc
     3841 cccgcatcga taccgagcgc tgctcgagag atctacgggt ggcatccctg tgacccctcc
     3901 ccagtgcctc tcctggccct ggaagttgcc actccagtgc ccaccagcct tgtcctaata
     3961 aaattaagtt gcatcatttt gtctgactag gtgtccttct ataatattat ggggtggagg
     4021 ggggtggtat ggagcaaggg gcaagttggg aagacaacct gtagggcctg cggggtctat
     4081 tgggaaccaa gctggagtgc agtggcacaa tcttggctca ctgcaatctc cgcctcctgg
     4141 gttcaagcga ttctcctgcc tcagcctccc gagttgttgg gattccaggc atgcatgacc
     4201 aggctcagct aatttttgtt tttttggtag agacggggtt tcaccatatt ggccaggctg
     4261 gtctccaact cctaatctca ggtgatctac ccaccttggc ctcccaaatt gctgggatta
     4321 caggcgtgaa ccactgctcc cttccctgtc cttctgattt tgtaggtaac cacgtgcgga
     4381 ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct
     4441 cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct
     4501 cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta
     4561 cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag
     4621 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     4681 cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4741 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4801 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     4861 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4921 aactggaaca acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc
     4981 gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     5041 caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc
     5101 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     5161 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     5221 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     5281 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     5341 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5401 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     5461 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5521 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     5581 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5641 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     5701 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5761 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5821 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5881 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5941 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     6001 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     6061 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     6121 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6181 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6241 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6301 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6361 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6421 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     6481 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     6541 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6601 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     6661 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6721 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6781 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6841 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6901 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6961 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     7021 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     7081 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcac
//
