LOCUS       sequence-410347-        5252 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses the membrane-localized enhanced green fluorescent protein 
            EGFP-CAAX in neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5252)
  AUTHORS   Wang H, Qian T, Zhao Y, Zhuo Y, Wu C, Osakada T, Chen P, Chen Z, Ren
            H, Yan Y, Geng L, Fu S, Mei L, Li G, Wu L, Jiang Y, Qian W, Zhang L,
            Peng W, Xu M, Hu J, Jiang M, Chen L, Tang C, Zhu Y, Lin D, Zhou JN, 
            Li Y
  TITLE     A tool kit of highly selective and sensitive genetically encoded 
            neuropeptide sensors.
  JOURNAL   Science. 2023 Nov 17;382(6672):eabq8173. doi: 
            10.1126/science.abq8173. Epub 2023 Nov 17.
  PUBMED    37972184
REFERENCE   2  (bases 1 to 5252)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science."; 
            date: "2023-11-17"; volume: "382(6672):eabq8173. doi"; pages: " 
            10.1126/science.abq8173. Epub 2023 Nov 17"
COMMENT     Sequence Label: pAAV-hSyn-EGFP-CAAX
FEATURES             Location/Qualifiers
     source          1..5252
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      581..590
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(587..589,590..592,593..1303)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(587..589,590..592,593..1303)
                     /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(632..653)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(893..912)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1240..1261
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1379..1967
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1432..1452)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1850..1861)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    1999..2475
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2198..2217)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   2515..2655
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2515..2644
                     /label=AAV2 ITR
     rep_origin      2730..3185
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2817..2836)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3027..3048
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3202..3221)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3321..3343
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3381..3399)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3467..3571
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3572..3640,3641..4432)
                     /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(3790..3809)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4603..5191
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5092..5111
                     /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 gcgtatgagg cggggtgggg gtgcctacct gacgaccgac
      181 cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag
      241 agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg
      301 acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg
      361 ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc
      421 gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc
      481 gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga
      541 ggagtcgtgt cgtgcctgag agcgcagtcg agaaaccggt gccaccatgg tgagcaaggg
      601 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
      661 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
      721 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
      781 gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt
      841 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
      901 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
      961 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     1021 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     1081 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     1141 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     1201 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     1261 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aaggatggta aaaagaagaa
     1321 aaagaagtca aagacaaagt gtgtaattat gtaagaattc gatatcaagc ttatcgataa
     1381 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     1441 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     1501 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     1561 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     1621 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     1681 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     1741 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     1801 ctatgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     1861 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     1921 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgagc
     1981 gctgctcgag agatctacgg gtggcatccc tgtgacccct ccccagtgcc tctcctggcc
     2041 ctggaagttg ccactccagt gcccaccagc cttgtcctaa taaaattaag ttgcatcatt
     2101 ttgtctgact aggtgtcctt ctataatatt atggggtgga ggggggtggt atggagcaag
     2161 gggcaagttg ggaagacaac ctgtagggcc tgcggggtct attgggaacc aagctggagt
     2221 gcagtggcac aatcttggct cactgcaatc tccgcctcct gggttcaagc gattctcctg
     2281 cctcagcctc ccgagttgtt gggattccag gcatgcatga ccaggctcag ctaatttttg
     2341 tttttttggt agagacgggg tttcaccata ttggccaggc tggtctccaa ctcctaatct
     2401 caggtgatct acccaccttg gcctcccaaa ttgctgggat tacaggcgtg aaccactgct
     2461 cccttccctg tccttctgat tttgtaggta accacgtgcg gaccgagcgg ccgcaggaac
     2521 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     2581 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     2641 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     2701 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     2761 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
     2821 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     2881 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     2941 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     3001 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     3061 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     3121 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     3181 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     3241 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     3301 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     3361 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     3421 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3481 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3541 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     3601 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     3661 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3721 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     3781 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     3841 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     3901 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     3961 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     4021 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     4081 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     4141 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     4201 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     4261 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     4321 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     4381 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     4441 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4501 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4561 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4621 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4681 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4741 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4801 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4861 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4921 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4981 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     5041 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     5101 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     5161 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5221 cctggccttt tgctggcctt ttgctcacat gt
//
