LOCUS       sequence-349856-        6136 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Develop a family of genetically encoded fluorescent biosensors for 
            nicotinic agonists, termed iDrugSnFRs, thus enabling optical 
            subcellular pharmacokinetics for nicotinic agonists.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6136)
  AUTHORS   Nichols AL, Blumenfeld Z, Fan C, Luebbert L, Blom AE, Cohen BN, 
            Marvin JS, Borden PM, Kim CH, Muthusamy AK, Shivange AV, Knox HJ, 
            Campello HR, Wang JH, Dougherty DA, Looger LL, Gallagher T, Rees DC,
            Lester HA
  TITLE     Fluorescence activation mechanism and imaging of drug permeation 
            with new sensors for smoking-cessation ligands.
  JOURNAL   eLife. 2022 Jan 4;11. pii: 74648. doi: 10.7554/eLife.74648.
  PUBMED    34982029
REFERENCE   2  (bases 1 to 6136)
  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: "eLife."; 
            date: "2022-01-4"; volume: "11. pii: 74648. doi"; pages: " 
            10.7554/eLife.74648"
COMMENT     Sequence Label: pAAV9-hSyn-iCyt_F_SnFR_PM
FEATURES             Location/Qualifiers
     source          1..6136
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   76..205
                     /label=AAV2 ITR
     promoter        287..734
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     sig_peptide     761..820
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     primer_bind     1273..1294
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1397..1418)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             2399..2428
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     misc_feature    2649..3237
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2702..2722)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3120..3131)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3240..3256)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3241..3257)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    3286..3407
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3297..3316)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3351..3370
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3569..3698
                     /label=AAV2 ITR
     rep_origin      complement(3935..4523)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4015..4034)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4694..5485,5486..5554))
                     /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     5317..5336
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5555..5659)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(join(5685..6136,1..4))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5822..5843)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6034..6053
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcgtcccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct
       61 tcgctattac gccagctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg
      121 cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg
      181 gccaactcca tcactagggg ttccttgtag ttaatgatta acccgccatg ctacttatct
      241 acgtagccat gctctaggaa gatctctgca gagggccctg cgtatgagtg caagtgggtt
      301 ttaggaccag gatgaggcgg ggtgggggtg cctacctgac gaccgacccc gacccactgg
      361 acaagcaccc aacccccatt ccccaaattg cgcatcccct atcagagagg gggaggggaa
      421 acaggatgcg gcgaggcgcg tgcgcactgc cagcttcagc accgcggaca gtgccttcgc
      481 ccccgcctgg cggcgcgcgc caccgccgcc tcagcactga aggcgcgctg acgtcactcg
      541 ccggtccccc gcaaactccc cttcccggcc accttggtcg cgtccgcgcc gccgccggcc
      601 cagccggacc gcaccacgcg aggcgcgaga taggggggca cgggcgcgac catctgcgct
      661 gcggcgccgg cgactcagcg ctgcctcagt ctgcggtggg cagcggagga gtcgtgtcgt
      721 gcctgagagc gcagtcgaat tcggcttggg gatatccacc atggagacag acacactcct
      781 gctatgggta ctgctgctct gggttccagg ttccactggt gacagatctg cgaacgacac
      841 cgtagttgtg ggctcgatca tctttacaga agggattatc gtcgcaaaca tggtggcaga
      901 gatgattgag gcgcatacag accttaaggt ggttcgcaaa ctgaaccttg gcgggaatgg
      961 tgttaacttt gaagccatta aacgcggagg tgcgaataat ggtattgaca tttacgtgga
     1021 gtacactggg cacggtcttg tggatattct ggggttcccg gagccgaacg tctatatcac
     1081 cgccgacaag cagaagaacg gcatcaaggc gaacttcaag atccgccaca acgtggagga
     1141 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     1201 gctgctgccc gacaaccact acctgagcac ccagtccgtg ctgagcaaag accccaacga
     1261 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     1321 ggacgagctg tacaagggcg gtaccggagg gagcatgagc aagggcgagg agctgttcac
     1381 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     1441 gcgcggcgag ggcgagggcg atgccaccaa cggcaagctg accctgaagt tcatctgcac
     1501 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
     1561 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     1621 cgaaggctac gtccaggagc gcaccatcag cttcaaggac gacggcacct acaagacccg
     1681 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     1741 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactttc cgccgcccgc
     1801 gactactgat ccagaaggtg catacgaaac cgtgaagaag gagtacaaac gtaaatggaa
     1861 tattgtatgg ctcaaaccac tgggattcaa caatacgtat acgcttaccg ttaaagacga
     1921 actggcgaaa cagtataacc ttaaaacctt cagtgactta gcgaaaatct cggataagct
     1981 gattctgggt gcaacgatgt ggtttttaga agggcccgat ggttacccag gcctgcaaaa
     2041 actgtacaat ttcaaattca agcacaccaa aagcatggac atgggtattc gctataccgc
     2101 cattgataat aacgaagttc aggtaattga tgcctgggcc actgatggct tgctggtgag
     2161 ccacaaatta aaaattctgg aggatgataa agcgttcttc ccgccgtatt atgctgcccc
     2221 catcatccgt caggatgtct tagataagca tcctgaactg aaggacgtgc tgaacaaact
     2281 cgcgaatcaa atttcactgg aagaaatgca gaaactgaat tacaaggtgg acggtgaggg
     2341 tcaggaccca gcgaaagtag ctaaggagtt tttgaaagag aaaggtttaa ttacgcgtga
     2401 acaaaaactc atctcagaag aggatctgaa tgctgtgggc caggacacgc aggaggtcat
     2461 cgtggtgcca cactccttgc cctttaaggt ggtggtgatc tcagccatcc tggccctggt
     2521 ggtgctcacc atcatctccc ttatcatcct catcaaatac ttgcagaaga ggcgggaaag
     2581 gcgacgccag gagaacgcta actccttttg ctatgaaaat gaggtagccc tgtaggcggc
     2641 ctatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     2701 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     2761 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     2821 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     2881 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     2941 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     3001 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
     3061 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     3121 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     3181 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
     3241 gataccgtcg acctcgaccc gggcggccgc ttcgagcaga catgataaga tacattgatg
     3301 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     3361 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     3421 gcattcattt tatgtttcag gttcaggggg agatgtggga ggttttttaa agcaagtaaa
     3481 acctctacaa atgtggtaaa atcgataagg atcttcctag agcatggcta cgtagataag
     3541 tagcatggcg ggttaatcat taactacaag gaacccctag tgatggagtt ggccactccc
     3601 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     3661 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcattaatga atcggccaac
     3721 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     3781 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3841 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     3901 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3961 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4021 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4081 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4141 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4201 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4261 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4321 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     4381 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4441 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4501 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4561 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     4621 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     4681 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     4741 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     4801 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     4861 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     4921 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     4981 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     5041 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     5101 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     5161 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     5221 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     5281 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     5341 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     5401 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     5461 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     5521 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     5581 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     5641 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg taagcgttaa
     5701 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     5761 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     5821 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     5881 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     5941 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     6001 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     6061 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     6121 tgcgccgcta cagggc
//
