LOCUS       sequence-348679-        4444 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 4444)
  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 4444)
  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: iCytSnFR
FEATURES             Location/Qualifiers
     source          1..4444
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     55..74
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        55..73
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             105..127
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             138..155
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             159..185
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     650..671
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(774..795)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             1782..1811
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1880..1898)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1894..1941
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2038..2493
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2125..2144)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2335..2356
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2519..2623
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2624..2692,2693..3484)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRFDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2842..2861)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3655..4243
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4144..4163
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4397..4414
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cctctccccg cgcgttggcc gattcattaa tgcaggatct cgatcccgcg aaattaatac
       61 gactcactat agggagacca caacggtttc cctctagaaa taattttgtt taactttaag
      121 aaggagatat acatatgcat catcatcatc atcatggtta tccctatgat gttccagatt
      181 atgctggggc ccagccggcc agatctgcga acgacaccgt agttgtgggc tcgatcatct
      241 ttacagaagg gattatcgtc gcaaacatgg tggcagagat gattgaggcg catacagacc
      301 ttaaggtggt tcgcaaactg aaccttggcg ggtacaacgt taactttgaa gccattaaac
      361 gcggaggtgc gaataatggt attgacattt acgtggagta cactgggcac ggtcttgtgg
      421 atattctggg gttcccggag ccgaacgtct atatcaccgc cgacaagcag aagaacggca
      481 tcaaggcgaa cttcaagatc cgccacaacg tggaggacgg cagcgtgcag ctcgccgacc
      541 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
      601 tgagcaccca gtccgtgctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
      661 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagggcggta
      721 ccggagggag catgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
      781 agctggacgg cgacgtaaac ggccacaagt tcagcgtgcg cggcgagggc gagggcgatg
      841 ccaccaacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
      901 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
      961 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     1021 ccatcagctt caaggacgac ggcacctaca agacccgcgc cgaggtgaag ttcgagggcg
     1081 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1141 tggggcacaa gctggagtac aactttccgc cgcccgcgac tactgatcca gaaggtgcat
     1201 acgaaaccgt gaagaaggag tacaaacgta aatggaatat tgtatggctc aaaccactgg
     1261 gattcaacaa tacgtatacg cttaccgtta aagacgaact ggcgaaacag tataacctta
     1321 aaaccttcag tgacttagcg aaaatctcgg ataagctgat tctgggtgca acgatgtggt
     1381 ttttagaagg gcccgatggt tacccaggcc tgcaaaaact gtacaatttc aaattcaagc
     1441 acaccaaaag catggacatg ggtattcgct ataccgccat tgataataac gaagttcagg
     1501 taattgatgc ctgggccact gatggcttgc tggtgagcca caaattaaaa attctggagg
     1561 atgataaagc gttcttcccg ccgtattatg ctgcccccat catccgtcag gatgtcttag
     1621 ataagcatcc tgaactgaag gacgtgctga acaaactcgc gaatcaaatt tcactggaag
     1681 aaatgcagaa actgaattac aaggtggacg gtgagggtca ggacccagcg aaagtagcta
     1741 aggagttttt gaaagagaaa ggtttaattc tgcaggtcga cgaacaaaaa ctcatctcag
     1801 aagaggatct gaattaagaa ttcgaagctt gatccggctg ctaacaaagc ccgaaaggaa
     1861 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     1921 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggatctgg cgtaatagcg
     1981 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     2041 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2101 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     2161 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     2221 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     2281 cgccctgata gacggttttt cgccctttga cgttggagtc catgttcttt aatagtggac
     2341 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     2401 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     2461 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     2521 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     2581 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     2641 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     2701 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     2761 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     2821 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     2881 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     2941 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     3001 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     3061 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc tttgatcgtt gggaaccgga
     3121 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     3181 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     3241 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     3301 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     3361 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     3421 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     3481 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     3541 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     3601 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3661 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3721 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3781 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     3841 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3901 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3961 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4021 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4081 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4141 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4201 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4261 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4321 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4381 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     4441 accg
//
