LOCUS       sequence-349553-        5200 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 5200)
  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 5200)
  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: _pCMV(MinDis)-iCyt_F_SnFR_PM
FEATURES             Location/Qualifiers
     source          1..5200
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        389..592
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     543..563
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     637..656
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        637..655
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     736..795
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     primer_bind     1248..1269
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1372..1393)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             2374..2403
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2629..2646)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2635..2859
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2991..3579)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3071..3090)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3750..4541,4542..4610))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGTRVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4373..4392
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4611..4715)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4742..5197
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4829..4848)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5039..5060
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 cgcgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      421 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      481 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      541 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagaa ctctctggct
      601 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
      661 ccaagcttgg taccgagctc ggatccacta gtaacggccg ccagtgtgct ggaattcggc
      721 ttggggatat ccaccatgga gacagacaca ctcctgctat gggtactgct gctctgggtt
      781 ccaggttcca ctggtgacag atctgcgaac gacaccgtag ttgtgggctc gatcatcttt
      841 acagaaggga ttatcgtcgc aaacatggtg gcagagatga ttgaggcgca tacagacctt
      901 aaggtggttc gcaaactgaa ccttggcggg aatggtgtta actttgaagc cattaaacgc
      961 ggaggtgcga ataatggtat tgacatttac gtggagtaca ctgggcacgg tcttgtggat
     1021 attctggggt tcccggagcc gaacgtctat atcaccgccg acaagcagaa gaacggcatc
     1081 aaggcgaact tcaagatccg ccacaacgtg gaggacggca gcgtgcagct cgccgaccac
     1141 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     1201 agcacccagt ccgtgctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     1261 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gggcggtacc
     1321 ggagggagca tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     1381 ctggacggcg acgtaaacgg ccacaagttc agcgtgcgcg gcgagggcga gggcgatgcc
     1441 accaacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     1501 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
     1561 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
     1621 atcagcttca aggacgacgg cacctacaag acccgcgccg aggtgaagtt cgagggcgac
     1681 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     1741 gggcacaagc tggagtacaa ctttccgccg cccgcgacta ctgatccaga aggtgcatac
     1801 gaaaccgtga agaaggagta caaacgtaaa tggaatattg tatggctcaa accactggga
     1861 ttcaacaata cgtatacgct taccgttaaa gacgaactgg cgaaacagta taaccttaaa
     1921 accttcagtg acttagcgaa aatctcggat aagctgattc tgggtgcaac gatgtggttt
     1981 ttagaagggc ccgatggtta cccaggcctg caaaaactgt acaatttcaa attcaagcac
     2041 accaaaagca tggacatggg tattcgctat accgccattg ataataacga agttcaggta
     2101 attgatgcct gggccactga tggcttgctg gtgagccaca aattaaaaat tctggaggat
     2161 gataaagcgt tcttcccgcc gtattatgct gcccccatca tccgtcagga tgtcttagat
     2221 aagcatcctg aactgaagga cgtgctgaac aaactcgcga atcaaatttc actggaagaa
     2281 atgcagaaac tgaattacaa ggtggacggt gagggtcagg acccagcgaa agtagctaag
     2341 gagtttttga aagagaaagg tttaattacg cgtgaacaaa aactcatctc agaagaggat
     2401 ctgaatgctg tgggccagga cacgcaggag gtcatcgtgg tgccacactc cttgcccttt
     2461 aaggtggtgg tgatctcagc catcctggcc ctggtggtgc tcaccatcat ctcccttatc
     2521 atcctcatca aatacttgca gaagaggcgg gaaaggcgac gccaggagaa cgctaactcc
     2581 ttttgctatg aaaatgaggt agccctgtag gcggccgctc gagatcagcc tcgactgtgc
     2641 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     2701 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     2761 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     2821 acaatagcag gcgtgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     2881 gtggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2941 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3001 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3061 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3121 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3181 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3241 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3301 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3361 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3421 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3481 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3541 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3601 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3661 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3721 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3781 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3841 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3901 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3961 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4021 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4081 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4141 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4201 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4261 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4321 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4381 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4441 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgtacccaac
     4501 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4561 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4621 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4681 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4741 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4801 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4861 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4921 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     4981 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     5041 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     5101 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     5161 aacgcgaatt ttaacaaaat attaacgctt acaatttacg
//
