LOCUS       sequence-358318-        6654 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 6654)
  AUTHORS   Nichols AL, Blumenfeld Z, Luebbert L, Knox HJ, Muthusamy AK, Marvin 
            JS, Kim CH, Grant SN, Walton DP, Cohen BN, Hammar R, Looger L, 
            Artursson P, Dougherty DA, Lester HA
  TITLE     Selective Serotonin Reuptake Inhibitors within Cells: Temporal 
            Resolution in Cytoplasm, Endoplasmic Reticulum, and Membrane.
  JOURNAL   J Neurosci. 2023 Mar 2:JN-RM-1519-22. doi: 
            10.1523/JNEUROSCI.1519-22.2022.
  PUBMED    36868853
REFERENCE   2  (bases 1 to 6654)
  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: "J Neurosci.
            2023 Mar 2:JN-RM-1519-22. doi: 10.1523/JNEUROSCI.1519-22.2022."
COMMENT     Sequence Label: pCMV(MinDis)-iFluoxSnFR_ER
FEATURES             Location/Qualifiers
     source          1..6654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        388..591
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     542..562
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     636..655
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        636..654
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1235..1256
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1359..1380)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2403..2420)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2409..2633
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2765..3353)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2845..2864)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3682..3729
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3754..3773
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3961..4755)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASKPDGEDLAVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4072..4091)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4682..4701
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(4854..4873)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     enhancer        4952..5143
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5100..5120
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(5203..5994,5995..6063))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSTQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5826..5845
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6064..6168)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6195..6650
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6282..6301)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6492..6513
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcgcgttgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
      421 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
      481 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
      541 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagaac tctctggcta
      601 actagagaac ccactgctta ctggcttatc gaaattaata cgactcacta tagggagacc
      661 caagcttggt accgaactcg gatccactag taacggccgc cagtgtgctg gaattcggct
      721 tggggatatc caccatgctg ctatccgtgc cgctgctgct cggcctcctc ggcctggcca
      781 tcgccagatc tgcgaacgac accgtagttg tgggctcgat catctttaca gaagggatta
      841 tcgtcgcaaa catggtggca gagatgattg aggcgcatac agaccttaag gtggttcgca
      901 aactgaacct tggcggggtc aacgttaact ttgaagccat taaacgcgga ggtgcgaata
      961 atggtattga catttacgtg gagtacactg ggcacggtct tgtggatatt ctggggttcc
     1021 cggagccgaa cgtctatatc accgccgaca agcagaagaa cggcatcaag gcgaacttca
     1081 agatccgcca caacgtggag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     1141 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     1201 tgctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     1261 ccgccgggat cactctcggc atggacgagc tgtacaaggg cggtaccgga gggagcatga
     1321 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     1381 taaacggcca caagttcagc gtgcgcggcg agggcgaggg cgatgccacc aacggcaagc
     1441 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     1501 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     1561 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc agcttcaagg
     1621 acgacggcac ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     1681 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     1741 agtacaactt tccgccgccc gcgactactg atccagaagg tgcatacgaa accgtgaaga
     1801 aggagtacaa acgtaaatgg aatattgtat ggctcaaacc actgggattc aacaatacgt
     1861 atacgcttac cgttaaagac gaactggcga aacagtataa ccttaaaacc ttcagtgact
     1921 tagcgaaaat ctcggataag ctgattctgg gtgcaacgat gcatttttta gaaagtcccg
     1981 atggttaccc aggcctgcaa aaactgtaca atttcaaatt caagcacacc aaaagcatgg
     2041 acatgggtat tcgctatacc gccattgata ataacgaagt tcaggtaatt gatgcctccg
     2101 ccactgatgg cttgctggtg agccacaaat taaaaattct ggaggatgat aaagcgttct
     2161 tcccgccgta ttatgctgcc cccatcatcc gtcaggatgt cttagataag catcctgaac
     2221 tgaaggacgt gctgaacaaa ctcgcgaatc aaatttcact ggaagaaatg cagaaactga
     2281 attacaaggt ggacggtgag ggtcaggacc cagcgaaagt agctaaggag tttttgaaag
     2341 agaaaggttt aattctgcag acagcagaaa aagatgagtt gtgagcggcc gctcgagatc
     2401 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     2461 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     2521 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     2581 ggaggattgg gaagacaata gcaggcgtgc tggggatgcg gtgggctcta tggcttctga
     2641 ggcggaaaga accagtggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     2701 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctga
     2761 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     2821 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     2881 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     2941 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     3001 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     3061 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     3121 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     3181 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     3241 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     3301 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     3361 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     3421 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     3481 ttaaatcaat ctaaagtata tatgagtaac ctgaggctat ggcagggcct gccgccccga
     3541 cgttggctgc gagccctggg ccttcacccg aacttggggg gtggggtggg gaaaaggaag
     3601 aaacgcgggc gtattggccc caatggggtc tcggtggggt atcgacagag tgccagccct
     3661 gggaccgaac cccgcgttta tgaacaaacg acccaacacc gtgcgtttta ttctgtcttt
     3721 ttattgccgt catagcgcgg gttccttccg gtattgtctc cttccgtgtt tcagttagcc
     3781 tccccctagg gtgggcgaag aactccagca tgagatcccc gcgctggagg atcatccagc
     3841 cggcgtcccg gaaaacgatt ccgaagccca acctttcata gaaggcggcg gtggaatcga
     3901 aatctcgtga tggcaggttg ggcgtcgctt ggtcggtcat ttcgaacccc agagtcccgc
     3961 tcagaagaac tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat
     4021 accgtaaagc acgaggaagc ggtcagccca ttcgccgcca agctcttcag caatatcacg
     4081 ggtagccaac gctatgtcct gatagcggtc cgccacaccc agccggccac agtcgatgaa
     4141 tccagaaaag cggccatttt ccaccatgat attcggcaag caggcatcgc catgggtcac
     4201 ggcgagatcc tcgccgtcgg gcttgctcgc cttgagcctg gcgaacagtt cggctggcgc
     4261 gagcccctga tgctcttcgt ccagatcatc ctgatcgaca agaccagctt ccatccgagt
     4321 acgtgctcgc tcgatgcgat gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag
     4381 cgtatgcagc cgccgcattg catcagccat gatggatact ttctcggcag gagcaaggtg
     4441 agatgacagg agatcctgcc ccggcacttc gcccaatagc agccagtccc ttcccgcttc
     4501 agtgacaacg tcgagcacag ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg
     4561 cgctgcctcg tcttgcagtt cattcagggc accggacagg tcggtcttga caaaaagaac
     4621 cgggcgcccc tgcgctgaca gccggaacac ggcggcatca gagcagccga ttgtctgttg
     4681 tgcccagtca tagccgaata gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc
     4741 atcttgttca atcatgcgaa acgatcctca tcctgtctct tgatcgatct ttgcaaaagc
     4801 ctaggcctcc aaaaaagcct cctcactact tctggaatag ctcagaggcc gggcctcggc
     4861 ctctgcataa ataaaaaaaa ttagtcagcc atggggcgga gaatgggcgg aactgggcgg
     4921 agttaggggc gggatgggcg gagttagggg cgggactatg gttgctgact aattgagatg
     4981 cttgctttgc atacttctgc ctgctgggga gcctggggac tttccacacc tggttgctga
     5041 ctaattgaga tgcttgcttt gcatacttct gcctgctggg gagcctgggg actttccaca
     5101 ccctaactga cacacattcc acagctggtt ctttccgcct caggactctt cctttttcaa
     5161 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     5221 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5281 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5341 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5401 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5461 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5521 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5581 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5641 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5701 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5761 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5821 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5881 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5941 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6001 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgagtact
     6061 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     6121 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6181 aaaagtgcca cctgacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     6241 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     6301 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     6361 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     6421 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     6481 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     6541 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     6601 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt acgc
//
