LOCUS       sequence-353959-        6622 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 6622)
  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 6622)
  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_cyto
FEATURES             Location/Qualifiers
     source          1..6622
                     /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"
     primer_bind     1185..1206
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1309..1330)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2371..2388)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2377..2601
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2733..3321)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2813..2832)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3650..3697
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3722..3741
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3929..4717)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDQEHQGL
                     APAELFARLKASKPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIALA
                     TRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4040..4059)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4644..4663
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4752..5112)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4766..4904
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        4921..5112
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5069..5089
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(5172..5963,5964..6032))
                     /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     5795..5814
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6033..6137)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6164..6619
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6251..6270)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6461..6482
                     /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 ggactattta 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 taccgaactc ggatccacta gtaacggccg ccagtgtgct ggaattcggc
      721 ttggggatat ccaccatgtc tgcgaacgac accgtagttg tgggctcgat catctttaca
      781 gaagggatta tcgtcgcaaa catggtggca gagatgattg aggcgcatac agaccttaag
      841 gtggttcgca aactgaacct tggcggggtc aacgttaact ttgaagccat taaacgcgga
      901 ggtgcgaata atggtattga catttacgtg gagtacactg ggcacggtct tgtggatatt
      961 ctggggttcc cggagccgaa cgtctatatc accgccgaca agcagaagaa cggcatcaag
     1021 gcgaacttca agatccgcca caacgtggag gacggcagcg tgcagctcgc cgaccactac
     1081 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     1141 acccagtccg tgctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     1201 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggg cggtaccgga
     1261 gggagcatga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
     1321 gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg cgatgccacc
     1381 aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
     1441 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
     1501 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     1561 agcttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     1621 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1681 cacaagctgg agtacaactt tccgccgccc gcgactactg atccagaagg tgcatacgaa
     1741 accgtgaaga aggagtacaa acgtaaatgg aatattgtat ggctcaaacc actgggattc
     1801 aacaatacgt atacgcttac cgttaaagac gaactggcga aacagtataa ccttaaaacc
     1861 ttcagtgact tagcgaaaat ctcggataag ctgattctgg gtgcaacgat gcatttttta
     1921 gaaagtcccg atggttaccc aggcctgcaa aaactgtaca atttcaaatt caagcacacc
     1981 aaaagcatgg acatgggtat tcgctatacc gccattgata ataacgaagt tcaggtaatt
     2041 gatgcctccg ccactgatgg cttgctggtg agccacaaat taaaaattct ggaggatgat
     2101 aaagcgttct tcccgccgta ttatgctgcc cccatcatcc gtcaggatgt cttagataag
     2161 catcctgaac tgaaggacgt gctgaacaaa ctcgcgaatc aaatttcact ggaagaaatg
     2221 cagaaactga attacaaggt ggacggtgag ggtcaggacc cagcgaaagt agctaaggag
     2281 tttttgaaag agaaaggttt aattctggat attgatgaac tggcgctgaa atttgcgggc
     2341 ctggatctgt gagcggccgc tcgagatcag cctcgactgt gccttctagt tgccagccat
     2401 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     2461 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     2521 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcgtgctg
     2581 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagtggcggt aatacggtta
     2641 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2701 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2761 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     2821 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     2881 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     2941 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3001 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3061 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3121 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
     3181 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3241 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3301 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3361 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3421 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaacct
     3481 gaggctatgg cagggcctgc cgccccgacg ttggctgcga gccctgggcc ttcacccgaa
     3541 cttggggggt ggggtgggga aaaggaagaa acgcgggcgt attggcccca atggggtctc
     3601 ggtggggtat cgacagagtg ccagccctgg gaccgaaccc cgcgtttatg aacaaacgac
     3661 ccaacaccgt gcgttttatt ctgtcttttt attgccgtca tagcgcgggt tccttccggt
     3721 attgtctcct tccgtgtttc agttagcctc cccctagggt gggcgaagaa ctccagcatg
     3781 agatccccgc gctggaggat catccagccg gcgtcccgga aaacgattcc gaagcccaac
     3841 ctttcataga aggcggcggt ggaatcgaaa tctcgtgatg gcaggttggg cgtcgcttgg
     3901 tcggtcattt cgaaccccag agtcccgctc agaagaactc gtcaagaagg cgatagaagg
     3961 cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt
     4021 cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga tagcggtccg
     4081 ccacacccag ccggccacag tcgatgaatc cagaaaagcg gccattttcc accatgatat
     4141 tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc ttgctcgcct
     4201 tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttgatca tcctgatcga
     4261 caagaccggc ttccatccga gtacgtgctc gctcgatgcg atgtttcgct tggtggtcga
     4321 atgggcaggt agccggatca agcgtatgca gccgccgcat tgcatcagcc atgatggata
     4381 ctttctcggc aggagcaagg tgagatgaca ggagatcctg ccccggcact tcgcccaata
     4441 gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa ggaacgcccg
     4501 tcgtggccag ccacgatagc cgcgctgcct cgtcttgcag ttcattcagg gcaccggaca
     4561 ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga cagccggaac acggcggcat
     4621 cagagcagcc gattgtctgt tgtgcccagt catagccgaa tagcctctcc acccaagcgg
     4681 ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg aaacgatcct catcctgtct
     4741 cttgatcgat ctttgcaaaa gcctaggcct ccaaaaaagc ctcctcacta cttctggaat
     4801 agctcagagg ccgaggaggc ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca
     4861 tggggcggag aatgggcgga actgggcgga gttaggggcg ggatgggcgg agttaggggc
     4921 gggactatgg ttgctgacta attgagatgc ttgctttgca tacttctgcc tgctggggag
     4981 cctggggact ttccacacct ggttgctgac taattgagat gcttgctttg catacttctg
     5041 cctgctgggg agcctgggga ctttccacac cctaactgac acacattcca cagctggttc
     5101 tttccgcctc aggactcttc ctttttcaat aaatcaatct aaagtatata tgagtaaact
     5161 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     5221 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     5281 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     5341 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     5401 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     5461 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     5521 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     5581 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     5641 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     5701 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     5761 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     5821 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     5881 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     5941 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     6001 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     6061 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     6121 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgcgcc ctgtagcggc
     6181 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     6241 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     6301 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     6361 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     6421 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     6481 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     6541 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     6601 atattaacgc ttacaattta cg
//
