LOCUS       sequence-329216-        6845 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cysteine mutated nAChR beta2 subunit (E61C) for the attachment of a 
            photoswitchable tethered ligand .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6845)
  AUTHORS   Durand-de Cuttoli R, Mondoloni S, Marti F, Lemoine D, Nguyen C, 
            Naude J, d'Izarny-Gargas T, Pons S, Maskos U, Trauner D, Kramer RH, 
            Faure P, Mourot A
  TITLE     Manipulating midbrain dopamine neurons and reward-related behaviors 
            with light-controllable nicotinic acetylcholine receptors.
  JOURNAL   Elife. 2018 Sep 4;7. pii: 37487. doi: 10.7554/eLife.37487.
  PUBMED    30176987
REFERENCE   2  (bases 1 to 6845)
  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: "2018-09-4"; volume: "7. pii: 37487. doi"; pages: " 
            10.7554/eLife.37487"
COMMENT     Sequence Label: pIRES2-CMV-nAChRb2(E61C)-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..6845
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      256..265
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1777..1786
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    join(1840..2414,2415..2417,2418..2426)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2007..2024)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2234..2253
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2427..2429,2430..2432,2433..3146)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2427..2429,2430..2432,2433..3143)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2472..2493)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2733..2752)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3080..3101
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3269..3390
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3306..3325)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3360..3379
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3397..3852)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3534..3555)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3746..3765
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3879..3983
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3985..4342
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3985..4176
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4008..4028)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4193..4328
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4377..5171
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4377..5171
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4431..4450)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5041..5060
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5359..5378)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5403..5450
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5779..6367
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6268..6287
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        6542..6845
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg
      241 actcagatct cgagcgccac catggcccgg tgctccaact ctatggcgct gctgttcagc
      301 tttggcctcc tttggctgtg ttcaggggtt ttgggtactg acacagagga gcggctggtg
      361 gagcatctct tggatccttc ccgctataac aagctgatcc gtccagctac taatggctct
      421 gagctggtga ctgtacagct catggtatca ttggcacagc tcatcagtgt gcacgagcgg
      481 gagcagatca tgaccaccaa cgtctggctg acccagtgct gggaagatta tcgcctcaca
      541 tggaagcctg aggatttcga caatatgaag aaagtccgac ttccttctaa gcacatctgg
      601 ctcccagatg tggttctata caacaatgct gacggcatgt acgaagtctc cttctattcc
      661 aatgctgtgg tctcctatga tggcagcatc ttttggctac cgcctgccat ctacaagagc
      721 gcatgcaaga ttgaggtgaa gcacttccca tttgaccagc agaactgcac catgaagttc
      781 cgctcctgga cctacgaccg cactgagatt gacctggtgc tcaaaagcga tgtggccagc
      841 ctggacgact tcacacctag tggggagtgg gacatcatcg cactgccagg ccgacgcaac
      901 gagaacccag acgactccac ctacgtggac atcacctacg acttcatcat tcgtcgcaaa
      961 ccgctcttct acaccatcaa cctcatcatc ccctgcgtac tcatcacctc gctggccatc
     1021 ctggtcttct acctgccctc agactgtggt gaaaagatga cactttgtat ttctgtgctg
     1081 ctggcgctca cggtgttcct gctgctcatc tccaagattg tgcctcccac ctccctcgac
     1141 gtaccgctgg tgggaaagta cctcatgttc accatggtgc tagtcacctt ctccatcgtc
     1201 actagcgtgt gtgtgctcaa tgtgcaccac cgttcgccta ccacgcacac catggcgccc
     1261 tgggtcaagg tggtcttcct ggagaagctg cccaccctcc tcttcctgca gcagccacgc
     1321 caccgctgtg cacgtcaacg cctgcgcttg cgaaggcgcc agcgggaacg tgagggggca
     1381 ggcacactat tcttccgcga aggtcctgca gctgacccat gtacctgctt tgtcaatcct
     1441 gcatcaatgc agggcttggc tggggctttc caggctgagc ccgctgcagc cggtctgggg
     1501 cgctcgatgg gcccatgcag ctgtggcctc cgggaagctg tggacggtgt acgcttcatt
     1561 gcggaccata tgcgaagtga agatgatgac cagagtgtga gggaggattg gaaatacgtt
     1621 gccatggtga tcgaccgcct gttcctctgg atctttgtct ttgtctgtgt ctttgggacc
     1681 attggcatgt tcctgcagcc tctcttccag aactacactg ccacgacctt ccttcactct
     1741 gaccactcag ctcccagctc caagtgaatc ccgcggccat ggcggccggg agcatgcgac
     1801 gtcgggcccg gtaccgaatt catagctagc atcgatccgc ccctctccct cccccccccc
     1861 taacgttact ggccgaagcc gcttggaata aggccggtgt gcgtttgtct atatgttatt
     1921 ttccaccata ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc ctgtcttctt
     1981 gacgagcatt cctaggggtc tttcccctct cgccaaagga atgcaaggtc tgttgaatgt
     2041 cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa acaacgtctg tagcgaccct
     2101 ttgcaggcag cggaaccccc cacctggcga caggtgcctc tgcggccaaa agccacgtgt
     2161 ataagataca cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt ggatagttgt
     2221 ggaaagagtc aaatggctct cctcaagcgt attcaacaag gggctgaagg atgcccagaa
     2281 ggtaccccat tgtatgggat ctgatctggg gcctcggtac acatgcttta catgtgttta
     2341 gtcgaggtta aaaaaacgtc taggcccccc gaaccacggg gacgtggttt tcctttgaaa
     2401 aacacgatga taatatggcc acaaccatgg tgagcaaggg cgaggagctg ttcaccgggg
     2461 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     2521 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     2581 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct
     2641 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     2701 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     2761 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     2821 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct
     2881 atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca
     2941 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     3001 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc
     3061 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     3121 tcggcatgga cgagctgtac aagtaaagcg gccgcgactc tagatcataa tcagccatac
     3181 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     3241 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     3301 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     3361 tggtttgtcc aaactcatca atgtatctta aggcgtaaat tgtaagcgtt aatattttgt
     3421 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     3481 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     3541 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     3601 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     3661 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     3721 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     3781 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     3841 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     3901 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     3961 aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca
     4021 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4081 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4141 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4201 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4261 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4321 tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga
     4381 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4441 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4501 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag
     4561 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4621 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4681 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4741 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4801 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4861 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg
     4921 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4981 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     5041 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     5101 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     5161 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     5221 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     5281 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     5341 ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat
     5401 gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg
     5461 ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata
     5521 cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct
     5581 cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta
     5641 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     5701 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     5761 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5821 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5881 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5941 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     6001 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6061 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     6121 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     6181 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     6241 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     6301 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     6361 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     6421 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgccatgca
     6481 ttagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
     6541 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
     6601 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
     6661 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
     6721 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
     6781 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
     6841 ccatg
//
