LOCUS       sequence-202568-        7074 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid to target the ecAT3.10 extracellular 
            ATP sensor to the cell surface..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7074)
  AUTHORS   Conley JM, Radhakrishnan S, Valentino SA, Tantama M
  TITLE     Imaging extracellular ATP with a genetically-encoded, ratiometric 
            fluorescent sensor.
  JOURNAL   PLoS One. 2017 Nov 9;12(11):e0187481. doi: 
            10.1371/journal.pone.0187481. eCollection 2017.
  PUBMED    29121644
REFERENCE   2  (bases 1 to 7074)
  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: "PLoS One.";
            date: "2017-11-9"; volume: "12(11):e0187481. doi"; pages: " 
            10.1371/journal.pone.0187481. eCollection 2017"
COMMENT     Sequence Label: ecAT3.10
FEATURES             Location/Qualifiers
     source          1..7074
                     /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"
     CDS             join(802..804,805..807,808..1320)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIE"
     CDS             join(1900..1902,1903..1905,1906..2616)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2623..2652
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2656..2802
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(2824..2841)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2830..3054
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(3186..3774)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3266..3285)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4103..4150
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4175..4194
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4382..5176)
                     /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
                     GLAPAELFARLKASKPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4493..4512)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5103..5122
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(5275..5294)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     enhancer        5373..5564
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5521..5541
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(5624..6415,6416..6484))
                     /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     6247..6266
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6485..6589)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6616..7071
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6703..6722)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6913..6934
                     /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 taccgaactc ggatccacta gtaacggccg ccagtgtgct ggaattcggc
      721 ttggggatat ccaccatgga gacagacaca ctcctgctat gggtactgct gctctgggtt
      781 ccaggttcca ctggtgacag aatggtgagc aagggcgagg agctgttcac cggggtggtg
      841 cccatcctgg tcgagctgga cggcgacgta aacggccaca ggttcagcgt gtccggcgag
      901 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
      961 ctgcccgtgc cctggcccac cctcgtgacc accctgacct ggggcgtgca gtgcttcgcc
     1021 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1081 gtccaggagc gtaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     1141 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1201 gacggcaaca tcctggggca caagctggag tacaactaca tcagccacaa cgtctatatc
     1261 accgccgaca agcagaagaa cggcatcaag gcccacttca agatccgcca caacatcgag
     1321 gacggcggcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1381 gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac
     1441 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccggtat cgatatgaaa
     1501 acgatccacg tgagcgtcac tactcctgat ggcccggtgt acgaagacga tgttgaaatg
     1561 gtcagcgtca aagcgaaaag cggcgagctc ggcattttgc cggggcacat tccgcttaag
     1621 gccccgctcg agatcagcgc ggcccggctg aaaaaaggcg gcaaaacgca atacattgcc
     1681 gtcagcggcg gcaatttgga agtccgcccg gacaaagtga cgattaacgc tcaagctgct
     1741 gaacgggcgg aggacattga cgtcctccgc gccaaagcgg cgaaagagcg ggcggagcgc
     1801 cgcctgcaaa gccagcagga cgacatcgac ttcaaacggg ccgaactggc gttaaaacgc
     1861 gccatgaacc gtttgagcgt tgccgaaatg aaggaattca tggtgagcaa gggcgaggag
     1921 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1981 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagctg
     2041 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgggctac
     2101 ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     2161 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     2221 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     2281 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     2341 agccacaacg tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag
     2401 atccgccaca acatcgagga cggcggcgtg cagctcgccg accactacca gcagaacacc
     2461 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccaag
     2521 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     2581 gccgggatca ctctcggcat ggacgagctg tacaaggtcg acgaacaaaa actcatctca
     2641 gaagaggatc tgaatgctgt gggccaggac acgcaggagg tcatcgtggt gccacactcc
     2701 ttgcccttta aggtggtggt gatctcagcc atcctggccc tggtggtgct caccatcatc
     2761 tcccttatca tcctcatcat gctttggcag aagaagccac gttaggcggc cgctcgagat
     2821 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     2881 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
     2941 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     3001 gggaggattg ggaagacaat agcaggcgtg ctggggatgc ggtgggctct atggcttctg
     3061 aggcggaaag aaccagtggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     3121 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3181 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3241 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3301 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3361 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3421 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3481 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3541 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3601 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3661 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3721 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     3781 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3841 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3901 tttaaatcaa tctaaagtat atatgagtaa cctgaggcta tggcagggcc tgccgccccg
     3961 acgttggctg cgagccctgg gccttcaccc gaacttgggg ggtggggtgg ggaaaaggaa
     4021 gaaacgcggg cgtattggcc ccaatggggt ctcggtgggg tatcgacaga gtgccagccc
     4081 tgggaccgaa ccccgcgttt atgaacaaac gacccaacac cgtgcgtttt attctgtctt
     4141 tttattgccg tcatagcgcg ggttccttcc ggtattgtct ccttccgtgt ttcagttagc
     4201 ctccccctag ggtgggcgaa gaactccagc atgagatccc cgcgctggag gatcatccag
     4261 ccggcgtccc ggaaaacgat tccgaagccc aacctttcat agaaggcggc ggtggaatcg
     4321 aaatctcgtg atggcaggtt gggcgtcgct tggtcggtca tttcgaaccc cagagtcccg
     4381 ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga
     4441 taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac
     4501 gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga
     4561 atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca
     4621 cgacgagatc ctcgccgtcg ggcttgctcg ccttgagcct ggcgaacagt tcggctggcg
     4681 cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag
     4741 tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa
     4801 gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt
     4861 gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt
     4921 cagtgacaac gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc
     4981 gcgctgcctc gtcttgcagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa
     5041 ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt
     5101 gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc
     5161 catcttgttc aatcatgcga aacgatcctc atcctgtctc ttgatcgatc tttgcaaaag
     5221 cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc cgggcctcgg
     5281 cctctgcata aataaaaaaa attagtcagc catggggcgg agaatgggcg gaactgggcg
     5341 gagttagggg cgggatgggc ggagttaggg gcgggactat ggttgctgac taattgagat
     5401 gcttgctttg catacttctg cctgctgggg agcctgggga ctttccacac ctggttgctg
     5461 actaattgag atgcttgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
     5521 accctaactg acacacattc cacagctggt tctttccgcc tcaggactct tcctttttca
     5581 ataaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5641 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5701 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5761 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5821 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5881 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5941 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6001 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6061 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6121 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6181 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6241 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6301 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6361 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6421 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6481 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6541 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6601 gaaaagtgcc acctgacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     6661 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     6721 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     6781 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     6841 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     6901 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     6961 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     7021 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt tacg
//
