LOCUS       sequence-257479-        7134 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7134)
  AUTHORS   Jin L, Han Z, Platisa J, Wooltorton JR, Cohen LB, Pieribone VA
  TITLE     Single action potentials and subthreshold electrical events imaged 
            in neurons with a fluorescent protein voltage probe.
  JOURNAL   Neuron. 2012 Sep 6;75(5):779-85.
  PUBMED    22958819
REFERENCE   2  (bases 1 to 7134)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Neuron."; 
            date: "2012-09-6"; volume: "75(5)"; pages: "779-85"
COMMENT     Sequence Label: ArcLight-Q239
FEATURES             Location/Qualifiers
     source          1..7134
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        7..984
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1019..1037
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1019..1036
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             1774..2490
                     /codon_start=1
                     /product="pH-sensitive mutant of green fluorescent protein 
                     (Ng et al., 2002)"
                     /label=superecliptic pHluorin
                     /note="excited strongly by blue light at neutral pH but 
                     minimally fluorescent at acidic pH"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNDHQVYIMADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLFTTSTLSKDPNEKRDHMVLLE
                     FVTADGITHGMDELYK"
     primer_bind     complement(1804..1832)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2427..2447
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    2520..2654
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2759..2779)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2764..3093
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2944..3079
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3006..3025
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3142..4167
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3142..4167
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4297..4418
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4456..4472)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4456..4472)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4469..4491)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4480..4496
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4504..4510,4511..4528,4529..4534))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4549..4570
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4687..4704)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4858..5446)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4938..4957)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5617..6408,6409..6477))
                     /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     6240..6259
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6478..6582)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6608..7063)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6745..6766)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6957..6976
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tcgaccgcca attcaatatg gcgtatatgg actcatgcca attcaatatg gtggatctgg
       61 acctgtgcca attcaatatg gcgtatatgg actcgtgcca attcaatatg gtggatctgg
      121 accccagcca attcaatatg gcggacttgg caccatgcca attcaatatg gcggacctgg
      181 cactgtgcca actggggagg ggtctacttg gcacggtgcc aagtttgagg aggggtcttg
      241 gccctgtgcc aagtccgcca tattgaattg gcatggtgcc aataatggcg gccatattgg
      301 ctatatgcca ggatcaatat ataggcaata tccaatatgg ccctatgcca atatggctat
      361 tggccaggtt caatactatg tattggccct atgccatata gtattccata tatgggtttt
      421 cctattgacg tagatagccc ctcccaatgg gcggtcccat ataccatata tggggcttcc
      481 taataccgcc catagccact cccccattga cgtcaatggt ctctatatat ggtctttcct
      541 attgacgtca tatgggcggt cctattgacg tatatggcgc ctcccccatt gacgtcaatt
      601 acggtaaatg gcccgcctgg ctcaatgccc attgacgtca ataggaccac ccaccattga
      661 cgtcaatggg atggctcatt gcccattcat atccgttctc acgcccccta ttgacgtcaa
      721 tgacggtaaa tggcccactt ggcagtacat caatatctat taatagtaac ttggcaagta
      781 cattactatt ggaagtacgc cagggtacat tggcagtact cccattgacg tcaatggcgg
      841 taaatggccc gcgatggctg ccaagtacat ccccattgac gtcaatgggg aggggcaatg
      901 acgcaaatgg gcgttccatt gacgtaaatg ggcggtaggc gtgcctaatg ggaggtctat
      961 ataagcaatg ctcgtttagg gaaccgccat tctgcctggg gacgtcggag ctagcttgat
     1021 ttaggtgaca ctatagaata caagcttatg gagggattcg acggttcaga ttttagtcct
     1081 ccagctgatt tagttggcgt tgacggtgca gtcatgcgga acgtcgttga cgtcacgata
     1141 aatggtgacg tcactgctcc gccgaaagct gcgccaagaa aaagtgaatc ggtaaagaaa
     1201 gttcattgga atgatgtaga ccaaggaccg agtgaaaaac cagagacaag acaggaggaa
     1261 cgaatagata tacccgagat ttcaggtcta tggtggggcg agaatgaaca tggagtggac
     1321 gatgggagaa tggagatacc tactactggt gtaggtcgcg tccagtttcg tgtccgagca
     1381 gtgattgatc atctagggat gcgagtcttt ggagtcttcc taattttctt ggacatcatc
     1441 ctcatgatca ttgatctcag tcttccagga aaaagtgaat cttcacaatc cttttatgac
     1501 gggatggctt tggctctttc ttgttatttc atgctggatt taggattaag gatatttgcc
     1561 tacgggccca agaatttctt caccaacccc tgggaggttg ctgatggttt gattatcgtt
     1621 gtcacattcg tcgtcacgat attttacact gtgttagatg aatacgtgca agaaacagga
     1681 gccgatggtt tggggcaatt ggttgtgttg gcccgtttgc tgcgtgtggt tcgattagca
     1741 agaatatttt attcccacca acaaggggat cccatgagta aaggagaaga acttttcact
     1801 ggagttgtcc caattcttgt tgaattagat ggtgatgtta atgggcacaa attttctgtc
     1861 agtggagagg gtgaaggtga tgcaacatac ggaaaactta cccttaaatt tatttgcact
     1921 actggaaaac tacctgttcc atggccaaca cttgtcacta ctttaactta tggtgttcaa
     1981 tgcttttcaa gatacccaga tcatatgaaa cggcatgact ttttcaagag tgccatgccc
     2041 gaaggttatg tacaggaaag aactatattt ttcaaagatg acgggaacta caagacacgt
     2101 gctgaagtca agtttgaagg tgataccctt gttaatagaa tcgagttaaa aggtattgat
     2161 tttaaagaag atggaaacat tcttggacac aaattggaat acaactataa cgatcaccag
     2221 gtgtacatca tggcagacaa acaaaagaat ggaatcaaag ctaacttcaa aattagacac
     2281 aacattgaag atggaggcgt tcaactagca gaccattatc aacaaaatac tccaattggc
     2341 gatgggcccg tccttttacc agacaaccat tacctgttta caacttctac tctttcgaaa
     2401 gatcccaacg aaaagagaga ccacatggtc cttcttgagt ttgtaacagc tgatgggatt
     2461 acacatggca tggatgaact atacaaatga tctagaacta tagtgagtcg tattacgtag
     2521 atccagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     2581 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     2641 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg
     2701 tgtgggaggt tttttaattg atatccgcgg cgccaatgca ttgggcccgg taccctgtgg
     2761 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2821 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     2881 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     2941 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3001 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     3061 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     3121 ttcggatctg atcagcacgt gatgaaaaag cctgaactca ccgcgacgtc tgtcgagaag
     3181 tttctgatcg aaaagttcga cagcgtctcc gacctgatgc agctctcgga gggcgaagaa
     3241 tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc
     3301 gccgatggtt tctacaaaga tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg
     3361 attccggaag tgcttgacat tggggaattc agcgagagcc tgacctattg catctcccgc
     3421 cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc tgttctgcag
     3481 ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc ttagccagac gagcgggttc
     3541 ggcccattcg gaccgcaagg aatcggtcaa tacactacat ggcgtgattt catatgcgcg
     3601 attgctgatc cccatgtgta tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc
     3661 gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg actgccccga agtccggcac
     3721 ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg acaatggccg cataacagcg
     3781 gtcattgact ggagcgaggc gatgttcggg gattcccaat acgaggtcgc caacatcttc
     3841 ttctggaggc cgtggttggc ttgtatggag cagcagacgc gctacttcga gcggaggcat
     3901 ccggagcttg caggatcgcc gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa
     3961 ctctatcaga gcttggttga cggcaatttc gatgatgcag cttgggcgca gggtcgatgc
     4021 gacgcaatcg tccgatccgg agccgggact gtcgggcgta cacaaatcgc ccgcagaagc
     4081 gcggccgtct ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc
     4141 agcactcgtc cgagggcaaa ggaatagcac gtgctacgag atttcgattc caccgccgcc
     4201 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4261 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     4321 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4381 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtacgcg
     4441 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     4501 acacaacata cgagccggga gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     4561 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     4621 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4681 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4741 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4801 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4861 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4921 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4981 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5041 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5101 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5161 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5221 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5281 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5341 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     5401 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     5461 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5521 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5581 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5641 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5701 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5761 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5821 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5881 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5941 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6001 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6061 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6121 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6181 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6241 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6301 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6361 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     6421 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6481 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6541 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6601 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     6661 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     6721 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     6781 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     6841 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     6901 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     6961 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     7021 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg
     7081 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccag
//
