LOCUS       sequence-240924-        7655 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the proton pump rhodopsin in mammalian cells under CAG 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7655)
  AUTHORS   Wu L, Dong A, Dong L, Wang SQ, Li Y
  TITLE     PARIS, an optogenetic method for functionally mapping gap junctions.
  JOURNAL   Elife. 2019 Jan 14;8. pii: 43366. doi: 10.7554/eLife.43366.
  PUBMED    30638447
REFERENCE   2  (bases 1 to 7655)
  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: "2019-01-14"; volume: "8. pii: 43366. doi"; pages: " 
            10.7554/eLife.43366"
COMMENT     Sequence Label: pcDNA3.1-ArchT-BFP2-TSERex
FEATURES             Location/Qualifiers
     source          1..7655
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      8..17
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(773..775,776..1486)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIKV
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLLE
                     FRTAAGITLGMDELYK"
     primer_bind     complement(815..836)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1685..1702)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1691..1915
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1961..2389
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2048..2067)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2258..2279
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2398..2418)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2403..2732
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2583..2718
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2645..2664
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2743..3342
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2743..2762)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3239..3259
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3502..3623
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3539..3558)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3593..3612
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3672..3688)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3672..3688)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3685..3707)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3696..3712
                     /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(3720..3726,3727..3744,3745..3750))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3765..3786
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3903..3920)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4074..4662)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4154..4173)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4833..5624,5625..5693))
                     /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     5456..5475
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5694..5798)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        5830..6209
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          6487..7503
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     7427..7449
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     7511..7530
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
ORIGIN
        1 gctagccgcc accatggatc ccatcgctct gcaggcagga tacgacctgc tgggcgacgg
       61 aaggccagag accctgtggc tgggaatcgg aaccctgctg atgctgatcg gcaccttcta
      121 cttcatcgtg aagggctggg gcgtgaccga caaggaggcc agggagtact acagcatcac
      181 aatcctggtg cccggcatcg ccagcgccgc ctacctgagc atgttcttcg gcatcggcct
      241 gaccgaggtg accgtggccg gcgaggtgct ggacatctac tacgccagat acgccgactg
      301 gctgttcacc acccctttgc ttctgctcga cctggccctg ctggctaagg tggacagggt
      361 gagcatcgga accctggtgg gagtggacgc cctgatgatc gtgaccggcc tgatcggcgc
      421 cctgagccac accccactgg ctaggtacag ctggtggctg ttcagcacca tctgcatgat
      481 cgtggtgctg tacttcctgg ctaccagcct gagggctgct gctaaggaga ggggaccaga
      541 ggtggctagc accttcaaca ccctgaccgc cctggtgctg gtgctgtgga ccgcctaccc
      601 catcctgtgg atcatcggaa ccgagggagc tggagtggtg ggactgggaa tcgagaccct
      661 gctgttcatg gtgctggacg tgaccgccaa agtgggcttc ggcttcatcc tgctgaggag
      721 cagggccatc ctgggcgaca ccgaggcccc cgagccccga ccggtagtag cagtgagcaa
      781 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
      841 cggccacaag ttcagcgtga ggggcgaggg cgagggcgat gccaccaacg gcaagctgac
      901 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
      961 cctgagccac ggcgtgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
     1021 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     1081 cggcacctac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     1141 cgagctgaag ggcgtcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     1201 caacttcaac agccacaaca tctatatcat ggccgtcaag cagaagaacg gcatcaaggt
     1261 gaacttcaag atccgccaca acgtggagga cggcagcgtg cagctcgccg accactacca
     1321 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacagccact acctgagcac
     1381 ccagtccgtg ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     1441 ccgcaccgcc gccgggatca ctctcggcat ggacgagctg tacaagaaaa gtagaatcac
     1501 aagcgaaggc gagtacatcc ccctggatca aatagacata aatgtaggtg gattttgtta
     1561 tgagaatgaa tgaaagcttg gtaccgagct cggatccact agtccagtgt ggtggaattc
     1621 tgcagatatc cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga
     1681 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     1741 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     1801 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     1861 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     1921 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     1981 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     2041 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     2101 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     2161 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     2221 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     2281 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     2341 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     2401 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     2461 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     2521 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     2581 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     2641 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     2701 gaggcttttt tggaggccta ggcttttgca aaaagctccc gcatgaccga gtacaagccc
     2761 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     2821 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     2881 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     2941 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     3001 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     3061 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     3121 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     3181 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc
     3241 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     3301 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gattcgaaat gaccgaccaa
     3361 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     3421 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     3481 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     3541 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3601 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     3661 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3721 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3781 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3841 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3901 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3961 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4021 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4081 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4141 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4201 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4261 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4321 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4381 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4441 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4501 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4561 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4621 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4681 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4741 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4801 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4861 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4921 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4981 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5041 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5101 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5161 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5221 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5281 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5341 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5401 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5461 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5521 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5581 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5641 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5701 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5761 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5821 cctgacgtcg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
     5881 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
     5941 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
     6001 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     6061 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     6121 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     6181 tctacgtatt agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc
     6241 tccccatctc ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt
     6301 gtgcagcgat gggggcgggg gggggggggg nnsgcgccrg gcggggsggg gsggggsgrg
     6361 gggsggggsg gggcgrggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga
     6421 aagtttcctt ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg
     6481 cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc
     6541 cgcccgcccc ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct
     6601 tctcctccgg gctgtaatta gcgcttggtt taatgacggc ttgtttcttt tctgtggctg
     6661 cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg gggggagcgg ctcggggggt
     6721 gcgtgcgtgt gtgtgtgcgt ggggagcgcc gcgtgcggct ccgcgctgcc cggcggctgt
     6781 gagcgctgcg ggcgcggcgc ggggctttgt gcgctccgca gtgtgcgcga ggggagcgcg
     6841 gccgggggcg gtgccccgcg gtgcgggggg ggctgcgagg ggaacaaagg ctgcgtgcgg
     6901 ggtgtgtgcg tgggggggtg agcagggggt gtgggcgcgt cggtcgggct gcaacccccc
     6961 ctgcaccccc ctccccgagt tgctgagcac ggcccggctt cgggtgcggg gctccgtacg
     7021 gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc
     7081 ggggcggggc cgcctcgggc cggggrgggc tcgggggrgg ggcgcggcgg cccccggagc
     7141 gccggcggct gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag
     7201 agggcgcagg gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc
     7261 cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc
     7321 ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc tccctctcca gcctcggggc
     7381 tgtccgcggg gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg
     7441 cgtgtgaccg gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta
     7501 cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattcctc
     7561 gaggaattca ctcctcaggt gcaggctgcc tatcagaagg tggtggctgg tgtggccaat
     7621 gccctggctc acaaatacca ctgagatctg catga
//
