LOCUS       Exported                7490 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Archon2 fluorescent voltage reporter in mammalian expression 
            plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    pCAG-Archon2-KGC-GFP-ER2-WPRE
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7490)
  AUTHORS   Piatkevich KD, Jung EE, Straub C, Linghu C, Park D, Suk HJ, Hochbaum
            DR, Goodwin D, Pnevmatikakis E, Pak N, Kawashima T, Yang CT, Rhoades
            JL, Shemesh O, Asano S, Yoon YG, Freifeld L, Saulnier JL, Riegler C,
            Engert F, Hughes T, Drobizhev M, Szabo B, Ahrens MB, Flavell SW, 
            Sabatini BL, Boyden ES
  TITLE     A robotic multidimensional directed evolution approach applied to 
            fluorescent voltage reporters.
  JOURNAL   Nat Chem Biol. 2018 Feb 26. pii: 10.1038/s41589-018-0004-9. doi: 
            10.1038/s41589-018-0004-9.
  PUBMED    29483642
REFERENCE   2  (bases 1 to 7490)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7490
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    24..35
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     enhancer        39..418
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        421..696
                     /label=chicken beta-actin promoter
     intron          697..1708
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1632..1654
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1716..1735
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1769..1778
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2642..3355
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2684..2705)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2945..2964)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3292..3313
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          3416..3548
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3531..3550
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    3557..4145
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3610..3630)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4028..4039)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4172..4189)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4178..4289
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    4581..4635
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4634..4653)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4994..5010)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4994..5010)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5007..5029)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5018..5034
                     /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(5042..5072)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5087..5108
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5167..5362
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5213..5348
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5275..5294
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    5368..5502
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5405..5424)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5459..5478
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5570..5587)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5741..6329)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5821..5840)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6798..7175)
                     /label=ISS
                     /note="immunostimulatory sequence from the AmpR gene; 
                     contains unmethylated CpG dinucleotides in the context of 
                     5'-AACGTT-3' (Sato et al., 1996)"
     primer_bind     7122..7141
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7360..7464)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agagtcatcc tcatcgtcga cattgattat tgactagtta
       61 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
      121 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
      181 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
      241 ggactattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
      301 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
      361 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggg
      421 tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc ccacccccaa
      481 ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg gggggggggg
      541 cgcgcgccag gcggggcggg gcgggggcga gggcggggcg gggcgaggcg gagaggtgcg
      601 gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg
      661 cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgtt gccttcgccc
      721 cgtgccccgc tccgcgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc
      781 ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagcg cttggtttaa
      841 tgacggctcg tttcttttct gtggctgcgt gaaagcctta aagggctccg ggagggccct
      901 ttgtgcgggg gggagcggct cggggggtgc gtgcgtgtgt gtgtgcgtgg ggagcgccgc
      961 gtgcggcccg cgctgcccgg cggctgtgag cgctgcgggc gcggcgcggg gctttgtgcg
     1021 ctccgcgtgt gcgcgagggg agcgcggccg ggggcggtgc cccgcggtgc gggggggctg
     1081 cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag ggggtgtggg
     1141 cgcggcggtc gggctgtaac ccccccctgc acccccctcc ccgagttgct gagcacggcc
     1201 cggcttcggg tgcggggctc cgtgcggggc gtggcgcggg gctcgccgtg ccgggcgggg
     1261 ggtggcggca ggtgggggtg ccgggcgggg cggggccgcc tcgggccggg gagggctcgg
     1321 gggaggggcg cggcggcccc ggagcgccgg cggctgtcga ggcgcggcga gccgcagcca
     1381 ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc aaatctgtgc
     1441 ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg cgaagcggtg
     1501 cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc gccgccgtcc
     1561 ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc gggggggacg
     1621 gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca
     1681 tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta ttgtgctgtc
     1741 tcatcatttt ggcaaagaat tcggtaccgc caccatggtt tctatcgcac tgcaggctgg
     1801 ttatgacctc ctcggagatg gtagacctga aaccctgtgg ctcggaattg gtacactgct
     1861 gatgctgatc ggaaccttct acttcctggt tagagggtgg ggagtgactg acaaggatgc
     1921 acgggagtac tacgcagtgc ccatcctggt cagcggtatt gccagtgcag cttatctgtc
     1981 aatgttcttt ggtatcggcc tcacagaggt gccagtcggt ggagagatgc tggacatcta
     2041 ctacgcaaga tacgcacact ggctgttcag cacaccactg ctgctgctgg acctggcact
     2101 cctggccaaa gttgatagag tgatcatcgg cactctcgtt ggcgtcgatg ctctgatgat
     2161 tgtcacaggc ctgataggag cactgtcaca tactgccatt gccagatact cttggtggct
     2221 gttctcaact atctgtatga tcgtcgttct gtatgtcctg gccacttctc tgcggtcagc
     2281 cgctaaggag agaggcccag aggttgcctc aaccttcaac atcctcaccg cactggttct
     2341 ggtcctgtgg actgcttacc ctatcatctg gattatcggc accgagggtg ccggagttgt
     2401 gggcctggga atcgaaactc tgctgttcat ggtgctggac gtcacctgca aggtgggctt
     2461 cgggtttatc ctgctcaggt caagagccat tctgggagat acagaagcac ctgaaccttc
     2521 agctggagct gataccggtg ccgccgaccg cccggtagta gcagtgagca aggcggccgc
     2581 caagagcagg atcaccagcg agggcgagta catccccctg gaccagatcg acatcaacgt
     2641 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     2701 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     2761 caagctgacc ctgaagttca tttgcaccac cggcaagctg cccgtgccct ggcccaccct
     2821 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     2881 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     2941 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     3001 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     3061 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     3121 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     3181 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     3241 cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     3301 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagttctg
     3361 ctacgagaac gaggtgtaat gtacaagtaa agcggccccg gactcgaggc cgcaggtaag
     3421 tatcaaggtt acaagacagg tttaaggaga ccaatagaaa ctgggcttgt cgagacagag
     3481 aagactcttg cgtttctgat aggcacctat tggtcttact gacatccact ttgcctttct
     3541 ctccacaggt gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat
     3601 tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca
     3661 tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc
     3721 tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc
     3781 tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt
     3841 cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg
     3901 gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga agctgacgtc
     3961 ctttccatgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta
     4021 cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg
     4081 gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc
     4141 cccgcctgga attcgagctc ggtacgatca gcctcgactg tgccttctag ttgccagcca
     4201 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     4261 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     4321 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggccagct
     4381 tttgttccct ttagggttaa ttgcgcgctt ggcgctaatc atggtcatag ctgtttccac
     4441 tcctcaggtg caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca
     4501 caaataccac tgagatcgtt ttccctctgc caaaaattat ggggacatca tgaagcccct
     4561 tgagcatctg acttctggct aataaaggaa atttattttc attgcaatag tgtgttgaat
     4621 tttttgtgtc tctcactcgg aaggacatat gggagggcaa atcatttaaa acatcagaat
     4681 gagtatttgg tttagagttt ggcaacatat gccatatgct ggctgccatg aacaaaggtg
     4741 gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag
     4801 aaaagccttg acttgaggtt agattttttt ttatattttg tttgtgttat ttttttcttt
     4861 aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact
     4921 actcccagtc atagctgtcc ctcttctctt atgaagatcc ctcgacctgc agcccaagct
     4981 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5041 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5101 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5161 ggatccgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     5221 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     5281 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     5341 tggaggccta ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa
     5401 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     5461 ggtttgtcca aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg
     5521 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     5581 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     5641 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     5701 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     5761 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     5821 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5881 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct
     5941 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     6001 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     6061 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     6121 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     6181 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     6241 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     6301 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     6361 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     6421 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     6481 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     6541 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     6601 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6661 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     6721 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6781 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     6841 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6901 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6961 tggccgcagt gttatcactc atggttatgg agcactgcat aattctctta ctgtcatgcc
     7021 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7081 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7141 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7201 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7261 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7321 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7381 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7441 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
//
