LOCUS       sequence-242234-        6579 bp    DNA     circular SYN 15-APR-2025
DEFINITION  AAV-mediated expression of Archon1-KGC-GFP-ER2 under the EF1-alpha 
            promoter (1.1kb short version). Using SV40 signal..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6579)
  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 6579)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Chem 
            Biol. 2018 Feb 26. pii: 10.1038/s41589-018-0004-9. doi: 
            10.1038/s41589-018-0004-9."
COMMENT     Sequence Label: pAAV-EF1-alpha-1.1-Archon1-KGC-GFP-ER2
FEATURES             Location/Qualifiers
     source          1..6579
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1932..2387)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2069..2090)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2281..2300
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2459..2588
                     /label=AAV2 ITR
     promoter        2661..3839
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2892..3830
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3787..3807
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      3846..3855
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4719..4721,4722..5432)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(4719..4721,4722..5432)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4761..4782)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5022..5041)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5369..5390
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    5481..6069
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5534..5554)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5952..5963)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    6112..6233
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6123..6142)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6177..6196
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   6395..6524
                     /label=AAV2 ITR
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     1981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
     2401 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     2461 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     2521 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     2581 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag
     2641 gaagatcgta ggttaattaa ggctccggtg cccgtcagtg ggcagagcgc acatcgccca
     2701 cagtccccga gaagttgggg ggaggggtcg gcaattgaac cggtgcctag agaaggtggc
     2761 gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc gagggtgggg
     2821 gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg
     2881 ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg
     2941 gcccttgcgt gccttgaatt acttccacct ggctgcagta cgtgattctt gatcccgagc
     3001 ttcgggttgg aagtgggtgg gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc
     3061 gtgcttgagt tgaggcctgg cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc
     3121 ttcgcgcctg tctcgctgct ttcgataagt ctctagccat ttaaaatttt tgatgacctg
     3181 ctgcgacgct ttttttctgg caagatagtc ttgtaaatgc gggccaagat ctgcacactg
     3241 gtatttcggt ttttggggcc gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt
     3301 cgacgaggcg gggcctgcga gcgcggccac cgagaatcgg acgggggtag tctcaagctg
     3361 gccggcctgc tctggtgcct ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa
     3421 ggctggcccg gtcggcacca gttgcgtgag cggaaagatg gccgcttccc ggccctgctg
     3481 cagggagctc aaaatggagg acgcggcgct cgggagagcg ggcgggtgag tcacccacac
     3541 aaaggaaaag ggcctttccg tcctcagccg tcgcttcatg tgactccacg gagtaccggg
     3601 cgccgtccag gcacctcgat tagttctcga gcttttggag tacgtcgtct ttaggttggg
     3661 gggaggggtt ttatgcgatg gagtttcccc acactgagtg ggtggagact gaagttaggc
     3721 cagcttggca cttgatgtaa ttctccttgg aatttgccct ttttgagttt ggatcttggt
     3781 tcattctcaa gcctcagaca gtggttcaaa gtttttttct tccatttcag gtgtcgtgag
     3841 gatccgccac catggtttct atcgcactgc aggctggtta tgacctcctc ggagatggta
     3901 gacctgaaag tctgtggctc ggaattggta cactgctgat gctgatcgga accttctact
     3961 tcctggttag agcgtgggga gagactgaca aggatgcacg ggagtactac gcagtgacaa
     4021 tcctggtcag cggtattgcc agtgcagctt atctgtcaat gttctttggt attggcctca
     4081 cagaggtgcc agtaggtgga gagatgctga acatctacta cgcaagatac gcacagtggc
     4141 tgttcactac accactgttg ctgctgcacc tggcactcct ggccaaagtt gatagagtga
     4201 ccatcggcac tctcgttggc gtcgatgctc tgatgattgt cacaggcctg ataggaacac
     4261 tgtcacatac tgccattgcc agatactctt ggtggctgtt ctcaactatc tgtatgatcg
     4321 tcgttctgta tgtcctggcc acttctctgc ggtcagccgc taaggagaga ggcccagagg
     4381 ttgcctcaac cttcaacatc ctcaccgcac tggttctggt cctgtggact gcttacccta
     4441 tcatctggat tatcggcacc gagggtgccg gagttgtggg cctgggaatc gaaactttgc
     4501 tgttcatggt gctggacgtc accgccaagg tgggcttcgg ttttatcctg ctcaggtcaa
     4561 gagccattct gcaagataca gaagcacctg aaccttcagc tggagctgat accggtgccg
     4621 ccgaccgccc ggtagtagca gtgagcaagg cggccgccaa gagcaggatc accagcgagg
     4681 gcgagtacat ccccctggac cagatcgaca tcaacgtggt gagcaagggc gaggagctgt
     4741 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     4801 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcattt
     4861 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
     4921 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     4981 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     5041 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     5101 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     5161 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     5221 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     5281 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga
     5341 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     5401 ggatcactct cggcatggac gagctgtaca agttctgcta cgagaacgag gtgtaagaat
     5461 tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg
     5521 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     5581 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     5641 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     5701 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     5761 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     5821 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     5881 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     5941 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     6001 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     6061 cctccccgca tcgataccgt cgacccgggc ggccgcttcg agcagacatg ataagataca
     6121 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     6181 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
     6241 acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt ttttaaagca
     6301 agtaaaacct ctacaaatgt ggtaaaatcg ataaggatct tcctagagca tggctacgta
     6361 gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat ggagttggcc
     6421 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     6481 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcat taatgaatcg
     6541 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgct
//
