LOCUS       Exported                6972 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV-mediated expression of Archon1-KGC-GFP-ER2 under the EF1-alpha 
            promoter (1.1kb short version), in floxed/reversed (Cre-dependent) 
            manner. Using bGHpA signal..
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-EF1-alpha-1.1-FLEX-rc [Archon1-KGC-GFP-ER2]
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6972)
  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 6972)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6972
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(11..28)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     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(941..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"
     protein_bind    complement(3848..3881)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(3944..3977)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             complement(4015..4728)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4057..4078)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4406..4425
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4665..4686
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      5592..5601
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    5612..5645
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    5716..5749
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    5782..6370
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5835..5855)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6253..6264)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    6411..6618
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6788..6917
                     /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 cggcgaggcg 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 ctagcccata acttcgtata aagtatccta tacgaagtta tatcaaaata ggaagaccaa
     3901 tgcttcacca tcgacccgaa ttgccaagca tcaccatcga cccataactt cgtataatgt
     3961 atgctatacg aagttatact agaattctca ttacacctcg ttctcgtagc agaacttgta
     4021 cagctcgtcc atgccgagag tgatcccggc ggcggtcacg aactccagca ggaccatgtg
     4081 atcgcgcttc tcgttggggt ctttgctcag ggcggactgg gtgctcaggt agtggttgtc
     4141 gggcagcagc acggggccgt cgccgatggg ggtgttctgc tggtagtggt cggcgagctg
     4201 cacgctgccg tcctcgatgt tgtggcggat cttgaagttc accttgatgc cgttcttctg
     4261 cttgtcggcc atgatataga cgttgtggct gttgtagttg tactccagct tgtgccccag
     4321 gatgttgccg tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc
     4381 gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt
     4441 gcgctcctgg acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg
     4501 gtcggggtag cggctgaagc actgcacgcc gtaggtcagg gtggtcacga gggtgggcca
     4561 gggcacgggc agcttgccgg tggtgcaaat gaacttcagg gtcagcttgc cgtaggtggc
     4621 atcgccctcg ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc cgtccagctc
     4681 gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca cgttgatgtc
     4741 gatctggtcc agggggatgt actcgccctc gctggtgatc ctgctcttgg cggccgcctt
     4801 gctcactgct actaccgggc ggtcggcggc accggtatca gctccagctg aaggttcagg
     4861 tgcttctgta tcttgcagaa tggctcttga cctgagcagg ataaaaccga agcccacctt
     4921 ggcggtgacg tccagcacca tgaacagcaa agtttcgatt cccaggccca caactccggc
     4981 accctcggtg ccgataatcc agatgatagg gtaagcagtc cacaggacca gaaccagtgc
     5041 ggtgaggatg ttgaaggttg aggcaacctc tgggcctctc tccttagcgg ctgaccgcag
     5101 agaagtggcc aggacataca gaacgacgat catacagata gttgagaaca gccaccaaga
     5161 gtatctggca atggcagtat gtgacagtgt tcctatcagg cctgtgacaa tcatcagagc
     5221 atcgacgcca acgagagtgc cgatggtcac tctatcaact ttggccagga gtgccaggtg
     5281 cagcagcaac agtggtgtag tgaacagcca ctgtgcgtat cttgcgtagt agatgttcag
     5341 catctctcca cctactggca cctctgtgag gccaatacca aagaacattg acagataagc
     5401 tgcactggca ataccgctga ccaggattgt cactgcgtag tactcccgtg catccttgtc
     5461 agtctctccc cacgctctaa ccaggaagta gaaggttccg atcagcatca gcagtgtacc
     5521 aattccgagc cacagacttt caggtctacc atctccgagg aggtcataac cagcctgcag
     5581 tgcgatagaa accatggtgg cggatcctag tataacttcg tataggatac tttatacgaa
     5641 gttatcattg ggattcttcc tattttgatc caagcatcac catcgaccct ctagtccaga
     5701 tctcaccatc gacccataac ttcgtatagc atacattata cgaagttatg tccctcgaag
     5761 aggttcgcga agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact
     5821 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     5881 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg
     5941 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg
     6001 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccggg
     6061 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc
     6121 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca
     6181 tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc
     6241 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct
     6301 ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc
     6361 gcctccccgc atcgataccg tcgacccggg cggccgctca taaagctcga ctgtgccttc
     6421 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     6481 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     6541 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     6601 tagcaggcat gctggggatg cggtccggac tgtacaattg cattcatttt atgtttcagg
     6661 ttcaggggga gatgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa
     6721 tcgataagga tcttcctaga gcatggctac gtagataagt agcatggcgg gttaatcatt
     6781 aactacaagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     6841 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     6901 agcgagcgag cgcgcagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     6961 gtattgggcg ct
//
