LOCUS       Exported                6630 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  tLuc-mCherry in AAV backbone.
ACCESSION   .
VERSION     .
KEYWORDS    AAV-tLuc-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6630)
  AUTHORS   Liao HK, Hatanaka F, Araoka T, Reddy P, Wu MZ, Sui Y, Yamauchi T, 
            Sakurai M, O'Keefe DD, Nunez-Delicado E, Guillen P, Campistol JM, Wu
            CJ, Lu LF, Esteban CR, Izpisua Belmonte JC
  TITLE     In Vivo Target Gene Activation via CRISPR/Cas9-Mediated 
            Trans-epigenetic Modulation.
  JOURNAL   Cell. 2017 Dec 14;171(7):1495-1507.e15. doi: 
            10.1016/j.cell.2017.10.025. Epub 2017 Dec 7.
  PUBMED    29224783
REFERENCE   2  (bases 1 to 6630)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6630
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     primer_bind     263..287
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             331..1980
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     CDS             1993..2046
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2056..2757
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(2189..2207)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2457..2476
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2651..2674
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    2887..3008
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2924..2943)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2978..2997
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    3235..3823
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             complement(3706..3717)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3837..3885
                     /note="synthetic polyadenylation signal"
     repeat_region   3893..4033
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3893..4022
                     /label=AAV2 ITR
     rep_origin      4108..4563
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4195..4214)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4405..4426
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4580..4599)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4699..4721
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4759..4777)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4845..4949
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4950..5810
                     /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     complement(5168..5187)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5981..6569
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6470..6489
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggcctcta gaacgcgtcc tccacggcca ctagatcgat gccctttagg
      181 gataacaggg taatagtgtc ccctccaccc cacagtgcgg gaaactagtt aggcgtgtac
      241 ggtgggaggc ctatataagc agagctcgtt tagtgaaccg tcagatcgcc tggagaattc
      301 tgcagatatc aacaagtttg tacatccacc atggaagatg ccaaaaacat taagaagggc
      361 ccagcgccat tctacccact cgaagacggg accgccggcg agcagctgca caaagccatg
      421 aagcgctacg ccctggtgcc cggcaccatc gcctttaccg acgcacatat cgaggtggac
      481 attacctacg ccgagtactt cgagatgagc gttcggctgg cagaagctat gaagcgctat
      541 gggctgaata caaaccatcg gatcgtggtg tgcagcgaga atagcttgca gttcttcatg
      601 cccgtgttgg gtgccctgtt catcggtgtg gctgtggccc cagctaacga catctacaac
      661 gagcgcgagc tgctgaacag catgggcatc agccagccca ccgtcgtatt cgtgagcaag
      721 aaagggctgc aaaagatcct caacgtgcaa aagaagctac cgatcataca aaagatcatc
      781 atcatggata gcaagaccga ctaccagggc ttccaaagca tgtacacctt cgtgacttcc
      841 catttgccac ccggcttcaa cgagtacgac ttcgtgcccg agagcttcga ccgggacaaa
      901 accatcgccc tgatcatgaa cagtagtggc agtaccggat tgcccaaggg cgtagcccta
      961 ccgcaccgca ccgcttgtgt ccgattcagt catgcccgcg accccatctt cggcaaccag
     1021 atcatccccg acaccgctat cctcagcgtg gtgccatttc accacggctt cggcatgttc
     1081 accacgctgg gctacttgat ctgcggcttt cgggtcgtgc tcatgtaccg cttcgaggag
     1141 gagctattct tgcgcagctt gcaagactat aagattcaat ctgccctgct ggtgcccaca
     1201 ctatttagct tcttcgctaa gagcactctc atcgacaagt acgacctaag caacttgcac
     1261 gagatcgcca gcggcggggc gccgctcagc aaggaggtag gtgaggccgt ggccaaacgc
     1321 ttccacctac caggcatccg ccagggctac ggcctgacag aaacaaccag cgccattctg
     1381 atcacccccg aaggggacga caagcctggc gcagtaggca aggtggtgcc cttcttcgag
     1441 gctaaggtgg tggacttgga caccggtaag acactgggtg tgaaccagcg cggcgagctg
     1501 tgcgtccgtg gccccatgat catgagcggc tacgttaaca accccgaggc tacaaacgct
     1561 ctcatcgaca aggacggctg gctgcacagc ggcgacatcg cctactggga cgaggacgag
     1621 cacttcttca tcgtggaccg gctgaagagc ctgatcaaat acaagggcta ccaggtagcc
     1681 ccagccgaac tggagagcat cctgctgcaa caccccaaca tcttcgacgc cggggtcgcc
     1741 ggcctgcccg acgacgatgc cggcgagctg cccgccgcag tcgtcgtgct ggaacacggt
     1801 aaaaccatga ccgagaagga gatcgtggac tatgtggcca gccaggttac aaccgccaag
     1861 aagctgcgcg gtggtgttgt gttcgtggac gaggtgccta aaggactgac cggcaagttg
     1921 gacgcccgca agatccgcga gattctcatt aaggccaaga agggcggcaa gatcgccgtg
     1981 ttagcggccg ctgagggcag aggaagtctt ctaacatgcg gtgacgtgga ggagaatccc
     2041 ggcccttccg gaaccatggt gagcaagggc gaggaggtca tcaaagagtt catgcgcttc
     2101 aaggtgcgca tggagggctc catgaacggc cacgagttcg agatcgaggg cgagggcgag
     2161 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg cggccccctg
     2221 cccttcgcct gggacatcct gtccccccag ttcatgtacg gctccaaggc gtacgtgaag
     2281 caccccgccg acatccccga ttacaagaag ctgtccttcc ccgagggctt caagtgggag
     2341 cgcgtgatga acttcgagga cggcggtctg gtgaccgtga cccaggactc ctccctgcag
     2401 gacggcacgc tgatctacaa ggtgaagatg cgcggcacca acttcccccc cgacggcccc
     2461 gtaatgcaga agaagaccat gggctgggag gcctccaccg agcgcctgta cccccgcgac
     2521 ggcgtgctga agggcgagat ccaccaggcc ctgaagctga aggacggcgg ccactacctg
     2581 gtggagttca agaccatcta catggccaag aagcccgtgc aactgcccgg ctactactac
     2641 gtggacacca agctggacat cacctcccac aacgaggact acaccatcgt ggaacagtac
     2701 gagcgctccg agggccgcca ccacctgttc ctgtacggca tggacgagct gtacaagtaa
     2761 tgtacaagtg aaatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     2821 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcgt
     2881 taactaaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     2941 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     3001 gtatcttatc atgtctggaa ttgactcaaa tgatgtcaat tagtctatca gaagctcatc
     3061 tggtctccct tccgggggac aagacatccc tgtttaatat ttaaacagca gtgttcccaa
     3121 actgggttct tatatccctt gctctggtca accaggttgc agggtttcct gtcctcacag
     3181 gaacgaagtc cctaaagaaa cagtggcagc caggtttagc cctagtcata tgataatcaa
     3241 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt
     3301 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct
     3361 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc
     3421 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg
     3481 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc
     3541 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
     3601 actgacaatt ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct gctcgcctgt
     3661 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca
     3721 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt
     3781 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcacaattg gaattcaata
     3841 aaagatcttt attttcatta gatctgtgtg ttggtttttt gtgtgcggcc gcaggaaccc
     3901 ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga
     3961 ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc
     4021 agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     4081 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     4141 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt
     4201 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4261 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     4321 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4381 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact
     4441 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa
     4501 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     4561 ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     4621 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     4681 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     4741 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     4801 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     4861 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     4921 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     4981 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5041 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     5101 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     5161 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     5221 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     5281 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     5341 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     5401 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     5461 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     5521 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     5581 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     5641 ataaatctgg agccggtgag cgtggaagcc gcggtatcat tgcagcactg gggccagatg
     5701 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     5761 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     5821 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     5881 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     5941 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6001 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6061 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6121 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     6181 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     6241 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     6301 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     6361 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     6421 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     6481 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     6541 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     6601 tggccttttg ctggcctttt gctcacatgt
//
