LOCUS       sequence-156883-        8730 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes DMNT3 under LITE2.0 system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8730)
  AUTHORS   Lo CL, Choudhury SR, Irudayaraj J, Zhou FC
  TITLE     Epigenetic Editing of Ascl1 Gene in Neural Stem Cells by 
            Optogenetics.
  JOURNAL   Sci Rep. 2017 Feb 9;7:42047. doi: 10.1038/srep42047.
  PUBMED    28181538
REFERENCE   2  (bases 1 to 8730)
  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: "Sci Rep."; 
            date: "2017-02-9"; volume: "7:42047. doi"; pages: " 
            10.1038/srep42047"
COMMENT     Sequence Label: DNMT3ACD-CRY2-EGFP
FEATURES             Location/Qualifiers
     source          1..8730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             2298..2345
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             3846..3866
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3891..4040
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             4062..4115
                     /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             join(4116..4118,4119..4832)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(4116..4118,4119..4829)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4158..4179)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4419..4438)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4766..4787
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    4857..5445
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4910..4930)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5328..5339)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5477..5953
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(5676..5695)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5993..6133
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5993..6122
                     /label=AAV2 ITR
     rep_origin      6208..6663
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6295..6314)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6505..6526
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6680..6699)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6799..6821
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6859..6877)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6945..7049
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7050..7118,7119..7910)
                     /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(7268..7287)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8081..8669
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8570..8589
                     /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 gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccgccac catgccagct gagaagagga
     1441 agcccatccg cgtgctgtct ctctttgatg ggattgctac agggctcctg gtgctgaagg
     1501 acctgggcat ccaagtggac cgctacattg cctccgaggt gtgtgaggac tccatcacgg
     1561 tgggcatggt gcggcaccag ggaaagatca tgtacgtcgg ggacgtccgc agcgtcacac
     1621 agaagcatat ccaggagtgg ggcccattcg acctggtgat tggaggcagt ccctgcaatg
     1681 acctctccat tgtcaaccct gcccgcaagg gactttatga gggtactggc cgcctcttct
     1741 ttgagttcta ccgcctcctg catgatgcgc ggcccaagga gggagatgat cgccccttct
     1801 tctggctctt tgagaatgtg gtggccatgg gcgttagtga caagagggac atctcgcgat
     1861 ttcttgagtc taaccccgtg atgattgacg ccaaagaagt gtctgctgca cacagggccc
     1921 gttacttctg gggtaacctt cctggcatga acaggccttt ggcatccact gtgaatgata
     1981 agctggagct gcaagagtgt ctggagcacg gcagaatagc caagttcagc aaagtgagga
     2041 ccattaccac caggtcaaac tctataaagc agggcaaaga ccagcatttc cccgtcttca
     2101 tgaacgagaa ggaggacatc ctgtggtgca ctgaaatgga aagggtgttt ggcttccccg
     2161 tccactacac agacgtctcc aacatgagcc gcttggcgag gcagagactg ctgggccgat
     2221 cgtggagcgt gccggtcatc cgccacctct tcgctccgct gaaggaatat tttgcttgtg
     2281 tgggatccgc caccatgaaa aggccggcgg ccacgaaaaa ggccggccag gcaaaaaaga
     2341 aaaagaagat ggacaaaaag acgattgtgt ggtttagacg tgaccttcgc attgaggaca
     2401 accctgcgtt ggcagcggca gcgcatgaag gttcggtatt ccctgtcttt atctggtgtc
     2461 ccgaagagga ggggcaattc tatcccggtc gcgcgtcccg atggtggatg aaacagtcgc
     2521 tcgcacatct tagccagtcg ttgaaggcgc tcggctctga tctgacgctc atcaaaacac
     2581 acaatacaat ttctgccatt ctggattgca ttagggtgac tggtgccacc aaagttgtgt
     2641 tcaatcacct ttatgatccc gtaagcctgg tcagggatca tacagtaaaa gagaaacttg
     2701 tagagagggg aatcagcgtc caatcgtaca atggggactt gttgtatgaa ccgtgggaaa
     2761 tctattgtga gaaaggcaaa cctttcactt ccttcaattc gtactggaag aaatgcctcg
     2821 acatgtccat cgagtcagtc atgctgccgc caccttggag acttatgccc attactgctg
     2881 cggcagaggc gatctgggct tgcagcatcg aagaacttgg tctggaaaac gaagcggaaa
     2941 aacccagtaa cgcgctcctg acgcgagcgt ggtcgccagg ctggtcgaat gctgataagc
     3001 tccttaatga gtttatcgag aaacagttga tcgactatgc gaagaatagc aagaaggtgg
     3061 tgggaaattc cacttccctc ctgtcgccgt atctccactt tggggagatt agcgtgaggc
     3121 atgtgttcca atgtgcgcgg atgaagcaaa tcatttgggc tcgcgacaaa aactctgaag
     3181 gagaggagtc tgccgatttg tttttgagag gaatcggcct tagggagtat tcacgataca
     3241 tctgcttcaa ctttccgttt acacatgaac aatccctcct cagccacctc cgcttctttc
     3301 cgtgggacgc ggacgtggac aaattcaagg cctggcggca ggggcgcacg ggatacccgt
     3361 tggtggatgc gggtatgaga gagctgtggg caacgggttg gatgcacaac cgtattcgag
     3421 tcatcgttag tagctttgcc gttaagttcc tcttgttgcc ctggaaatgg gggatgaaat
     3481 acttttggga cacgcttctg gatgcggact tggagtgcga tattcttggg tggcagtaca
     3541 tttcgggttc catccctgac gggcatgaac ttgaccgatt ggataaccct gcactgcaag
     3601 gagcaaagta cgacccggag ggagaataca tcaggcagtg gctcccagag ctggcccgtc
     3661 tgcccaccga gtggattcac cacccgtggg atgctcccct gaccgtcctt aaagcctcag
     3721 gggtcgaatt ggggacgaat tacgcgaaac ccatcgtcga tattgacacc gcacgagaat
     3781 tgctggcgaa agccatctca agaacacgcg aagcgcagat tatgatcgga gcagcgccgg
     3841 ctagccccaa gaagaagaga aaggtggagg ccagcggttc cggacgggct gacgcattgg
     3901 acgattttga tctggatatg ctgggaagtg acgccctcga tgattttgac cttgacatgc
     3961 ttggttcgga tgcccttgat gactttgacc tcgacatgct cggcagtgac gcccttgatg
     4021 atttcgacct ggacatgctg attaactcta gaggcagtgg agagggcaga ggaagtctgc
     4081 taacatgcgg tgacgtcgag gagaatcctg gcccagtgag caagggcgag gagctgttca
     4141 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     4201 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     4261 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     4321 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     4381 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     4441 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     4501 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     4561 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     4621 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     4681 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     4741 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     4801 tcactctcgg catggacgag ctgtacaagt aagaattcga tatcaagctt atcgataatc
     4861 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     4921 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     4981 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
     5041 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
     5101 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
     5161 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     5221 gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct
     5281 gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
     5341 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc
     5401 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgagcgc
     5461 tgctcgagag atctacgggt ggcatccctg tgacccctcc ccagtgcctc tcctggccct
     5521 ggaagttgcc actccagtgc ccaccagcct tgtcctaata aaattaagtt gcatcatttt
     5581 gtctgactag gtgtccttct ataatattat ggggtggagg ggggtggtat ggagcaaggg
     5641 gcaagttggg aagacaacct gtagggcctg cggggtctat tgggaaccaa gctggagtgc
     5701 agtggcacaa tcttggctca ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc
     5761 tcagcctccc gagttgttgg gattccaggc atgcatgacc aggctcagct aatttttgtt
     5821 tttttggtag agacggggtt tcaccatatt ggccaggctg gtctccaact cctaatctca
     5881 ggtgatctac ccaccttggc ctcccaaatt gctgggatta caggcgtgaa ccactgctcc
     5941 cttccctgtc cttctgattt tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc
     6001 ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga
     6061 ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc
     6121 agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     6181 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     6241 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     6301 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     6361 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     6421 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     6481 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     6541 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     6601 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     6661 ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     6721 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     6781 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     6841 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     6901 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     6961 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     7021 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     7081 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     7141 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     7201 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     7261 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     7321 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     7381 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     7441 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     7501 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     7561 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     7621 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     7681 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     7741 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     7801 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     7861 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     7921 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     7981 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     8041 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     8101 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     8161 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     8221 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     8281 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     8341 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     8401 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     8461 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     8521 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     8581 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     8641 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     8701 tggccttttg ctggcctttt gctcacatgt
//
