LOCUS       sequence-69627-e        6818 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LITE2.0 CRY2PHR-VP64. Changes from LITE1.0: N-terminal 
            alpha-importin nuclear localization sequence. Synapsin promoter for 
            neuronal expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6818)
  AUTHORS   Konermann S, Brigham MD, Trevino A, Hsu PD, Heidenreich M, Le Cong, 
            Platt RJ, Scott DA, Church GM, Zhang F
  TITLE     Optical control of mammalian endogenous transcription and epigenetic
            states.
  JOURNAL   Nature. 2013 Jul 23. doi: 10.1038/nature12466.
  PUBMED    23877069
REFERENCE   2  (bases 1 to 6818)
  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: "Nature. 
            2013 Jul 23. doi: 10.1038/nature12466."
COMMENT     Sequence Label: LITE2.0
            pAAV_hSyn_NLS(alpha-imp)-CRY2PHR-NLS-V...
FEATURES             Location/Qualifiers
     source          1..6818
                     /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        181..628
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             651..698
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             2199..2219
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2244..2393
                     /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             2415..2468
                     /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(2469..2471,2472..3185)
                     /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(2469..2471,2472..3182)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2511..2532)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2772..2791)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3119..3140
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3210..3798
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3263..3283)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3681..3692)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3840..3857)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3846..4053
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4082..4222
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4082..4211
                     /label=AAV2 ITR
     rep_origin      4296..4751
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4383..4402)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4593..4614
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4768..4787)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4887..4909
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4947..4965)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5033..5137
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5138..5206,5207..5998)
                     /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(5356..5375)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6169..6757
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6658..6677
                     /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 gcgttgtaca gtgtctagac tgcagagggc cctgcgtatg
      181 agtgcaagtg ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga
      241 ccccgaccca ctggacaagc acccaacccc cattccccaa attgcgcatc ccctatcaga
      301 gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg
      361 gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc
      421 gctgacgtca ctcgccggtc ccccgcaaac tccccttccc ggccaccttg gtcgcgtccg
      481 cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc gagatagggg ggcacgggcg
      541 cgaccatctg cgctgcggcg ccggcgactc agcgctgcct cagtctgcgg tgggcagcgg
      601 aggagtcgtg tcgtgcctga gagcgcagtc gagaaggatc cgccaccatg aaaaggccgg
      661 cggccacgaa aaaggccggc caggcaaaaa agaaaaagaa gatggacaaa aagacgattg
      721 tgtggtttag acgtgacctt cgcattgagg acaaccctgc gttggcagcg gcagcgcatg
      781 aaggttcggt attccctgtc tttatctggt gtcccgaaga ggaggggcaa ttctatcccg
      841 gtcgcgcgtc ccgatggtgg atgaaacagt cgctcgcaca tcttagccag tcgttgaagg
      901 cgctcggctc tgatctgacg ctcatcaaaa cacacaatac aatttctgcc attctggatt
      961 gcattagggt gactggtgcc accaaagttg tgttcaatca cctttatgat cccgtaagcc
     1021 tggtcaggga tcatacagta aaagagaaac ttgtagagag gggaatcagc gtccaatcgt
     1081 acaatgggga cttgttgtat gaaccgtggg aaatctattg tgagaaaggc aaacctttca
     1141 cttccttcaa ttcgtactgg aagaaatgcc tcgacatgtc catcgagtca gtcatgctgc
     1201 cgccaccttg gagacttatg cccattactg ctgcggcaga ggcgatctgg gcttgcagca
     1261 tcgaagaact tggtctggaa aacgaagcgg aaaaacccag taacgcgctc ctgacgcgag
     1321 cgtggtcgcc aggctggtcg aatgctgata agctccttaa tgagtttatc gagaaacagt
     1381 tgatcgacta tgcgaagaat agcaagaagg tggtgggaaa ttccacttcc ctcctgtcgc
     1441 cgtatctcca ctttggggag attagcgtga ggcatgtgtt ccaatgtgcg cggatgaagc
     1501 aaatcatttg ggctcgcgac aaaaactctg aaggagagga gtctgccgat ttgtttttga
     1561 gaggaatcgg ccttagggag tattcacgat acatctgctt caactttccg tttacacatg
     1621 aacaatccct cctcagccac ctccgcttct ttccgtggga cgcggacgtg gacaaattca
     1681 aggcctggcg gcaggggcgc acgggatacc cgttggtgga tgcgggtatg agagagctgt
     1741 gggcaacggg ttggatgcac aaccgtattc gagtcatcgt tagtagcttt gccgttaagt
     1801 tcctcttgtt gccctggaaa tgggggatga aatacttttg ggacacgctt ctggatgcgg
     1861 acttggagtg cgatattctt gggtggcagt acatttcggg ttccatccct gacgggcatg
     1921 aacttgaccg attggataac cctgcactgc aaggagcaaa gtacgacccg gagggagaat
     1981 acatcaggca gtggctccca gagctggccc gtctgcccac cgagtggatt caccacccgt
     2041 gggatgctcc cctgaccgtc cttaaagcct caggggtcga attggggacg aattacgcga
     2101 aacccatcgt cgatattgac accgcacgag aattgctggc gaaagccatc tcaagaacac
     2161 gcgaagcgca gattatgatc ggagcagcgc cggctagccc caagaagaag agaaaggtgg
     2221 aggccagcgg ttccggacgg gctgacgcat tggacgattt tgatctggat atgctgggaa
     2281 gtgacgccct cgatgatttt gaccttgaca tgcttggttc ggatgccctt gatgactttg
     2341 acctcgacat gctcggcagt gacgcccttg atgatttcga cctggacatg ctgattaact
     2401 ctagaggcag tggagagggc agaggaagtc tgctaacatg cggtgacgtc gaggagaatc
     2461 ctggcccagt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2521 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2581 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2641 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2701 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     2761 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     2821 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     2881 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     2941 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     3001 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3061 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     3121 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3181 agtaagaatt cgatatcaag cttatcgata atcaacctct ggattacaaa atttgtgaaa
     3241 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     3301 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     3361 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     3421 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     3481 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     3541 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     3601 ggaaatcatc gtcctttcct tggctgctcg cctatgttgc cacctggatt ctgcgcggga
     3661 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     3721 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     3781 tttgggccgc ctccccgcat cgataccgag cgctgctcga gctagagctc gctgatcagc
     3841 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     3901 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     3961 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     4021 ggattgggaa gacaatagca ggcatgctgg ggaggtaacc acgtgcggac cgagcggccg
     4081 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     4141 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     4201 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     4261 ggtattcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag
     4321 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     4381 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4441 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4501 aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     4561 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     4621 actcaaccct atctcgggct attcttttga tttataaggg attttgccga tttcggccta
     4681 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     4741 gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     4801 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     4861 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     4921 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     4981 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     5041 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     5101 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     5161 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     5221 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     5281 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     5341 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     5401 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     5461 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     5521 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     5581 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     5641 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     5701 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     5761 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     5821 tattgctgat aaatctggag ccggtgagcg tggaagccgc ggtatcattg cagcactggg
     5881 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5941 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     6001 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     6061 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     6121 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     6181 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     6241 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     6301 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     6361 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     6421 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     6481 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     6541 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     6601 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     6661 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     6721 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     6781 acggttcctg gccttttgct ggccttttgc tcacatgt
//
