LOCUS       sequence-183935-        5200 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CIB1 (Full-length, with endogenous NLS) fused to VP64 
            activation domain (4xVP16 inserts).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5200)
  AUTHORS   Pathak GP, Spiltoir JI, Hoglund C, Polstein LR, Heine-Koskinen S, 
            Gersbach CA, Rossi J, Tucker CL
  TITLE     Bidirectional approaches for optogenetic regulation of gene 
            expression in mammalian cells using Arabidopsis cryptochrome 2.
  JOURNAL   Nucleic Acids Res. 2017 Apr 20. doi: 10.1093/nar/gkx260.
  PUBMED    28431041
REFERENCE   2  (bases 1 to 5200)
  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: "Nucleic 
            Acids Res. 2017 Apr 20. doi: 10.1093/nar/gkx260."
COMMENT     Sequence Label: CIB-VP64 (B1016)
FEATURES             Location/Qualifiers
     source          1..5200
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        58..361
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        362..565
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     516..536
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             625..1134
                     /codon_start=1
                     /product="N-terminal portion of the CIB1 transcription 
                     factor from Arabidopsis thaliana"
                     /label=CIBN
                     /note="binds to CRY2 (cryptochrome 2) that has absorbed 
                     blue light (Kennedy et al., 2010)"
                     /translation="MNGAIGGDLLLNFPDMSVLERQRAHLKYLNPTFDSPLAGFFADSS
                     MITGGEMDSYLSTAGLNLPMMYGETTVEGDSRLSISPETTLGTGNFKKRKFDTETKDCN
                     EKKKKMTMNRDDLVEEGEEEKSKITEQNNGSTKSIKKMKHKAKKEENNFSNDSSKVTKE
                     LEKTDYI"
     CDS             1663..1683
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1693..1842
                     /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"
     polyA_signal    1985..2106
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2022..2041)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2076..2095
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2113..2568)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2250..2271)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2462..2481
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2595..2699
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2701..3058
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2701..2892
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2724..2744)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2909..3044
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3093..3887
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3093..3887
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3147..3166)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3757..3776
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4075..4094)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4119..4166
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4495..5083
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4984..5003
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt
       61 tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac
      121 gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg
      181 ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag
      241 tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat
      301 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
      361 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
      421 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
      481 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
      541 gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct agcgctaccg
      601 gactcagatc tcgagctcgc caccatgaat ggagctatag gaggtgacct tttgctcaat
      661 tttcctgaca tgtcggtcct agagcgccaa agggctcacc tcaagtacct caatcccacc
      721 tttgattctc ctctcgccgg cttctttgcc gattcttcaa tgattaccgg cggcgagatg
      781 gacagctatc tttcgactgc cggtttgaat cttccgatga tgtacggtga gacgacggtg
      841 gaaggtgatt caagactctc aatttcgccg gaaacgacgc ttgggactgg aaatttcaag
      901 aaacggaagt ttgatacaga gactaaggat tgtaatgaga agaagaagaa gatgacgatg
      961 aacagagatg acctagtaga agaaggagaa gaagagaagt cgaaaataac agagcaaaac
     1021 aatgggagca caaaaagcat caagaagatg aaacacaaag ccaagaaaga agagaacaat
     1081 ttctctaatg attcatctaa agtgacgaag gaattggaga aaacggatta tattcatgtt
     1141 cgtgcacgac gaggccaagc cactgatagt cacagcatag cagaacgagt tagaagagaa
     1201 aagatcagtg agagaatgaa gtttctacaa gatttggttc ctggatgcga caagatcaca
     1261 ggcaaagcag ggatgcttga tgaaatcatt aactatgttc agtctcttca gagacaaatc
     1321 gagttcttat cgatgaaact agcaattgtg aatccaaggc cggattttga tatggatgac
     1381 atttttgcca aagaggttgc ctcaactcca atgactgtgg tgccatctcc tgaaatggtt
     1441 ctttccggtt attctcatga gatggttcac tctggttatt ctagtgagat ggttaactcc
     1501 ggttaccttc atgtcaatcc aatgcagcaa gtgaatacca gttctgatcc attgtcatgc
     1561 ttcaacaatg gcgaagctcc ttcgatgtgg gactctcatg tgcagaatct ctatggcaat
     1621 ttaggagttg gtggcggtgg ctctggaggt ggtgggtccg gaccgaaaaa gaaacgcaaa
     1681 gttgggcgcg ccgacgcgct ggacgatttc gatctcgaca tgctgggttc tgatgccctc
     1741 gatgactttg acctggatat gttgggaagc gacgcattgg atgactttga tctggacatg
     1801 ctcggctccg atgctctgga cgatttcgat ctcgatatgt taattaactg atgtacaagt
     1861 aaagcggccg cgactctaga tcataatcag ccataccaca tttgtagagg ttttacttgc
     1921 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
     1981 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     2041 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     2101 atcttaaggc gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt
     2161 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     2221 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     2281 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     2341 aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     2401 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     2461 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     2521 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt
     2581 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     2641 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc
     2701 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     2761 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     2821 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2881 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     2941 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     3001 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga
     3061 tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     3121 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     3181 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     3241 agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc
     3301 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     3361 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     3421 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     3481 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     3541 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     3601 tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg
     3661 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     3721 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     3781 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     3841 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     3901 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     3961 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     4021 ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac
     4081 acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa
     4141 aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc
     4201 tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc
     4261 accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg
     4321 ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta
     4381 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     4441 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     4501 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     4561 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     4621 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     4681 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4741 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4801 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4861 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4921 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4981 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     5041 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     5101 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     5161 ccctgattct gtggataacc gtattaccgc catgcattag
//
