LOCUS       sequence-183936-        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        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             628..1137
                     /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             1666..1686
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1696..1845
                     /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    1988..2109
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2025..2044)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2079..2098
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2116..2571)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2253..2274)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2465..2484
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2598..2702
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2704..3061
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2704..2895
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2727..2747)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2912..3047
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3096..3890
                     /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             3096..3890
                     /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(3150..3169)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3760..3779
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4078..4097)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4122..4169
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4498..5086
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4987..5006
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct cgccaccatg aatggagcta taggaggtga ccttttgctc
      661 aattttcctg acatgtcggt cctagagcgc caaagggctc acctcaagta cctcaatccc
      721 acctttgatt ctcctctcgc cggcttcttt gccgattctt caatgattac cggcggcgag
      781 atggacagct atctttcgac tgccggtttg aatcttccga tgatgtacgg tgagacgacg
      841 gtggaaggtg attcaagact ctcaatttcg ccggaaacga cgcttgggac tggaaatttc
      901 aagaaacgga agtttgatac agagactaag gattgtaatg agaagaagaa gaagatgacg
      961 atgaacagag atgacctagt agaagaagga gaagaagaga agtcgaaaat aacagagcaa
     1021 aacaatggga gcacaaaaag catcaagaag atgaaacaca aagccaagaa agaagagaac
     1081 aatttctcta atgattcatc taaagtgacg aaggaattgg agaaaacgga ttatattcat
     1141 gttcgtgcac gacgaggcca agccactgat agtcacagca tagcagaacg agttagaaga
     1201 gaaaagatca gtgagagaat gaagtttcta caagatttgg ttcctggatg cgacaagatc
     1261 acaggcaaag cagggatgct tgatgaaatc attaactatg ttcagtctct tcagagacaa
     1321 atcgagttct tatcgatgaa actagcaatt gtgaatccaa ggccggattt tgatatggat
     1381 gacatttttg ccaaagaggt tgcctcaact ccaatgactg tggtgccatc tcctgaaatg
     1441 gttctttccg gttattctca tgagatggtt cactctggtt attctagtga gatggttaac
     1501 tccggttacc ttcatgtcaa tccaatgcag caagtgaata ccagttctga tccattgtca
     1561 tgcttcaaca atggcgaagc tccttcgatg tgggactctc atgtgcagaa tctctatggc
     1621 aatttaggag ttggtggcgg tggctctgga ggtggtgggt ccggaccgaa aaagaaacgc
     1681 aaagttgggc gcgccgacgc gctggacgat ttcgatctcg acatgctggg ttctgatgcc
     1741 ctcgatgact ttgacctgga tatgttggga agcgacgcat tggatgactt tgatctggac
     1801 atgctcggct ccgatgctct ggacgatttc gatctcgata tgttaattaa ctgatgtaca
     1861 agtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag aggttttact
     1921 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     1981 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     2041 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     2101 tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     2161 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     2221 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     2281 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     2341 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     2401 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     2461 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     2521 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     2581 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     2641 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     2701 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2761 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2821 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2881 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     2941 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     3001 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3061 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     3121 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     3181 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     3241 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     3301 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     3361 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     3421 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     3481 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     3541 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     3601 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     3661 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     3721 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     3781 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     3841 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     3901 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     3961 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4021 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     4081 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     4141 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     4201 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     4261 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     4321 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     4381 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     4441 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     4501 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     4561 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     4621 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     4681 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     4741 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     4801 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     4861 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     4921 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     4981 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5041 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5101 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     5161 atcccctgat tctgtggata accgtattac cgccatgcat
//
