LOCUS       sequence-183932-        6752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CIB1 (full length, with endogenous NLS) fused to VP16 
            activation domain..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6752)
  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 6752)
  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-VP16 (B1014)
FEATURES             Location/Qualifiers
     source          1..6752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        7..386
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        387..590
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     541..561
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     635..654
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        635..653
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             746..1255
                     /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"
     regulatory      1766..1775
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1778..1798
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1799..2032
                     /codon_start=1
                     /gene="UL48"
                     /product="transcriptional activation domain of herpes 
                     simplex virus protein VP16 (Triezenberg et al., 1988; 
                     Cousens et al., 1989)"
                     /label=VP16 AD
                     /translation="APPTDVSLGDELHLDGEDVAMAHADALDDFDLDMLGDGDSPGPGF
                     TPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(2115..2132)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2121..2345
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2391..2819
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2478..2497)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2688..2709
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2828..2848)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2833..3162
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3013..3148
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3075..3094
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3229..4023
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3283..3302)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3893..3912
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4197..4318
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4234..4253)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4288..4307
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4367..4383)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4367..4383)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4380..4402)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4391..4407
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4415..4421,4422..4439,4440..4445))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4460..4481
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4598..4615)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4769..5357)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4849..4868)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5528..6319,6320..6388))
                     /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     6151..6170
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6389..6493)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6568..6587)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
       61 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      121 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      181 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      241 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      301 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      361 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      421 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      481 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      541 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa
      601 ctagagaacc cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc
      661 aagctggcta gcgtttaaac ttaagcttgg taccgagctc ggatccacta gtccagtgtg
      721 gtggaattct gcagatatcg ccaccatgaa tggagctata ggaggtgacc ttttgctcaa
      781 ttttcctgac atgtcggtcc tagagcgcca aagggctcac ctcaagtacc tcaatcccac
      841 ctttgattct cctctcgccg gcttctttgc cgattcttca atgattaccg gcggcgagat
      901 ggacagctat ctttcgactg ccggtttgaa tcttccgatg atgtacggtg agacgacggt
      961 ggaaggtgat tcaagactct caatttcgcc ggaaacgacg cttgggactg gaaatttcaa
     1021 gaaacggaag tttgatacag agactaagga ttgtaatgag aagaagaaga agatgacgat
     1081 gaacagagat gacctagtag aagaaggaga agaagagaag tcgaaaataa cagagcaaaa
     1141 caatgggagc acaaaaagca tcaagaagat gaaacacaaa gccaagaaag aagagaacaa
     1201 tttctctaat gattcatcta aagtgacgaa ggaattggag aaaacggatt atattcatgt
     1261 tcgtgcacga cgaggccaag ccactgatag tcacagcata gcagaacgag ttagaagaga
     1321 aaagatcagt gagagaatga agtttctaca agatttggtt cctggatgcg acaagatcac
     1381 aggcaaagca gggatgcttg atgaaatcat taactatgtt cagtctcttc agagacaaat
     1441 cgagttctta tcgatgaaac tagcaattgt gaatccaagg ccggattttg atatggatga
     1501 catttttgcc aaagaggttg cctcaactcc aatgactgtg gtgccatctc ctgaaatggt
     1561 tctttccggt tattctcatg agatggttca ctctggttat tctagtgaga tggttaactc
     1621 cggttacctt catgtcaatc caatgcagca agtgaatacc agttctgatc cattgtcatg
     1681 cttcaacaat ggcgaagctc cttcgatgtg ggactctcat gtgcagaatc tctatggcaa
     1741 tttaggagtt ggtggcggtg gtaccgccac catgggccct aaaaagaagc gtaaagtcgc
     1801 ccccccgacc gatgtcagcc tgggggacga gctccactta gacggcgagg acgtggcgat
     1861 ggcgcatgcc gacgcgctag acgatttcga tctggacatg ttgggggacg gggattcccc
     1921 gggtccggga tttacccccc acgactccgc cccctacggc gctctggata tggccgactt
     1981 cgagtttgag cagatgttta ccgatgccct tggaattgac gagtacggtg gggaattccc
     2041 ggggatccgt cgacgcgtct gcagaagctt cggcggcagc taatctagag ggcccgttta
     2101 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2161 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2221 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     2281 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     2341 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     2401 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     2461 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     2521 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     2581 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     2641 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     2701 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     2761 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     2821 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     2881 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     2941 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3001 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3061 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     3121 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     3181 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     3241 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3301 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     3361 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     3421 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     3481 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     3541 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     3601 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     3661 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     3721 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     3781 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     3841 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     3901 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     3961 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4021 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     4081 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     4141 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     4201 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4261 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4321 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     4381 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     4441 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     4501 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     4561 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4621 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     4681 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     4741 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     4801 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     4861 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     4921 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     4981 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5041 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5101 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5161 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5221 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5281 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5341 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5401 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5461 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     5521 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     5581 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     5641 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5701 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     5761 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5821 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     5881 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     5941 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6001 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6061 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6121 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6181 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6241 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6301 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6361 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6421 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     6481 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     6541 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     6601 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     6661 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
     6721 tgcgctgctt cgcgatgtac gggccagata ta
//
