LOCUS       sequence-350882-        6012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CIBN-EGFP-KRasCT in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6012)
  AUTHORS   Yamamoto K, Miura H, Ishida M, Mii Y, Kinoshita N, Takada S, Ueno N,
            Sawai S, Kondo Y, Aoki K
  TITLE     Optogenetic relaxation of actomyosin contractility uncovers 
            mechanistic roles of cortical tension during cytokinesis.
  JOURNAL   Nat Commun. 2021 Dec 8;12(1):7145. doi: 10.1038/s41467-021-27458-3.
  PUBMED    34880255
REFERENCE   2  (bases 1 to 6012)
  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: "Nat 
            Commun."; date: "2021-12-8"; volume: "12(1):7145. doi"; pages: " 
            10.1038/s41467-021-27458-3"
COMMENT     Sequence Label: pCAGGS-CIBN-EGFP-KRasCT
FEATURES             Location/Qualifiers
     source          1..6012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          666..1686
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1606..1628
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1694..1713
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1744..2253
                     /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
                     MITGGEMDSYLSTAGLNLPMMYGETTVEGDSRLSISPETTLGTGNFKAAKFDTETKDCN
                     EAAKKMTMNRDDLVEEGEEEKSKITEQNNGSTKSIKKMKHKAKKEENNFSNDSSKVTKE
                     LEKTDYI"
     CDS             join(2272..2274,2275..2277,2278..2790)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(2317..2338)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2578..2597)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2925..2946
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3034..3053)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3099..3154
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3153..3172)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3515..3531)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3515..3531)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3528..3550)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3539..3555
                     /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(3563..3569,3570..3587,3588..3593))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3608..3629
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3688..3883
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3734..3869
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3889..4023
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3926..3945)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3980..3999
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4091..4108)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4262..4850)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4342..4361)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5021..5812,5813..5881))
                     /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     5644..5663
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5882..5986)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgatcga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
      421 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      481 tgcagcgatg ggggcggggg gggggggggs gcgcgccrgg sggggsgggg sggggsgrgg
      541 ggsggggsgg ggsgrggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa
      601 agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc
      661 gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc
      721 gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg gacggccctt
      781 ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt ctgtggctgc
      841 gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc tcggggggtg
      901 cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg
      961 agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg
     1021 ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg
     1081 gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg caaccccccc
     1141 tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg ctccgtacgg
     1201 ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg gtgccgggcg
     1261 gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg
     1321 ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa tcgtgcgaga
     1381 gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc
     1441 gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg
     1501 gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct
     1561 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1621 gtgtgaccgg cggctctagc tagagcctct gctaaccatg ttcatgcctt cttctttttc
     1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattc
     1741 accatgaatg gagctatagg aggtgacctt ttgctcaatt ttcctgacat gtcggtccta
     1801 gagcgccaaa gggctcacct caagtacctc aatcccacct ttgattctcc tctcgccggc
     1861 ttctttgccg attcttcaat gattaccggc ggcgagatgg acagctatct ttcgactgcc
     1921 ggtttgaatc ttccgatgat gtacggtgag acgacggtgg aaggtgattc aagactctca
     1981 atttcgccgg aaacgacgct tgggactgga aatttcaagg cagcgaagtt tgatacagag
     2041 actaaggatt gtaatgaggc ggcgaagaag atgacgatga acagagatga cctagtagaa
     2101 gaaggagaag aagagaagtc gaaaataaca gagcaaaaca atgggagcac aaaaagcatc
     2161 aagaagatga aacacaaagc caagaaagaa gagaacaatt tctctaatga ttcatctaaa
     2221 gtgacgaagg aattggagaa aacggattat attcatctcg agcgcggccg catggtgagc
     2281 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     2341 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     2401 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     2461 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
     2521 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     2581 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     2641 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     2701 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
     2761 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
     2821 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     2881 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     2941 ttcgtgaccg ccgcctctag aaagatgagc aaagatggta aaaagaagaa aaagaagtca
     3001 aagacaaagt gtgtaattat gtaagtcgac ggatcttttt ccctctgcca aaaattatgg
     3061 ggacatcatg aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat
     3121 tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa
     3181 tcatttaaaa catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct
     3241 ggctgccatg aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc
     3301 tgtccattcc ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg
     3361 ttttgtgtta tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag
     3421 atttttcctc ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc
     3481 cctcgacctg cagcccaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     3541 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     3601 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     3661 cgggaaacct gtcgtgccag cggatccgca tctcaattag tcagcaacca tagtcccgcc
     3721 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     3781 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     3841 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctaa cttgtttatt
     3901 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     3961 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     4021 atccgctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     4081 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     4141 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     4201 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     4261 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     4321 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     4381 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     4441 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     4501 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     4561 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     4621 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     4681 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     4741 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     4801 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4861 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     4921 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     4981 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     5041 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5101 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     5161 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     5221 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5281 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     5341 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5401 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     5461 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     5521 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     5581 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     5641 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     5701 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     5761 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     5821 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     5881 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     5941 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6001 aagtgccacc tg
//
