LOCUS       sequence-361474-        4081 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of humanized mKO2 in plant cell.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4081)
  AUTHORS   Fujii Y, Yoshimura A, Kodama Y
  TITLE     A novel orange-colored bimolecular fluorescence complementation 
            (BiFC) assay using monomeric Kusabira-Orange protein.
  JOURNAL   Biotechniques. 2018 Apr;64(4):153-161. doi: 10.2144/btn-2017-0121.
  PUBMED    29661017
REFERENCE   2  (bases 1 to 4081)
  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"; doi: "10"; journalName: 
            "Biotechniques"; date: "2018-04"; volume: "64"; issue: "4"; pages: 
            "153-161"
COMMENT     Sequence Label: p35S-hmKO2
FEATURES             Location/Qualifiers
     source          1..4081
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1..18)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(172..760)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(252..271)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(931..1722,1723..1791))
                     /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     1554..1573
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1792..1896)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1964..1982
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2020..2042)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2142..2161
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2355..2377
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2369..2386
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2370..2386
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2446..2791
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     2734..2754
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     misc_feature    2811..2866
                     /label=TMV Omega
                     /note="translational enhancer from the tobacco mosaic virus
                     5'-leader sequence (Gallie et al., 1988)"
     CDS             2886..3542
                     /codon_start=1
                     /product="monomeric Kusabira-Orange 2 fluorescent protein"
                     /label=mKO2
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     CDS             2886..3542
                     /codon_start=1
                     /product="humanized monomeric Kusabira-Orange fluorescent 
                     protein"
                     /label=hmKO
                     /note="mammalian codon-optimized"
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     terminator      3561..3813
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     primer_bind     complement(3851..3867)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3851..3867)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3864..3886)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3875..3891
                     /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(3899..3905,3906..3923,3924..3929))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3944..3965
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
       61 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
      121 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
      181 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
      241 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
      301 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
      361 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
      421 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
      481 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
      541 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
      601 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
      661 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
      721 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
      781 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
      841 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
      901 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
      961 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     1021 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     1081 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     1141 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     1201 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     1261 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     1321 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     1381 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     1441 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     1501 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     1561 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     1621 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     1681 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     1741 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     1801 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     1861 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     1921 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     1981 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     2041 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     2101 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     2161 actgagagtg caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     2221 catcaggcgc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     2281 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
     2341 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgcca agcttgcatg
     2401 cctgcaggct ctagaggatc ccccctcaga agaccagagg gctattgaga cttttcaaca
     2461 aagggtaata tcgggaaacc tcctcggatt ccattgccca gctatctgtc acttcatcga
     2521 aaggacagta gaaaaggaag gtggctccta caaatgccat cattgcgata aaggaaaggc
     2581 tatcgttcaa gatgcctcta ccgacagtgg tcccaaagat ggacccccac ccacgaggaa
     2641 catcgtggaa aaagaagacg ttccaaccac gtcttcaaag caagtggatt gatgtgatat
     2701 ctccactgac gtaagggatg acgcacaatc ccactatcct tcgcaagacc cttcctctat
     2761 ataaggaagt tcatttcatt tggagaggac aggcttcttg agatccttca acaattacca
     2821 acaacaacaa acaacaaaca acattacaat tactatttac aattacagtc gactctagag
     2881 gatccatggt gagcgtgatc aagcccgaga tgaagatgag gtactacatg gacggctccg
     2941 tcaatgggca tgagttcaca atcgagggtg agggcacagg cagaccttac gagggacatc
     3001 aggagatgac actgcgcgtc acaatggccg agggcgggcc aatgcctttc gccttcgacc
     3061 tggtgtccca cgtgttctgt tacggccaca gagtttttac caagtaccca gaagagatcc
     3121 cagactattt caagcaggcc tttcctgagg gcctgtcctg ggagaggtcc ctggagttcg
     3181 aggacggcgg ctccgcctcc gtgagcgccc acatcagcct gaggggcaac accttctacc
     3241 acaagtccaa gttcaccggc gtgaacttcc ccgccgacgg ccccatcatg cagaaccaga
     3301 gcgtggactg ggagccctcc accgagaaga tcaccgccag cgacggcgtg ctgaagggcg
     3361 acgtgaccat gtacctgaag ctggagggcg gcggcaacca caagtgccag atgaagacca
     3421 cctacaaggc cgccaaggag atcctggaga tgcccggcga ccactacatc ggccacaggc
     3481 tggtgaggaa gaccgagggc aacatcaccg agcaggtgga ggacgccgtg gcccactcct
     3541 aaagcggccg cccggctgca gatcgttcaa acatttggca ataaagtttc ttaagattga
     3601 atcctgttgc cggtcttgcg atgattatca tataatttct gttgaattac gttaagcatg
     3661 taataattaa catgtaatgc atgacgttat ttatgagatg ggtttttatg attagagtcc
     3721 cgcaattata catttaatac gcgatagaaa acaaaatata gcgcgcaaac taggataaat
     3781 tatcgcgcgc ggtgtcatct atgttactag atccgatgat aagctgtcaa acatgagaat
     3841 tcgtaatcat gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     3901 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     3961 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4021 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4081 c
//
