LOCUS       sequence-203371-        5888 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of your protein of interest in fusion with orange 
            fluorescent protein at the N-terminus (cleavable by TEV) (PMID: 
            27240196). CyOFP1 is useful for multi-channel imaging..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5888)
  AUTHORS   Szczesny RJ, Kowalska K, Klosowska-Kosicka K, Chlebowski A, Owczarek
            EP, Warkocki Z, Kulinski TM, Adamska D, Affek K, Jedroszkowiak A, 
            Kotrys AV, Tomecki R, Krawczyk PS, Borowski LS, Dziembowski A
  TITLE     Versatile approach for functional analysis of human proteins and 
            efficient stable cell line generation using FLP-mediated 
            recombination system.
  JOURNAL   PLoS One. 2018 Mar 28;13(3):e0194887. doi: 
            10.1371/journal.pone.0194887. eCollection 2018.
  PUBMED    29590189
REFERENCE   2  (bases 1 to 5888)
  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: "PLoS One.";
            date: "2018-03-28"; volume: "13(3):e0194887. doi"; pages: " 
            10.1371/journal.pone.0194887. eCollection 2018"
COMMENT     Sequence Label: pKK-CyOFP1-TEV
FEATURES             Location/Qualifiers
     source          1..5888
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(42..61)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        233..612
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        613..816
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     767..787
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    818..836
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    839..857
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     867..891
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             988..1689
                     /codon_start=1
                     /product="orange-red fluorescent protein that is excitable 
                     by cyan light and suitable as an acceptor for blue-emitting
                     luciferase (Chu et al., 2016)"
                     /label=CyOFP1
                     /translation="MVSKGEELIKENMRSKLYLEGSVNGHQFKCTHEGEGKPYEGKQTN
                     RIKVVEGGPLPFAFDILATHFMYGSKVFIKYPADLPDYFKQSFPEGFTWERVMVFEDGG
                     VLTATQDTSLQDGELIYNVKVRGVNFPANGPVMQKKTLGWEPSTETMYPADGGLEGRCD
                     KALKLVGGGHLHVNFKTTYKSKKPVKMPGVHYVDRRLERIKEADNETYVEQYEHAVARY
                     SNLGGGMDELYK"
     CDS             1696..1716
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1757..1777
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(1838..1855)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1844..2068
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2201..2220)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2352..2399
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2407..3429
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2407..3429
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    3559..3680
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3596..3615)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3650..3669
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3729..3745)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3729..3745)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3742..3764)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3753..3769
                     /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(3777..3783,3784..3801,3802..3807))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3822..3843
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3960..3977)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4131..4719)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4211..4230)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4890..5681,5682..5750))
                     /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     5513..5532
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5751..5855)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
       61 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
      121 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
      181 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga
      241 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
      301 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
      361 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      421 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      481 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      541 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      601 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      661 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      721 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      781 aggcgtgtac ggtgggaggt ctatataagc agagctctcc ctatcagtga tagagatctc
      841 cctatcagtg atagagatcg tcgacgagct cgtttagtga accgtcagat cgcctggaga
      901 cgccatccac gctgttttga cctccataga agacaccggg accgatccag cctccggact
      961 ctggcgttta aacttaagct tggtaccatg gtgagcaagg gcgaggagct gatcaaggag
     1021 aacatgagaa gcaagctgta cctggaaggc agcgtgaacg gccaccagtt caagtgcacc
     1081 cacgaagggg agggcaagcc ctacgagggc aagcagacca acaggatcaa ggtggtggag
     1141 ggaggccccc tgccgttcgc attcgacatc ctggccaccc actttatgta cgggagcaag
     1201 gtgttcatca agtaccccgc cgacctcccc gattatttta agcagtcctt ccctgagggc
     1261 ttcacatggg agagagtcat ggtgttcgaa gacgggggcg tgctgaccgc cacccaggac
     1321 accagcctcc aggacggcga gctcatctac aacgtcaagg tcagaggggt gaacttccca
     1381 gccaacggcc ccgtgatgca gaagaaaaca ctgggctggg agcccagcac cgagaccatg
     1441 taccccgctg acggcggcct ggaaggcaga tgcgacaagg ccctgaagct cgtgggcggg
     1501 ggccacctgc acgtcaactt caagaccaca tacaagtcca agaaacccgt gaagatgccc
     1561 ggcgtccact acgtggaccg cagactggaa agaatcaagg aggccgacaa cgagacctac
     1621 gtcgagcagt acgagcacgc tgtggccaga tactccaacc tgggcggagg catggacgag
     1681 ctgtacaagg gatccgaaaa cctgtacttc caaggaaccg gtggtcacct gtacaatcga
     1741 tcctgcagga gctagcgaga atttgtattt tcagggtgat atccagcaca gtggcggccg
     1801 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     1861 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     1921 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     1981 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     2041 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     2101 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     2161 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     2221 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     2281 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     2341 tcacgtacct agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc
     2401 ttggccaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc
     2461 gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc
     2521 gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa
     2581 gatcgttatg tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac
     2641 attggggaat tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg
     2701 ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg
     2761 gatgcgatcg ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa
     2821 ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg
     2881 tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat
     2941 gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc
     3001 ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag
     3061 gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg
     3121 gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg
     3181 ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt
     3241 gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc
     3301 ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat
     3361 ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca
     3421 aaggaatagc acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3481 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3541 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3601 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3661 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     3721 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3781 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3841 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3901 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3961 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4021 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4081 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4141 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4201 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4261 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4321 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4381 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4441 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4501 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4561 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4621 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4681 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4741 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4801 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4861 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4921 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4981 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5041 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5101 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5161 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5221 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5281 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5341 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5401 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5461 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5521 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5581 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5641 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5701 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5761 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5821 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5881 gacgtcga
//
