LOCUS       sequence-203375-        5205 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of your protein of interest in fusion with HA at the 
            C-terminus. The tag is cleavable by TEV protease..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5205)
  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 5205)
  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-TEV-HA
FEATURES             Location/Qualifiers
     source          1..5205
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(38..57)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        229..608
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        609..812
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     763..783
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    814..832
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    835..853
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     863..887
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             996..1016
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1057..1077
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1084..1110
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1151..1168)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1157..1381
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1514..1533)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    1665..1712
                     /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             1720..2742
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             1720..2742
                     /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    2872..2993
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2909..2928)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2963..2982
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3042..3058)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3042..3058)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3055..3077)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3066..3082
                     /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(3090..3096,3097..3114,3115..3120))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3135..3156
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3273..3290)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3444..4032)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3524..3543)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4203..4994,4995..5063))
                     /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     4826..4845
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5064..5168)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
       61 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
      121 aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg
      181 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
      241 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
      301 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
      361 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      421 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      481 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
      541 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      601 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
      661 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
      721 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
      781 gtgtacggtg ggaggtctat ataagcagag ctctccctat cagtgataga gatctcccta
      841 tcagtgatag agatcgtcga cgagctcgtt tagtgaaccg tcagatcgcc tggagacgcc
      901 atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc cggactctgg
      961 cgtttaaact taagcttggt accgagctcg gatccgaaaa cctgtacttc caaggaaccg
     1021 gtggtcacct gtacaatcga tcctgcagga gctagcgaga atttgtattt tcagggtgat
     1081 atctacccct acgacgtgcc cgactacgct tgactcgagt ctagagggcc cgtttaaacc
     1141 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     1201 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     1261 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     1321 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     1381 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     1441 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     1501 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     1561 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     1621 ctcgacccca aaaaacttga ttagggtgat ggttcacgta cctagaagtt cctattccga
     1681 agttcctatt ctctagaaag tataggaact tccttggcca aaaagcctga actcaccgcg
     1741 acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg tctccgacct gatgcagctc
     1801 tcggagggcg aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg
     1861 cgggtaaata gctgcgccga tggtttctac aaagatcgtt atgtttatcg gcactttgca
     1921 tcggccgcgc tcccgattcc ggaagtgctt gacattgggg aattcagcga gagcctgacc
     1981 tattgcatct cccgccgtgc acagggtgtc acgttgcaag acctgcctga aaccgaactg
     2041 cccgctgttc tgcagccggt cgcggaggcc atggatgcga tcgctgcggc cgatcttagc
     2101 cagacgagcg ggttcggccc attcggaccg caaggaatcg gtcaatacac tacatggcgt
     2161 gatttcatat gcgcgattgc tgatccccat gtgtatcact ggcaaactgt gatggacgac
     2221 accgtcagtg cgtccgtcgc gcaggctctc gatgagctga tgctttgggc cgaggactgc
     2281 cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca acaatgtcct gacggacaat
     2341 ggccgcataa cagcggtcat tgactggagc gaggcgatgt tcggggattc ccaatacgag
     2401 gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta tggagcagca gacgcgctac
     2461 ttcgagcgga ggcatccgga gcttgcagga tcgccgcggc tccgggcgta tatgctccgc
     2521 attggtcttg accaactcta tcagagcttg gttgacggca atttcgatga tgcagcttgg
     2581 gcgcagggtc gatgcgacgc aatcgtccga tccggagccg ggactgtcgg gcgtacacaa
     2641 atcgcccgca gaagcgcggc cgtctggacc gatggctgtg tagaagtact cgccgatagt
     2701 ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat agcacgtact acgagatttc
     2761 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     2821 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     2881 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     2941 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     3001 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
     3061 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     3121 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     3181 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     3241 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     3301 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3361 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3421 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3481 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3541 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3601 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3661 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3721 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3781 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3841 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     3901 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3961 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4021 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     4081 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     4141 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     4201 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4261 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4321 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     4381 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4441 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4501 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4561 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4621 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4681 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4741 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4801 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4861 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4921 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4981 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     5041 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     5101 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     5161 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acgga
//
