LOCUS       sequence-441418-        7259 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of MAC3-tagged CDK8.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7259)
  AUTHORS   Liu X, Salokas K, Keskitalo S, Martinez-Botia P, Varjosalo M
  TITLE     Analyzing Protein Interactions by MAC-Tag Approaches.
  JOURNAL   Methods Mol Biol. 2023;2690:281-297. doi: 
            10.1007/978-1-0716-3327-4_24.
  PUBMED    37450155
REFERENCE   2  (bases 1 to 7259)
  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: "Methods Mol
            Biol."; date: "2023"; volume: "2690:281-297. doi"; pages: " 
            10.1007/978-1-0716-3327-4_24"
COMMENT     Sequence Label: MAC3-CDK8
FEATURES             Location/Qualifiers
     source          1..7259
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    586..604
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    607..625
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     635..659
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1343..1369
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1400..1423
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1460..1483
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     protein_bind    1496..1520
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2915..2939)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2977..2994)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2983..3207
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3340..3359)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3491..3538
                     /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             3546..4568
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3546..4568
                     /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    4698..4819
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4735..4754)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4789..4808
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4868..4884)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4868..4884)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4881..4903)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4892..4908
                     /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(4916..4922,4923..4940,4941..4946))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4961..4982
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5099..5116)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5270..5858)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5350..5369)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6029..6820,6821..6889))
                     /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     6652..6671
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6890..6994)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7069..7088)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctccct atcagtgata
      601 gagatctccc tatcagtgat agagatcgtc gacgagctcg tttagtgaac cgtcagatcg
      661 cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc
      721 tccggactct agcgtttaaa cttaagcttg gtaccgagct cggatccact agtccagtgt
      781 ggtggaattc tgcagatatc cagcacagtg gcggccgctc gagaccatgt tcaagaacct
      841 gatctggctg aaggaggtgg acagcaccca ggagagactg aaggagtgga acgtgtccta
      901 cggcaccgcc ctggtggccg acagacagac caagggcaga ggcggcccgg gcagaaagtg
      961 gctgagccag gagggcggcc tgtacttcag cttcctgctg aaccccaagg agttcgagaa
     1021 cctgctgcag ctgcccctgg tgctgggcct gagcgtgagc gaggccctgg aggagatcac
     1081 cgagatcccc ttcagcctga agtggcccaa cgacgtgtac ttccaggaga agaaggtgag
     1141 cggcgtgctg tgcgagctga gcaaggacaa gctgatcgtg ggcatcggca tcaacgtgaa
     1201 ccagagagag atccccgagg agatcaagga cagagccacc accctgtacg agatcaccgg
     1261 caaggactgg gacagaaagg aggtgctgct gaaggtgctg aagagaatca gcgagaacct
     1321 gaagaagttc aaggagaagg actacccata cgatgttcct gactatgccg gtaccgagct
     1381 cggatccacc atggctagct ggagccaccc gcagttcgag aaaggtggag gttccggagg
     1441 tggatcggga ggtggatcgt ggagccaccc gcagttcgaa aaagcggccg atatcacaag
     1501 tttgtacaaa aaagcaggct ccaccatgga ctatgacttt aaagtaaagc tgagcagcga
     1561 gcgggagcgg gtcgaggacc tgtttgaata cgagggctgc aaagttggcc gaggcactta
     1621 tggtcacgtc tacaaagcca agaggaaaga tgggaaggat gataaagact atgctttaaa
     1681 acaaatagaa ggaactggga tctctatgtc ggcatgtaga gaaatagcat tacttcgaga
     1741 gcttaagcat ccaaacgtca tttctcttca aaaggtgttt ctgtctcatg ctgataggaa
     1801 ggtgtggctt ctgtttgact atgctgaaca tgacctctgg catataatca agtttcacag
     1861 agcttctaaa gcaaacaaga agccagttca gttacctcgg ggaatggtga agtcactatt
     1921 atatcagatc ctagatggta ttcactacct gcatgctaac tgggtgttgc acagagattt
     1981 gaaacctgct aatattttag ttatgggtga aggtcctgag cgaggaagag taaaaattgc
     2041 tgacatgggc tttgcccgat tatttaattc acctttgaag cctttagcag atttggatcc
     2101 agtggttgtt acattctggt accgagcccc tgaactactt cttggagcaa ggcattatac
     2161 caaagctatt gatatttggg ctatagggtg tatatttgca gaactactaa cgtcagaacc
     2221 aatatttcac tgtcgacaag aggacatcaa aactagtaat ccttatcacc atgaccagct
     2281 ggacagaata ttcaatgtaa tgggatttcc tgcagataaa gattgggaag atataaaaaa
     2341 gatgcctgaa cattcaacat taatgaaaga tttcagaaga aatacgtata ccaactgcag
     2401 ccttatcaag tatatggaaa aacataaagt taaaccagat agtaaagcat tccacttgct
     2461 tcagaagctg cttaccatgg acccaataaa gcgaattacc tcagaacagg ctatgcagga
     2521 cccctatttc ttagaagacc cacttcctac atcagacgtt tttgccggtt gtcaaatccc
     2581 ttacccaaaa cgagaatttt taacggaaga agaacctgat gacaaaggag acaaaaacca
     2641 gcagcagcag cagggcaata accacactaa tggaactggc cacccaggga atcaagacag
     2701 cagtcacaca cagggacccc cgttgaagaa agtgagagtt gttcctccta ccactacctc
     2761 aggtggactt atcatgacct cagactatca gcgttccaat ccacatgctg cctatcccaa
     2821 ccctggacca agcacatcac agccgcagag cagcatggga tactcagcta cctcccagca
     2881 gcctccacag tactcacatc agacacacgg tacgacccag ctttcttgta caaagtggtg
     2941 acgtaagcta ggggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc
     3001 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     3061 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     3121 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     3181 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     3241 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     3301 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     3361 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     3421 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3481 cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata ggaacttcct
     3541 tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg
     3601 acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg
     3661 atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag
     3721 atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca
     3781 ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt
     3841 tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggccatgg
     3901 atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag
     3961 gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt
     4021 atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg
     4081 agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg
     4141 gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg
     4201 cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg
     4261 cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc
     4321 cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg
     4381 acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg
     4441 gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg
     4501 gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt ccgagggcaa
     4561 aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4621 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4681 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     4741 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     4801 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     4861 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     4921 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     4981 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5041 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5101 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5161 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5221 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5281 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5341 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5401 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5461 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5521 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5581 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5641 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5701 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5761 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5821 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5881 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5941 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6001 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6061 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6121 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6181 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6241 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6301 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6361 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6421 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6481 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6541 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6601 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6661 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6721 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6781 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6841 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6901 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6961 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7021 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     7081 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     7141 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
     7201 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
