LOCUS       sequence-463622-        6641 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression in Flp-IN human cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6641)
  AUTHORS   Lopez Martinez D, Todorovski I, Noe Gonzalez M, Rusimbi C, Blears D,
            Khallou N, Han Z, Dirac-Svejstrup AB, Svejstrup JQ
  TITLE     PAF1C-mediated activation of CDK12/13 kinase activity is critical 
            for CTD phosphorylation and transcript elongation.
  JOURNAL   Mol Cell. 2025 May 15;85(10):1952-1967.e8. doi: 
            10.1016/j.molcel.2025.04.012. Epub 2025 May 1.
  PUBMED    40315851
REFERENCE   2  (bases 1 to 6641)
  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: 
            "10.1016/j.molcel.2025.04.012"; journalName: "Mol Cell"; date: 
            "2025-05-15- 15"; volume: "85"; issue: "10"; pages: "1952-1967.e8"
COMMENT     Sequence Label: pFRT-TO-FLAG(N)-CDC73deltaKIM
FEATURES             Location/Qualifiers
     source          1..6641
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(112..131)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    263..310
                     /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             318..1340
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             318..1340
                     /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    1470..1591
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1507..1526)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1561..1580
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1640..1656)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1640..1656)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1653..1675)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1664..1680
                     /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(1688..1694,1695..1712,1713..1718))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1733..1754
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1871..1888)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2042..2630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2122..2141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2801..3592,3593..3661))
                     /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     3424..3443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3662..3766)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3841..3860)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4032..4411
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4412..4615
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4566..4586
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    4617..4635
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4638..4656
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     4666..4690
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4784..4807
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4814..4837
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(6390..6407)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6396..6620
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg
       61 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
      121 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
      181 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
      241 agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct ctagaaagta
      301 taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga
      361 tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg
      421 ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg
      481 gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg
      541 aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac
      601 agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg
      661 cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat
      721 tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg
      781 atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc
      841 aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc
      901 acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg
      961 actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga
     1021 ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc
     1081 ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc
     1141 agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa
     1201 tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg
     1261 tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc
     1321 gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg
     1381 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     1441 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     1501 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     1561 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     1621 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     1681 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     1741 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     1801 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     1861 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     1921 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     1981 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     2041 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     2101 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     2161 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     2221 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     2281 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     2341 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     2401 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     2461 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     2521 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     2581 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     2641 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     2701 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     2761 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     2821 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     2881 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     2941 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     3001 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     3061 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     3121 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     3181 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     3241 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     3301 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     3361 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     3421 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     3481 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     3541 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     3601 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     3661 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     3721 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     3781 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
     3841 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
     3901 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
     3961 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
     4021 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     4081 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     4141 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     4201 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     4261 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     4321 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     4381 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     4441 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     4501 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     4561 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc
     4621 tatcagtgat agagatctcc ctatcagtga tagagatcgt cgacgagctc gtttagtgaa
     4681 ccgtcagatc gcctggagac gccatccacg ctgttttgac ctccatagaa gacaccggga
     4741 ccgatccagc ctccggactc tagcgtttaa acttaagctt atggactaca aggacgacga
     4801 tgacaagctc gatcttgaag tcctctttca gggacccatg gcggacgtgc ttagcgtcct
     4861 gcgacagtac aacatccaga agaaggagat tgtggtgaag ggagacgaag tgatcttcgg
     4921 ggagttctcc tggcccaaga atgtgaagac caactatgtt gtttggggga ctggaaagga
     4981 aggccaaccc agagagtact acacattgga ttccatttta tttctactta ataacgtgca
     5041 cctttctcat cctgtttatg tccgacgtgc agctactgaa aatattcctg tggttagaag
     5101 acctgatcga aaagatctac ttggatatct caatggtgaa gcgtcaacat cggcaagtat
     5161 agacagaagc gctcccttag aaataggtct tcagcgatct actcaagtca aacgagctgc
     5221 agatgaagtt ttagcagaag caaagaaacc acgaattgag gatgaagagt gtgtgcgcct
     5281 tgataaagag agattggctg cccgtttgga gggtcacaaa gaagggattg ttcaaacgga
     5341 gcaaatcagg tctttgtctg aagctatgtc agtggaaaaa attgctgcaa tcaaagccaa
     5401 aattatggct aagaaaagat ctactatcaa gactgatcta gatgatgaca taactgccct
     5461 taaacagagg agttttgtgg atgctgaggt agatgtgacc cgagatattg tcagcagaga
     5521 gagagtatgg aggacacgaa caactatctt acaaagcaca ggaaagaatt tttccaagaa
     5581 catttttgca attcttcaat ctgtaaaagc cagagaagaa gggcgtgcac ctgaacagcg
     5641 acctgcccca aatgcagcac ctgtggatcc cactttgcgc accaaacagc ctatcccagc
     5701 tgccgagggt gcatctgccc ggaagactca gactcctgca gcccagccag taccaagacc
     5761 agtttctcaa gcaagacctc ccccaaatca gaagaaagga tctcgaacac ccattatcat
     5821 aattcctgca gctaccacct ctttaataac catgcttaat gcaaaagacc ttctacagga
     5881 cctgaaattt gtcccatcag atgaaaagaa gaaacaaggt tgtcaacgag aaaatgaaac
     5941 tctaatacaa agaagaaaag accagatgca accagggggc actgcaatta gtgttacagt
     6001 accttataga gtagtagacc agccccttaa acttatgcct caagactggg accgcgttgt
     6061 agccgttttt gtgcagggtc ctgcatggca gttcaaaggt tggccatggc ttttgcctga
     6121 tggatcacca gttgatatat ttgctaaaat taaagccttc catctgaagt atgatgaagt
     6181 tcgtctggat ccaaatgttc agaaatggga tgtaacagta ttagaactca gctatcacaa
     6241 acgtcatttg gatagaccag tgttcttacg gttttgggaa acattggaca ggtacatggt
     6301 aaaacacaag tcccatttaa ggttctgatg atatccagca cagtggcggc cgctcgagtc
     6361 tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     6421 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     6481 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     6541 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     6601 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc a
//
