LOCUS       sequence-217062-        6070 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid that can be used with the Flp-In T-REx 
            system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6070)
  AUTHORS   Shelton SB, Shah NM, Abell NS, Devanathan SK, Mercado M, Xhemalce B
  TITLE     Crosstalk between the RNA Methylation and Histone-Binding Activities
            of MePCE Regulates P-TEFb Activation on Chromatin.
  JOURNAL   Cell Rep. 2018 Feb 6;22(6):1374-1383. doi: 
            10.1016/j.celrep.2018.01.028.
  PUBMED    29425494
REFERENCE   2  (bases 1 to 6070)
  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.celrep.2018.01"; journalName: "Cell Rep"; date: 
            "2018-02-6- 6"; volume: "22"; issue: "6"; pages: "1374-1383"
COMMENT     Sequence Label: pcDNA5/FRT/TO/intron/GFP
FEATURES             Location/Qualifiers
     source          1..6070
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        152..531
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        532..735
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     686..706
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    737..755
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    758..776
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     786..810
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          929..1061
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1044..1063
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1106..1124
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1167..1176
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1173..1175,1176..1178,1179..1889)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1173..1175,1176..1178,1179..1889)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1218..1239)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1479..1498)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1826..1847
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1939..1956)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1945..2169
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2302..2321)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2453..2500
                     /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             2508..3530
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2508..3530
                     /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    3660..3781
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3697..3716)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3751..3770
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3830..3846)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3830..3846)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3843..3865)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3854..3870
                     /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(3878..3884,3885..3902,3903..3908))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3923..3944
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4061..4078)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4232..4820)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4312..4331)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4991..5782,5783..5851))
                     /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     5614..5633
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5852..5956)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6031..6050)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca
       61 aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg
      121 cttcgcgatg tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag
      181 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
      241 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      301 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      361 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
      421 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
      481 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      541 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      601 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      661 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      721 tatataagca gagctctccc tatcagtgat agagatctcc ctatcagtga tagagatcgt
      781 cgacgagctc gtttagtgaa ccgtcagatc gcctggagac gccatccacg ctgttttgac
      841 ctccatagaa gacaccggga ccgatccagc ctccggactc tagcgtttaa acttaagctg
      901 cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag gttacaagac aggtttaagg
      961 agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct gataggcacc
     1021 tattggtctt actgacatcc actttgcctt tctctccaca ggtgtccact cccagttcaa
     1081 ttacagctct taaggctaga gtacttaata cgactcacta taggctagcc tcgagaattc
     1141 acgcgtggta cctctagagt cgacccgcca ccatggtgag caagggcgag gagctgttca
     1201 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     1261 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     1321 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     1381 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     1441 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     1501 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     1561 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     1621 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     1681 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     1741 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     1801 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     1861 tcactctcgg catggacgag ctgtacaagc ccgggcggcc gctcgagtct agagggcccg
     1921 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     1981 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     2041 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     2101 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     2161 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc
     2221 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     2281 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     2341 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     2401 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtacc tagaagttcc
     2461 tattccgaag ttcctattct ctagaaagta taggaacttc cttggccaaa aagcctgaac
     2521 tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc tccgacctga
     2581 tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga gggcgtggat
     2641 atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa agatcgttat gtttatcggc
     2701 actttgcatc ggccgcgctc ccgattccgg aagtgcttga cattggggaa ttcagcgaga
     2761 gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa
     2821 ccgaactgcc cgctgttctg cagccggtcg cggaggccat ggatgcgatc gctgcggccg
     2881 atcttagcca gacgagcggg ttcggcccat tcggaccgca aggaatcggt caatacacta
     2941 catggcgtga tttcatatgc gcgattgctg atccccatgt gtatcactgg caaactgtga
     3001 tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg ctttgggccg
     3061 aggactgccc cgaagtccgg cacctcgtgc acgcggattt cggctccaac aatgtcctga
     3121 cggacaatgg ccgcataaca gcggtcattg actggagcga ggcgatgttc ggggattccc
     3181 aatacgaggt cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga
     3241 cgcgctactt cgagcggagg catccggagc ttgcaggatc gccgcggctc cgggcgtata
     3301 tgctccgcat tggtcttgac caactctatc agagcttggt tgacggcaat ttcgatgatg
     3361 cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc
     3421 gtacacaaat cgcccgcaga agcgcggccg tctggaccga tggctgtgta gaagtactcg
     3481 ccgatagtgg aaaccgacgc cccagcactc gtccgagggc aaaggaatag cacgtactac
     3541 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     3601 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     3661 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     3721 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     3781 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     3841 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     3901 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     3961 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     4021 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4081 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4141 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4201 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4261 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4321 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4381 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4441 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4501 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4561 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4621 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     4681 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4741 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4801 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4861 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     4921 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4981 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5041 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5101 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5161 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5221 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5281 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5341 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5401 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5461 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5521 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5581 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5641 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5701 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5761 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5821 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     5881 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     5941 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag
     6001 atctcccgat cccctatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     6061 ccagtatctg
//
