LOCUS       sequence-269891-        6920 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  DOX-inducible G3BP1 with RNA binding domains deleted inserted with 
            GFP tagged on the N-terminus and used with the Flp-In T-Rex System.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6920)
  AUTHORS   Fischer JW, Busa VF, Shao Y, Leung AKL
  TITLE     Structure-Mediated RNA Decay by UPF1 and G3BP1.
  JOURNAL   Mol Cell. 2020 Apr 2;78(1):70-84.e6. doi: 
            10.1016/j.molcel.2020.01.021. Epub 2020 Feb 3.
  PUBMED    32017897
REFERENCE   2  (bases 1 to 6920)
  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.2020.01.021"; journalName: "Mol Cell"; date: 
            "2020-04-2- 2"; volume: "78"; issue: "1"; pages: "70-84.e6"
COMMENT     Sequence Label: pcDNA5 FRT TO-EGFP-G3BP1-DeltaRBP
FEATURES             Location/Qualifiers
     source          1..6920
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        139..518
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        519..722
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     673..693
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    724..742
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    745..763
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     regulatory      921..930
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(927..929,930..932,933..1643)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(927..929,930..932,933..1643)
                     /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(972..993)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1233..1252)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1580..1601
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2776..2793)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2782..3006
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3139..3158)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3290..3337
                     /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             3345..4367
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3345..4367
                     /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    4497..4618
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4534..4553)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4588..4607
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4667..4683)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4667..4683)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4680..4702)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4691..4707
                     /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(4715..4721,4722..4739,4740..4745))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4760..4781
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4898..4915)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5069..5657)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5149..5168)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5828..6619,6620..6688))
                     /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     6451..6470
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6689..6793)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6868..6887)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga
       61 ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac
      121 gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
      181 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
      241 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
      301 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      361 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      421 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      481 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
      541 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
      601 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
      661 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
      721 ctctccctat cagtgataga gatctcccta tcagtgatag agatcgtcga cgagctcgtt
      781 tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac
      841 accgggaccg atccagcctc cggactctag cgtttaaact taagcttagt gaaccgtcag
      901 atccgctagc gctaccggtc gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg
      961 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     1021 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     1081 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct
     1141 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     1201 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     1261 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     1321 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct
     1381 atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca
     1441 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     1501 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc
     1561 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     1621 tcggcatgga cgagctgtac aagtccggac tcagatctat ggtgatggag aagcctagtc
     1681 ccctgctggt cgggcgggaa tttgtgagac agtattacac actgctgaac caggccccag
     1741 acatgctgca tagattttat ggaaagaact cttcttatgt ccatggggga ttggattcaa
     1801 atggaaagcc agcagatgca gtctacggac agaaagaaat ccacaggaaa gtgatgtcac
     1861 aaaacttcac caactgccac accaagattc gccatgttga tgctcatgcc acgctaaatg
     1921 atggtgtggt agtccaggtg atggggcttc tctctaacaa caaccaggct ttgaggagat
     1981 tcatgcaaac gtttgtcctt gctcctgagg ggtctgttgc aaataaattc tatgttcaca
     2041 atgatatctt cagataccaa gatgaggtct ttggtgggtt tgtcactgag cctcaggagg
     2101 agtctgaaga agaagtagag gaacctgaag aaagacagca aacacctgag gtggtacctg
     2161 atgattctgg aactttctat gatcaggcag ttgtcagtaa tgacatggaa gaacatttag
     2221 aggagcctgt tgctgaacca gagcctgatc ctgaaccaga accagaacaa gaacctgtat
     2281 ctgaaatcca agaggaaaag cctgagccag tattagaaga aactgcccct gaggatgctc
     2341 agaagagttc ttctccagca cctgcagaca tagctcagac agtacaggaa gacttgagga
     2401 cattttcttg ggcatctgtg accagtaaga atcttccacc cagtggagct gttccagtta
     2461 ctgggatacc acctcatgtt gttaaagtac cagcttcaca gccccgtcca gagtctaagc
     2521 ctgaatctca gattccacca caaagacctc agcgggatca aagagtgcga gaacaacgaa
     2581 taaatattcc tccccaaagg ggacccagac caatccgtga ggctggtgag caaggtgaca
     2641 ttgaaccctg agaattctgc agtcgacggt accgcgggcc cgggatccac tagtccagtg
     2701 tggtggaatt ctgcagatat ccagcacagt ggcggccgct cgagtctaga gggcccgttt
     2761 aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     2821 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     2881 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     2941 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     3001 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     3061 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3121 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3181 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3241 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtacctag aagttcctat
     3301 tccgaagttc ctattctcta gaaagtatag gaacttcctt ggccaaaaag cctgaactca
     3361 ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc
     3421 agctctcgga gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg
     3481 tcctgcgggt aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact
     3541 ttgcatcggc cgcgctcccg attccggaag tgcttgacat tggggaattc agcgagagcc
     3601 tgacctattg catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg
     3661 aactgcccgc tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc
     3721 ttagccagac gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat
     3781 ggcgtgattt catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg
     3841 acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg
     3901 actgccccga agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg
     3961 acaatggccg cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat
     4021 acgaggtcgc caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc
     4081 gctacttcga gcggaggcat ccggagcttg caggatcgcc gcggctccgg gcgtatatgc
     4141 tccgcattgg tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag
     4201 cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta
     4261 cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa gtactcgccg
     4321 atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa ggaatagcac gtactacgag
     4381 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     4441 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     4501 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4561 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4621 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     4681 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     4741 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     4801 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     4861 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4921 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     4981 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5041 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     5101 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5161 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5221 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5281 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5341 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5401 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5461 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5521 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5581 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5641 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5701 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5761 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     5821 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     5881 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     5941 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     6001 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6061 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     6121 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6181 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6241 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6301 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6361 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6421 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6481 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6541 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6601 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6661 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6721 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     6781 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     6841 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     6901 gtatctgctc cctgcttgtg
//
