LOCUS       sequence-448479-        8051 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mEGFP-fusion protein of TOP3B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8051)
  AUTHORS   Chen X, Fansler MM, Janjos U, Ule J, Mayr C
  TITLE     The FXR1 network acts as signaling scaffold for actomyosin 
            remodeling.
  JOURNAL   Cell. 5 August 2024. doi: 10.1016/j.cell.2024.07.015.
  PUBMED    37961296
REFERENCE   2  (bases 1 to 8051)
  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: "Cell. 5 
            August 2024. doi: 10.1016/j.cell.2024.07.015."
COMMENT     Sequence Label: pcDNA3puro-mGFP-TOP3B
FEATURES             Location/Qualifiers
     source          1..8051
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     170..190
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    447..568
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(484..503)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     538..557
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(617..633)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(617..633)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(630..652)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    641..657
                     /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(665..671,672..689,690..695))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    710..731
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(848..865)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1019..1607)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1099..1118)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1778..2569,2570..2638))
                     /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     2401..2420
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2639..2743)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2818..2837)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3009..3388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3389..3592
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3543..3563
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3637..3656
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3637..3655
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(3679..3681,3682..3684,3685..4395)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3724..3745)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3985..4004)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4332..4353
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(6665..6682)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6671..6895
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6941..7369
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7028..7047)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7238..7259
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7378..7398)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7383..7712
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7563..7698
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7625..7644
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             join(7725..8051,1..273)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7725..7744)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
ORIGIN
        1 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
       61 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
      121 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
      181 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
      241 acctggtgca tgacccgcaa gcccggtgcc tgagcgggac tctggggttc gaaatgaccg
      301 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
      361 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
      421 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
      481 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
      541 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
      601 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
      661 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
      721 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
      781 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
      841 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
      901 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
      961 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     1021 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     1081 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     1141 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     1201 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     1261 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     1321 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     1381 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     1441 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     1501 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     1561 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     1621 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     1681 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     1741 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     1801 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     1861 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     1921 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     1981 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     2041 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     2101 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     2161 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     2221 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     2281 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     2341 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     2401 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     2461 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     2521 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     2581 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     2641 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     2701 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     2761 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
     2821 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
     2881 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
     2941 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
     3001 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
     3061 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     3121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     3181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     3241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     3301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     3361 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     3421 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     3481 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     3541 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct
     3601 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
     3661 ccaagctggc tagcaaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
     3721 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
     3781 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
     3841 cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc
     3901 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
     3961 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
     4021 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
     4081 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg
     4141 gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac
     4201 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     4261 ctgctgcccg acaaccacta cctgagcacc cagtccaagc tgagcaaaga ccccaacgag
     4321 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
     4381 gacgagctgt acaagaagac tgtgctcatg gttgctgaaa agccgtcctt ggcacagtca
     4441 attgccaaaa tcctctctag agggagcctg tcctcacaca aagggctgaa cggggcctgc
     4501 tcagtccacg agtacactgg gacctttgct ggccagccag tgcgcttcaa gatgacgtct
     4561 gtctgtggtc acgtgatgac cctggatttc ctgggaaaat acaacaaatg ggacaaagtg
     4621 gaccccgcag aactgttcag ccaagctccc acggagaaga aagaagctaa ccccaagctg
     4681 aacatggtga agttcctgca ggtggagggc agaggctgcg actacatcgt gctgtggctg
     4741 gactgcgaca aggaggggga gaacatctgc tttgaggtta acactgacaa agacagatct
     4801 ctccttttgg actgggaccg agtaagagtg tttgaccggg agatcgcaca gatgttttta
     4861 aacatgacaa agctggagaa ggaagcccag gtggaggcca caagcaggaa agaaaaggcc
     4921 aagcagaggc ccctggccct gaacactgtg gagatgctgc gtgtggccag ctcttctctg
     4981 ggcatggggc cgcagcacgc catgcagacg gctgagcggc tctacacgca aggctacatc
     5041 agctacccac ggacagagac cacccactac cctgagaact ttgacctgaa gggctctctg
     5101 cggcagcagg ccaaccaccc ctactgggcc gacacggtga agcggttgtt agcagaaggt
     5161 atcaaccgcc cgcggaaagg ccatgacgcc ggcgaccatc cccccatcac ccccatgaag
     5221 tctgccacag aggccgaatt agggggtgac gcgtggcggc tctatgagta catcaccaga
     5281 cacttcatcg ccacggtcag ccatgactgc aagtacctgc agagcaccat ctccttcaga
     5341 attgggcccg agctcttcac ctgctccggg aagaccgtcc tctcaccagg cttcacggag
     5401 gtcatgccct ggcagagcgt gcccctggag gagagcctgc ccacttgcca gcggggtgat
     5461 gccttccctg tgggcgaggt gaagatgctg gagaagcaga cgaacccacc cgactacctg
     5521 acggaggccg agctcatcac gctcatggag aagcatggca tcggcacgga tgccagcatc
     5581 cctgtgcata tcaacaacat ctgccagcgc aactatgtca cggtggagag cgggcgccgg
     5641 ctcaagccca ccaacctcgg catcgtcctg gtgcacggct actataagat tgatgcagag
     5701 ctggtgctcc ccaccatccg cagtgcagtg gagaagcagc tgaacctgat cgcccagggc
     5761 aaggccgact accgccaggt cctgggccac accctggacg tgttcaagag gaagttccac
     5821 tactttgtcg actccattgc tggcatggat gagttgatgg aggtgtcttt ctcgcccctg
     5881 gcggccacag gcaagcccct ctcacgctgt gggaagtgcc accgcttcat gaagtacatc
     5941 caggccaagc caagccgcct gcactgctcc cactgcgatg agacctacac gctcccccag
     6001 aacggcacca tcaagctcta caaggagctc cgctgccctc tggatgactt cgagctggtc
     6061 ctgtggtcat caggctctcg gggcaagagc tacccgctgt gcccctactg ctacaaccac
     6121 ccacccttcc gagacatgaa gaaaggcatg ggctgcaacg agtgtacgca cccctcctgc
     6181 cagcactcgc tgagcatgct gggcatcggc cagtgcgtgg aatgtgagag cggggtgctg
     6241 gtgctggacc ccacctcggg ccccaagtgg aaggtggcct gcaacaagtg caacgtggta
     6301 gcgcactgct tcgagaacgc ccaccgcgtg cgggtgtccg ccgacacctg cagtgtctgt
     6361 gaggccgcct tgcttgatgt ggacttcaac aaggccaagt ccccactccc gggcgatgag
     6421 acgcagcaca tgggctgcgt cttttgtgac cccgtcttcc aggagctggt ggagctgaag
     6481 catgcggcct cctgccaccc catgcaccgc ggtggaccag ggagaaggca gggtcgaggg
     6541 cggggccggg ccaggaggcc ccctgggaag cccaacccca gacggcccaa ggacaagatg
     6601 tcagccctgg ccgcctactt tgtatgactc gagtctagag ggcccgttta aacccgctga
     6661 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     6721 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     6781 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     6841 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     6901 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     6961 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     7021 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     7081 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     7141 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     7201 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     7261 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     7321 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     7381 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     7441 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     7501 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     7561 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     7621 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     7681 gaggcttttt tggaggccta ggcttttgca aaaagctccc aaccatgacc gagtacaagc
     7741 ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg
     7801 cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg
     7861 tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg
     7921 tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg
     7981 cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc
     8041 agcaacagat g
//
