LOCUS       sequence-251368-        5763 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5763)
  AUTHORS   Meyer C, Garzia A, Morozov P, Molina H, Tuschl T
  TITLE     The G3BP1-Family-USP10 Deubiquitinase Complex Rescues Ubiquitinated 
            40S Subunits of Ribosomes Stalled in Translation from Lysosomal 
            Degradation.
  JOURNAL   Mol Cell. 2020 Mar 19;77(6):1193-1205.e5. doi: 
            10.1016/j.molcel.2019.12.024. Epub 2020 Jan 24.
  PUBMED    31981475
REFERENCE   2  (bases 1 to 5763)
  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.2019.12.024"; journalName: "Mol Cell"; date: 
            "2020-03-19- 19"; volume: "77"; issue: "6"; pages: "1193-1205.e5"
COMMENT     Sequence Label: pcDNA5_FRT_GFP
FEATURES             Location/Qualifiers
     source          1..5763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        106..485
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        486..689
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     640..660
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     734..753
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        734..752
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(811..813,814..816,817..1527)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(811..813,814..816,817..1527)
                     /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(856..877)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1117..1136)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1464..1485
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1586..1603)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1592..1816
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1949..1968)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2100..2147
                     /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             2155..3177
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2155..3177
                     /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    3307..3428
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3344..3363)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3398..3417
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3477..3493)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3477..3493)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3490..3512)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3501..3517
                     /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(3525..3531,3532..3549,3550..3555))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3570..3591
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3708..3725)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3879..4467)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3959..3978)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4638..5429,5430..5498))
                     /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     5261..5280
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5499..5603)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5678..5697)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg cttagggtta
       61 ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat tgattattga
      121 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
      181 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      241 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      301 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      361 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      421 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      481 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
      541 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      601 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
      661 tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc actgcttact
      721 ggcttatcga aattaatacg actcactata gggagaccca agctggctag cgtttaaact
      781 taagcttggt accgagctcg gatcccgagt atggtgagca agggcgagga gctgttcacc
      841 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
      901 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
      961 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     1021 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     1081 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     1141 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     1201 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     1261 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     1321 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     1381 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa
     1441 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     1501 actctcggca tggacgagct gtacaagaag cttactagct aacggccgct cgagtctaga
     1561 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     1621 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     1681 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     1741 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     1801 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     1861 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     1921 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     1981 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     2041 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtacctag
     2101 aagttcctat tccgaagttc ctattctcta gaaagtatag gaacttcctt ggccaaaaag
     2161 cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc
     2221 gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg
     2281 cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga tcgttatgtt
     2341 tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat tggggaattc
     2401 agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg
     2461 cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct
     2521 gcggccgatc ttagccagac gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa
     2581 tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta tcactggcaa
     2641 actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt
     2701 tgggccgagg actgccccga agtccggcac ctcgtgcacg cggatttcgg ctccaacaat
     2761 gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc gatgttcggg
     2821 gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc ttgtatggag
     2881 cagcagacgc gctacttcga gcggaggcat ccggagcttg caggatcgcc gcggctccgg
     2941 gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga cggcaatttc
     3001 gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg agccgggact
     3061 gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa
     3121 gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa ggaatagcac
     3181 gtactacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     3241 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     3301 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     3361 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     3421 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     3481 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     3541 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     3601 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     3661 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     3721 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     3781 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     3841 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     3901 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     3961 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4021 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4081 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4141 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4201 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4261 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     4321 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4381 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4441 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4501 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     4561 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     4621 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     4681 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     4741 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     4801 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     4861 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     4921 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     4981 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5041 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5101 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5161 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5221 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     5281 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5341 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5401 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     5461 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     5521 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     5581 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
     5641 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
     5701 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
     5761 aaa
//
