LOCUS       sequence-251650-        7730 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 7730)
  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 7730)
  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: pFRT_TO_FlagHA_USP10_C424A
FEATURES             Location/Qualifiers
     source          1..7730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        103..482
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        483..686
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     637..657
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    688..706
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    709..727
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     737..761
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             861..884
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             885..911
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    927..951
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3352..3376)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3429..3470
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3550..3567)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3556..3780
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3913..3932)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4064..4111
                     /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             4119..5141
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4119..5141
                     /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    5271..5392
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5308..5327)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5362..5381
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5441..5457)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5441..5457)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5454..5476)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5465..5481
                     /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(5489..5495,5496..5513,5514..5519))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5534..5555
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5672..5689)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5843..6431)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5923..5942)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6602..7393,7394..7462))
                     /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     7225..7244
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7463..7567)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7642..7661)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc
       61 gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta
      121 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
      181 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      241 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      301 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      361 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
      421 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
      481 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
      541 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
      601 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac
      661 ggtgggaggt ctatataagc agagctctcc ctatcagtga tagagatctc cctatcagtg
      721 atagagatcg tcgacgagct cgtttagtga accgtcagat cgcctggaga cgccatccac
      781 gctgttttga cctccataga agacaccggg accgatccag cctccggact ctagcgttta
      841 aacttaagct tggtaccatg gactacaagg acgacgatga caagtaccct tatgacgtgc
      901 ccgattacgc tttgaaccag atatcaacaa gtttgtacaa aaaagcaggc ttcatggccc
      961 tccacagccc gcagtatatt tttggagatt ttagccctga tgaattcaat caattctttg
     1021 tgactcctcg atcttcagtt gagcttcctc catacagtgg aacagttctg tgtggcacac
     1081 aggctgtgga taaactacct gatggacaag aatatcagag aattgagttt ggtgtcgatg
     1141 aagtcattga acccagtgac actttgccga gaacccccag ctacagtatt tcaagcacac
     1201 tgaaccctca ggcccctgaa tttattctcg gttgtacagc ttccaaaata acccctgatg
     1261 gtatcactaa agaagcaagc tatggctcca tcgactgcca gtacccaggc tctgccctcg
     1321 ctttggatgg aagttctaat gtggaggcgg aagttttgga aaatgatggt gtctcaggtg
     1381 gtcttggaca aagggagcgt aaaaagaaga aaaagcggcc acctggatat tacagctatt
     1441 tgaaagatgg tggcgatgat agtatctcca cagaagccct ggtcaatggc catgccaatt
     1501 cagcagtccc gaacagtgtc agtgcagagg atgcagaatt tatgggtgac atgcccccgt
     1561 cagttacgcc caggacttgt aacagccccc agaactccac agactctgtc agtgacattg
     1621 tgcctgacag tcctttcccc ggagcactcg gcagtgacac caggactgca gggcagccag
     1681 aggggggccc cggggctgat tttggtcagt cctgcttccc tgcagaggct ggcagagaca
     1741 ccctgtcaag gacagctggg gctcagccct gcgttggtac cgatactact gaaaaccttg
     1801 gagttgctaa tggacaaata cttgaatcct cgggtgaggg cacagctacc aacggggtgg
     1861 agttgcacac cacggaaagc atagacttgg acccaaccaa acccgagagt gcatcacctc
     1921 ctgctgacgg cacgggctct gcatcaggca cccttcctgt cagccagccc aagtcctggg
     1981 ccagcctctt tcatgattct aagccctctt cctcctcgcc ggtggcctat gtggaaacta
     2041 agtattcccc tcccgccata tctcccctgg tttctgaaaa gcaggttgaa gtcaaagaag
     2101 ggcttgttcc ggtttcagag gatcctgtag ccataaagat tgcagagttg ctggagaatg
     2161 taaccctaat ccataaacca gtgtcgttgc aaccccgtgg gctgatcaat aaagggaact
     2221 gggcctacat taatgctaca ctgcaggcat tggttgcttg cccgccgatg taccacctga
     2281 tgaagttcat tcctctgtat tccaaagtgc aaaggccttg tacgtcaaca cccatgatag
     2341 acagctttgt tcggctaatg aatgagttca ctaatatgcc agtacctcca aaaccccgac
     2401 aagctcttgg agataaaatc gtgagggata ttcgccctgg agctgccttt gagcccacat
     2461 atatttacag actcctgaca gttaacaagt caagcctgtc tgaaaagggt cgacaagaag
     2521 atgctgagga atacttaggc ttcattctaa atggacttca tgaggaaatg ttgaacctaa
     2581 agaagcttct ctcaccaagt aatgaaaaac ttacgatttc caacggcccc aaaaaccact
     2641 cggtcaatga agaagagcag gaagaacaag gtgaaggaag cgaggatgaa tgggaacaag
     2701 tgggcccccg gaacaagact tccgtcaccc gccaggcgga ttttgttcag actccaatca
     2761 ccggcatttt tggtggacac atcaggtctg tggtttacca gcagagttca aaagaatctg
     2821 ccactttgca gccatttttc acgttgcagt tggatatcca gtcagacaag atacgcacag
     2881 tccaggatgc actggagagc ttggtggcaa gagaatctgt ccaaggttat accacaaaaa
     2941 ccaaacaaga ggttgagata agtcgaagag tgactctgga aaaactccct cctgtcctcg
     3001 tgctgcacct gaaacgattc gtttatgaga agactggtgg gtgccagaag cttatcaaaa
     3061 atattgaata tcctgtggac ttggaaatta gtaaagaact gctttctcca ggggttaaaa
     3121 ataagaattt taaatgccac cgaacctatc ggctgtttgc agtggtctac catcacggca
     3181 acagtgcgac gggcggccat tacactacag acgtcttcca gatcggtctg aatggctggc
     3241 tgcgcatcga tgaccagaca gtcaaggtga tcaaccagta ccaggtggtg aaaccaactg
     3301 ctgaacgcac agcctacctc ctgtattacc gccgagtgga cctgctgtaa tacccagctt
     3361 tcttgtacaa agtggttgat atccagcaca gtggcggccg ctcaagtcta gagggcccgc
     3421 ggttcgaagg taagcctatc cctaaccctc tcctcggtct cgattctacg cgtacctgat
     3481 ccgcggccgc atagataact gatccagtgt gcctcgagtc tagagggccc gtttaaaccc
     3541 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     3601 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     3661 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     3721 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     3781 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     3841 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     3901 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     3961 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     4021 tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa
     4081 gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga
     4141 cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct
     4201 cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc
     4261 gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat
     4321 cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct
     4381 attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc
     4441 ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc
     4501 agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg
     4561 atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca
     4621 ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc
     4681 ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg
     4741 gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg
     4801 tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact
     4861 tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca
     4921 ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg
     4981 cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa
     5041 tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg
     5101 gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg
     5161 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     5221 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     5281 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     5341 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     5401 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     5461 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     5521 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     5581 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5641 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5701 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     5761 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     5821 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     5881 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5941 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     6001 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6061 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6121 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6181 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     6241 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6301 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6361 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     6421 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     6481 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     6541 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6601 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6661 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6721 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6781 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6841 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6901 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6961 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     7021 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     7081 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     7141 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     7201 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     7261 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     7321 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7381 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7441 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     7501 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     7561 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
     7621 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     7681 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt
//
