LOCUS       sequence-251590-        8298 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 8298)
  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 8298)
  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: pLENTI_RPS3_K227R
FEATURES             Location/Qualifiers
     source          1..8298
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    133..250
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        333..712
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        713..916
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     942..961
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        942..960
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1774..1803
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1822..1845
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    1879..2452
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    1902..2454
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2046..2063)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2273..2292
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2454..3053
                     /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(2454..2473)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2950..2970
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(3083..3116)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3172..3760
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3225..3245)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3643..3654)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3763..3779)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3764..3780)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             4289..4469
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(4531..5119)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4611..4630)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5196..5855)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5770..5789
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(5856..5960)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6035..6054)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6225..6604
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6606..6804
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             6822..7002
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7049..7174
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7671..7904
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8089..8133
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
ORIGIN
        1 aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga
       61 tccattcgat tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt
      121 attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat
      181 agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat
      241 tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggttag taccgggccc
      301 gctctagaca tgtccaatat gaccgccatg ttgacattga ttattgacta gttattaata
      361 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
      421 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      481 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      541 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc
      601 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg
      661 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
      721 gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
      781 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      841 atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt
      901 ctatataagc agagctcgtt tagtgaaccg tcagaatttt gtaatacgac tcactatagg
      961 gcggccggga attcgtcgac tggatccggt accgaggaga tctgccgccg cgatcgccgg
     1021 cgcgccatgg cagtgcaaat atccaagaag aggaagtttg tcgctgatgg catcttcaaa
     1081 gctgaactga atgagtttct tactcgggag ctggctgaag atggctactc tggagttgag
     1141 gtgcgagtta caccaaccag gacagaaatc attatcttag ccaccagaac acagaatgtt
     1201 cttggtgaga agggccggcg gattcgggaa ctgactgctg tagttcagaa gaggtttggc
     1261 tttccagagg gcagtgtaga gctttatgct gaaaaggtgg ccactagagg tctgtgtgcc
     1321 attgcccagg cagagtctct gcgttacaaa ctcctaggag ggcttgctgt gcggagggcc
     1381 tgctatggtg tgctgcggtt catcatggag agtggggcca aaggctgcga ggttgtggtg
     1441 tctgggaaac tccgaggaca gagggctaaa tccatgaagt ttgtggatgg cctgatgatc
     1501 cacagcggag accctgttaa ctactacgtt gacactgctg tgcgccacgt gttgctcaga
     1561 cagggtgtgc tgggcatcaa ggtgaagatc atgctgccct gggacccaac tggtaagatt
     1621 ggccctaaga agcccctgcc tgaccacgtg agcattgtgg aacccaaaga tgagatactg
     1681 cccaccaccc ccatctcaga acagaggggt gggaagccag agccgcctgc catgccccag
     1741 ccagtcccca cagcaacgcg tacgcggccg ctcgagcaga aactcatctc agaagaggat
     1801 ctggcagcaa atgatatcct ggattacaag gatgacgacg ataaggttta aacctaggcg
     1861 tagcggccgc aaattccgcc cctctccctc ccccccccct aacgttactg gccgaagccg
     1921 cttggaataa ggccggtgtg cgtttgtcta tatgttattt tccaccatat tgccgtcttt
     1981 tggcaatgtg agggcccgga aacctggccc tgtcttcttg acgagcattc ctaggggtct
     2041 ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct
     2101 ggaagcttct tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc ggaacccccc
     2161 acctggcgac aggtgcctct gcggccaaaa gccacgtgta taagatacac ctgcaaaggc
     2221 ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg gaaagagtca aatggctctc
     2281 ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag gtaccccatt gtatgggatc
     2341 tgatctgggg cctcggtgca catgctttac atgtgtttag tcgaggttaa aaaaacgtct
     2401 aggccccccg aaccacgggg acgtggtttt cctttgaaaa acacgatgat aatatgaccg
     2461 agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt cccccgggca gtacgcaccc
     2521 tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatcca gaccgccaca
     2581 tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca
     2641 aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg
     2701 aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc
     2761 tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt
     2821 ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg
     2881 tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct
     2941 ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg
     3001 aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgatgtacaa
     3061 gtaggattcg tcgagggacc taataacttc gtatagcata cattatacga agttatacat
     3121 gtttaagggt tccggttcca ctaggtacaa ttcgatatca agcttatcga taatcaacct
     3181 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     3241 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     3301 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     3361 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     3421 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     3481 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     3541 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt
     3601 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     3661 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     3721 cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg tcgacctcga
     3781 tcgagaccta gaaaaacatg gagcaatcac aagtagcaat acagcagcta ccaatgctga
     3841 ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt tttccagtca cacctcaggt
     3901 acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa
     3961 ggggggactg gaagggctaa ttcactccca acgaagacaa gatatccttg atctgtggat
     4021 ctaccacaca caaggctact tccctgattg gcagaactac acaccagggc cagggatcag
     4081 atatccactg acctttggat ggtgctacaa gctagtacca gttgagcaag agaaggtaga
     4141 agaagccaat gaaggagaga acacccgctt gttacaccct gtgagcctgc atgggatgga
     4201 tgacccggag agagaagtat tagagtggag gtttgacagc cgcctagcat ttcatcacat
     4261 ggcccgagag ctgcatccgg actgtactgg gtctctctgg ttagaccaga tctgagcctg
     4321 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     4381 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     4441 cttttagtca gtgtggaaaa tctctagcag catgtgagca aaaggccagc aaaaggccag
     4501 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     4561 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     4621 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     4681 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     4741 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     4801 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     4861 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     4921 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     4981 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     5041 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     5101 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     5161 gaacgaaaac tcacgttaag ggattttggt catgattacg ccccgccctg ccactcatcg
     5221 cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca aacggcatga
     5281 tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata tttgcccatg
     5341 gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa actggtgaaa
     5401 ctcacccagg gattggctga gacgaaaaac atattctcaa taaacccttt agggaaatag
     5461 gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa ctgccggaaa
     5521 tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg gaaaacggtg
     5581 taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc catacggaac
     5641 tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata aaacttgtgc
     5701 ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt ctggttatag
     5761 gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca ttgggatata
     5821 tcaacggtgg tatatccagt gatttttttc tccatactct tcctttttca atattattga
     5881 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     5941 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt cgacggatcg
     6001 ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     6061 taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa
     6121 tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg cttagggtta
     6181 ggcgttttgc gctgcttcgc gatgtacggg ccagatatcg cgttgacatt gattattgac
     6241 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     6301 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     6361 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     6421 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     6481 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     6541 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     6601 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
     6661 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     6721 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
     6781 acggtgggag gtctatataa gcagcgcgtt ttgcctgtac tgggtctctc tggttagacc
     6841 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
     6901 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
     6961 gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc cgaacaggga
     7021 cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc ttgctgaagc
     7081 gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt tgactagcgg
     7141 aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga gaattagatc
     7201 gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat taaaacatat
     7261 agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt tagaaacatc
     7321 agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag gatcagaaga
     7381 acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa ggatagagat
     7441 aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa gtaagaccac
     7501 cgcacagcaa gcggccggcc gctgatcttc agacctggag gaggagatat gagggacaat
     7561 tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc
     7621 accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg
     7681 ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg
     7741 gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct
     7801 attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca
     7861 agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc
     7921 tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct
     7981 ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac
     8041 acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa
     8101 gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg
     8161 ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt
     8221 tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag
     8281 acccacctcc caaccccg
//
