LOCUS       sequence-251595-        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_K227/230R
FEATURES             Location/Qualifiers
     source          1..8298
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    175..292
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        375..754
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        755..958
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     984..1003
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        984..1002
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1816..1845
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1864..1887
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    1921..2494
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    1944..2496
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2088..2105)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2315..2334
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2496..3095
                     /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(2496..2515)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2992..3012
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(3125..3158)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3214..3802
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3267..3287)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3685..3696)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3805..3821)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3806..3822)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             4331..4511
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(4573..5161)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4653..4672)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5238..5897)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5812..5831
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(5898..6002)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6077..6096)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6267..6646
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6648..6846
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             6864..7044
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7091..7216
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7713..7946
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8131..8175
                     /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 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
       61 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
      121 ggatcggcac tgcgtgcgcc aattctgcag acaaatggca gtattcatcc acaattttaa
      181 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
      241 agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta
      301 ttacagggac agcagagatc cagtttggtt agtaccgggc ccgctctaga catgtccaat
      361 atgaccgcca tgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
      421 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      481 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      541 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
      601 cttggcagta catcaagtgt atcatatgcc aagtccgccc cctattgacg tcaatgacgg
      661 taaatggccc gcctggcatt atgcccagta catgacctta cgggactttc ctacttggca
      721 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacaccaa
      781 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
      841 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta ataaccccgc
      901 cccgttgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg
      961 tttagtgaac cgtcagaatt ttgtaatacg actcactata gggcggccgg gaattcgtcg
     1021 actggatccg gtaccgagga gatctgccgc cgcgatcgcc ggcgcgccat ggcagtgcaa
     1081 atatccaaga agaggaagtt tgtcgctgat ggcatcttca aagctgaact gaatgagttt
     1141 cttactcggg agctggctga agatggctac tctggagttg aggtgcgagt tacaccaacc
     1201 aggacagaaa tcattatctt agccaccaga acacagaatg ttcttggtga gaagggccgg
     1261 cggattcggg aactgactgc tgtagttcag aagaggtttg gctttccaga gggcagtgta
     1321 gagctttatg ctgaaaaggt ggccactaga ggtctgtgtg ccattgccca ggcagagtct
     1381 ctgcgttaca aactcctagg agggcttgct gtgcggaggg cctgctatgg tgtgctgcgg
     1441 ttcatcatgg agagtggggc caaaggctgc gaggttgtgg tgtctgggaa actccgagga
     1501 cagagggcta aatccatgaa gtttgtggat ggcctgatga tccacagcgg agaccctgtt
     1561 aactactacg ttgacactgc tgtgcgccac gtgttgctca gacagggtgt gctgggcatc
     1621 aaggtgaaga tcatgctgcc ctgggaccca actggtaaga ttggccctaa gaagcccctg
     1681 cctgaccacg tgagcattgt ggaacccaaa gatgagatac tgcccaccac ccccatctca
     1741 gaacagaggg gtgggaggcc agagccgcct gccatgcccc agccagtccc cacagcaacg
     1801 cgtacgcggc cgctcgagca gaaactcatc tcagaagagg atctggcagc aaatgatatc
     1861 ctggattaca aggatgacga cgataaggtt taaacctagg cgtagcggcc gcaaattccg
     1921 cccctctccc tccccccccc ctaacgttac tggccgaagc cgcttggaat aaggccggtg
     1981 tgcgtttgtc tatatgttat tttccaccat attgccgtct tttggcaatg tgagggcccg
     2041 gaaacctggc cctgtcttct tgacgagcat tcctaggggt ctttcccctc tcgccaaagg
     2101 aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct ctggaagctt cttgaagaca
     2161 aacaacgtct gtagcgaccc tttgcaggca gcggaacccc ccacctggcg acaggtgcct
     2221 ctgcggccaa aagccacgtg tataagatac acctgcaaag gcggcacaac cccagtgcca
     2281 cgttgtgagt tggatagttg tggaaagagt caaatggctc tcctcaagcg tattcaacaa
     2341 ggggctgaag gatgcccaga aggtacccca ttgtatggga tctgatctgg ggcctcggtg
     2401 cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc cgaaccacgg
     2461 ggacgtggtt ttcctttgaa aaacacgatg ataatatgac cgagtacaag cccacggtgc
     2521 gcctcgccac ccgcgacgac gtcccccggg cagtacgcac cctcgccgcc gcgttcgccg
     2581 actaccccgc cacgcgccac accgtcgatc cagaccgcca catcgagcgg gtcaccgagc
     2641 tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg
     2701 acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg
     2761 ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga
     2821 tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg
     2881 gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg
     2941 aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc
     3001 ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc
     3061 gcacctggtg catgacccgc aagcccggtg cctgatgtac aagtaggatt cgtcgaggga
     3121 cctaataact tcgtatagca tacattatac gaagttatac atgtttaagg gttccggttc
     3181 cactaggtac aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     3241 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     3301 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     3361 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     3421 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     3481 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     3541 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     3601 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     3661 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     3721 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     3781 tccctttggg ccgcctcccc gcatcgatac cgtcgacctc gatcgagacc tagaaaaaca
     3841 tggagcaatc acaagtagca atacagcagc taccaatgct gattgtgcct ggctagaagc
     3901 acaagaggag gaggaggtgg gttttccagt cacacctcag gtacctttaa gaccaatgac
     3961 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct
     4021 aattcactcc caacgaagac aagatatcct tgatctgtgg atctaccaca cacaaggcta
     4081 cttccctgat tggcagaact acacaccagg gccagggatc agatatccac tgacctttgg
     4141 atggtgctac aagctagtac cagttgagca agagaaggta gaagaagcca atgaaggaga
     4201 gaacacccgc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg agagagaagt
     4261 attagagtgg aggtttgaca gccgcctagc atttcatcac atggcccgag agctgcatcc
     4321 ggactgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     4381 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     4441 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     4501 aatctctagc agcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4561 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4621 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4681 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4741 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     4801 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     4861 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     4921 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     4981 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5041 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5101 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5161 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5221 agggattttg gtcatgatta cgccccgccc tgccactcat cgcagtactg ttgtaattca
     5281 ttaagcattc tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc
     5341 ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag
     5401 aagttgtcca tattggccac gtttaaatca aaactggtga aactcaccca gggattggct
     5461 gagacgaaaa acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa
     5521 cacgccacat cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc
     5581 cagagcgatg aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta
     5641 tcccatatca ccagctcacc gtctttcatt gccatacgga actccggatg agcattcatc
     5701 aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc
     5761 tttaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac
     5821 tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca
     5881 gtgatttttt tctccatact cttccttttt caatattatt gaagcattta tcagggttat
     5941 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     6001 cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct cccgatcccc
     6061 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc
     6121 ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct acaacaaggc
     6181 aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt gcgctgcttc
     6241 gcgatgtacg ggccagatat cgcgttgaca ttgattattg actagttatt aatagtaatc
     6301 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     6361 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     6421 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     6481 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     6541 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     6601 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     6661 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     6721 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     6781 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     6841 aagcagcgcg ttttgcctgt actgggtctc tctggttaga ccagatctga gcctgggagc
     6901 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc
     6961 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt
     7021 agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag cgaaagggaa
     7081 accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga
     7141 ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga aggagagaga
     7201 tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg
     7261 gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg caagcaggga
     7321 gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct gtagacaaat
     7381 actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat cattatataa
     7441 tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca ccaaggaagc
     7501 tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc aagcggccgg
     7561 ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat
     7621 aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga
     7681 gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga
     7741 gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta
     7801 ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat
     7861 ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa
     7921 agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc
     7981 accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat
     8041 cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc
     8101 ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat
     8161 aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta
     8221 ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata
     8281 gtgaatagag ttaggcag
//
