LOCUS       sequence-251612-        8447 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 8447)
  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 8447)
  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_RPS2_K275R
FEATURES             Location/Qualifiers
     source          1..8447
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    283..400
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        483..862
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        863..1066
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1092..1111
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1092..1110
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2073..2102
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2121..2144
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    2178..2751
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2201..2753
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2345..2362)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2572..2591
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2753..3352
                     /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(2753..2772)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3249..3269
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(3382..3415)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3471..4059
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3524..3544)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3942..3953)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4062..4078)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4063..4079)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             4588..4768
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(4830..5418)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4910..4929)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5495..6154)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     6069..6088
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(6155..6259)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6334..6353)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6524..6903
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6905..7103
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             7121..7301
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7348..7473
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7970..8203
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8388..8432
                     /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 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt
       61 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca
      121 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa
      181 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg
      241 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
      301 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
      361 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
      421 cagagatcca gtttggttag taccgggccc gctctagaca tgtccaatat gaccgccatg
      481 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
      541 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
      601 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
      661 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
      721 tcaagtgtat catatgccaa gtccgccccc tattgacgtc aatgacggta aatggcccgc
      781 ctggcattat gcccagtaca tgaccttacg ggactttcct acttggcagt acatctacgt
      841 attagtcatc gctattacca tggtgatgcg gttttggcag tacaccaatg ggcgtggata
      901 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
      961 ttggcaccaa aatcaacggg actttccaaa atgtcgtaat aaccccgccc cgttgacgca
     1021 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt tagtgaaccg
     1081 tcagaatttt gtaatacgac tcactatagg gcggccggga attcgtcgac tggatccggt
     1141 accgaggaga tctgccgccg cgatcgccgg cgcgtatggc ggatgacgcc ggtgcagcgg
     1201 gggggcccgg gggccctggt ggccctggga tggggaaccg cggtggcttc cgcggaggtt
     1261 tcggcagtgg catccggggc cggggtcgcg gccgtggacg gggccggggc cgaggccgcg
     1321 gagctcgcgg aggcaaggcc gaggataagg agtggatgcc cgtcaccaag ttgggccgct
     1381 tggtcaagga catgaagatc aagtccctgg aggagatcta tctcttctcc ctgcccatta
     1441 aggaatcaga gatcattgat ttcttcctgg gggcctctct caaggatgag gttttgaaga
     1501 ttatgccagt gcagaagcag acccgtgccg gccagcgcac caggttcaag gcatttgttg
     1561 ctatcgggga ctacaatggc cacgtcggtc tgggtgttaa gtgctccaag gaggtggcca
     1621 ccgccatccg tggggccatc atcctggcca agctctccat cgtccccgtg cgcagaggct
     1681 actgggggaa caagatcggc aagccccaca ctgtcccttg caaggtgaca ggccgctgcg
     1741 gctctgtgct ggtacgcctc atccctgcac ccaggggcac tggcatcgtc tccgcacctg
     1801 tgcctaagaa gctgctcatg atggctggta tcgatgactg ctacacctca gcccggggct
     1861 gcactgccac cctgggcaac ttcgccaagg ccacctttga tgccatttct aagacctaca
     1921 gctacctgac ccccgacctc tggaaggaga ctgtattcac caagtctccc tatcaggagt
     1981 tcactgacca cctcgtcagg acccacacca gagtctccgt gcagcggact caggctccag
     2041 ctgtggctac aacaacgcgt acgcggccgc tcgagcagaa actcatctca gaagaggatc
     2101 tggcagcaaa tgatatcctg gattacaagg atgacgacga taaggtttaa acctaggcgt
     2161 agcggccgca aattccgccc ctctccctcc ccccccccta acgttactgg ccgaagccgc
     2221 ttggaataag gccggtgtgc gtttgtctat atgttatttt ccaccatatt gccgtctttt
     2281 ggcaatgtga gggcccggaa acctggccct gtcttcttga cgagcattcc taggggtctt
     2341 tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg tgaaggaagc agttcctctg
     2401 gaagcttctt gaagacaaac aacgtctgta gcgacccttt gcaggcagcg gaacccccca
     2461 cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg
     2521 gcacaacccc agtgccacgt tgtgagttgg atagttgtgg aaagagtcaa atggctctcc
     2581 tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg taccccattg tatgggatct
     2641 gatctggggc ctcggtgcac atgctttaca tgtgtttagt cgaggttaaa aaaacgtcta
     2701 ggccccccga accacgggga cgtggttttc ctttgaaaaa cacgatgata atatgaccga
     2761 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc ccccgggcag tacgcaccct
     2821 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccag accgccacat
     2881 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     2941 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     3001 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     3061 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     3121 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     3181 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     3241 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     3301 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gatgtacaag
     3361 taggattcgt cgagggacct aataacttcg tatagcatac attatacgaa gttatacatg
     3421 tttaagggtt ccggttccac taggtacaat tcgatatcaa gcttatcgat aatcaacctc
     3481 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     3541 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     3601 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     3661 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     3721 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     3781 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     3841 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
     3901 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     3961 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     4021 ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt cgacctcgat
     4081 cgagacctag aaaaacatgg agcaatcaca agtagcaata cagcagctac caatgctgat
     4141 tgtgcctggc tagaagcaca agaggaggag gaggtgggtt ttccagtcac acctcaggta
     4201 cctttaagac caatgactta caaggcagct gtagatctta gccacttttt aaaagaaaag
     4261 gggggactgg aagggctaat tcactcccaa cgaagacaag atatccttga tctgtggatc
     4321 taccacacac aaggctactt ccctgattgg cagaactaca caccagggcc agggatcaga
     4381 tatccactga cctttggatg gtgctacaag ctagtaccag ttgagcaaga gaaggtagaa
     4441 gaagccaatg aaggagagaa cacccgcttg ttacaccctg tgagcctgca tgggatggat
     4501 gacccggaga gagaagtatt agagtggagg tttgacagcc gcctagcatt tcatcacatg
     4561 gcccgagagc tgcatccgga ctgtactggg tctctctggt tagaccagat ctgagcctgg
     4621 gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg
     4681 cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc
     4741 ttttagtcag tgtggaaaat ctctagcagc atgtgagcaa aaggccagca aaaggccagg
     4801 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4861 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4921 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4981 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5041 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5101 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5161 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5221 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     5281 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5341 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5401 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5461 aacgaaaact cacgttaagg gattttggtc atgattacgc cccgccctgc cactcatcgc
     5521 agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa acggcatgat
     5581 gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat ttgcccatgg
     5641 tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac
     5701 tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta gggaaatagg
     5761 ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac tgccggaaat
     5821 cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt
     5881 aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc atacggaact
     5941 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
     6001 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
     6061 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
     6121 caacggtggt atatccagtg atttttttct ccatactctt cctttttcaa tattattgaa
     6181 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6241 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg
     6301 gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     6361 aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat
     6421 ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag
     6481 gcgttttgcg ctgcttcgcg atgtacgggc cagatatcgc gttgacattg attattgact
     6541 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
     6601 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     6661 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     6721 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     6781 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     6841 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     6901 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
     6961 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
     7021 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
     7081 cggtgggagg tctatataag cagcgcgttt tgcctgtact gggtctctct ggttagacca
     7141 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     7201 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     7261 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
     7321 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
     7381 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
     7441 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
     7501 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
     7561 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca
     7621 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
     7681 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
     7741 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
     7801 gcacagcaag cggccggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     7861 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     7921 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     7981 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
     8041 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     8101 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     8161 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     8221 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     8281 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
     8341 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
     8401 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaac
//
