LOCUS       sequence-430573-        5584 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a circular RNA containing a Squash aptamer and an IRE (TfR
            derived) in mammalian cells, using the Tornado expression system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5584)
  AUTHORS   Hagen T, Litke JL, Nasir N, Hou Q, Jaffrey SR
  TITLE     Engineering acyclovir-induced RNA nanodevices for reversible and 
            tunable control of aptamer function.
  JOURNAL   Cell Chem Biol. 2024 Aug 23:S2451-9456(24)00319-2. doi: 
            10.1016/j.chembiol.2024.07.017.
  PUBMED    39191249
REFERENCE   2  (bases 1 to 5584)
  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"; journalName: "Cell Chem 
            Biol. 2024 Aug 23:S2451-9456(24)00319-2. doi: 
            10.1016/j.chembiol.2024.07.017."
COMMENT     Sequence Label: pAV-puro-U6+27-F30-Tornado-Squash-TfR-IRE
FEATURES             Location/Qualifiers
     source          1..5584
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     242..262
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        290..538
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     290..310
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     461..480
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     polyA_signal    1009..1130
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1227..1682)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1364..1385)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1576..1595
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1709..1813
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1814..1882,1883..2674)
                     /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     complement(2032..2051)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2845..3433
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3334..3353
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3587..3604
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    3702..3836
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(4261..4281)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          4411..4476
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             complement(4903..5502)
                     /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(4986..5006)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     5483..5502
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(join(5512..5584,1..257))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(5526..5584,1..77)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcctctgcat aaataaaaaa aattagtcag ccatggggcg gagaatgggc ggaactgggc
       61 ggagttaggg gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga
      121 tgcatgcttt gcatacttct gcctgctggg gagcctgggg actttccaca cctggttgct
      181 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
      241 caccctaact gacacacatt ccacagccgg atccaaggtc gggcaggaag agggcctatt
      301 tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga taattagaat
      361 taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt
      421 tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt
      481 aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccgt
      541 gctcgcttcg gcagcacata tactagtcga cgggccgcac tcgccggtcc caagcccgga
      601 taaaatggga gggggcggga aaccgcctaa ccatgccgag tgcggccgct tgccatgtgt
      661 atcggtccgc tacaaggtga gcccaataat acggtttggg ttaggatagg aagtagagcc
      721 gtaaactctc taagcggtag cggtccgata ctctgatgat gggtcccgat tatcgggagc
      781 agtgtcttcc ataatcgggt cccatcattc atggcaagtg gccgcggtcg gcgtggactg
      841 tagaacactg ccaatgccgg tcccaagccc ggataaaagt ggagggtaca gtccacgctc
      901 tagagcgcac ttcggtgcgc tttttactag gacctgcagg catgcaagct tgacgtcggt
      961 taccgatatc catatggcga ccgcatcgat ctcgagccga ggactagtaa cttgtttatt
     1021 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     1081 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctta
     1141 cgtagataag tagcatggcg ggttatagcg aagaggcccg caccgatcgc ccttcccaac
     1201 agttgcgcag cctgaatggc gaatggaaat tgtaagcgtt aatattttgt taaaattcgc
     1261 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     1321 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     1381 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     1441 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     1501 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     1561 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     1621 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     1681 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     1741 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     1801 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     1861 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     1921 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     1981 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     2041 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     2101 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     2161 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     2221 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     2281 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     2341 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     2401 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     2461 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     2521 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     2581 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     2641 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     2701 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     2761 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     2821 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     2881 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     2941 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     3001 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     3061 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     3121 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     3181 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     3241 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3301 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     3361 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     3421 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3481 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3541 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     3601 cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctggag gggtggagtc
     3661 gtgacgtgaa ttacgtcata gggttaggga ggtcctggat cgatccagac atgataagat
     3721 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     3781 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     3841 acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa
     3901 gcaagtaaaa cctctacaaa tgtggtatgg ctgattatga tctctagtca aggcactata
     3961 catcaaatat tccttattaa cccctttaca aattaaaaag ctaaaggtac acaatttttg
     4021 agcatagtta ttaatagcag acactctatg cctgtgtgga gtaagaaaaa acagtatgtt
     4081 atgattataa ctgttatgcc tacttataaa ggttacagaa tatttttcca taattttctt
     4141 gtatagcagt gcagcttttt cctttgtggt gtaaatagca aagcaagcaa gagttctatt
     4201 actaaacaca gcatgactca aaaaacttag caattctgaa ggaaagtcct tggggtcttc
     4261 tacctttctc ttcttttttg gaggagtaga atgttgagag tcagcagtag cctcatcatc
     4321 actagatggc atttcttctg agcaaaacag gttttcctca ttaaaggcat tccaccactg
     4381 ctcccattca tcagttccat aggttggaat ctaaaataca caaacaatta gaatcagtag
     4441 tttaacacat tatacactta aaaattttat atttacctta gagctttaaa tctctgtagg
     4501 tagtttgtcc aattatgtca caccacagaa gtaaggttcc ttcacaaaga tccggaccaa
     4561 agcggccatc gtgcctcccc actcctgcag ttcgggggca tggatgcgcg gatagccgct
     4621 gctggtttcc tggatgccga cggatttgca ctgccggtag aactccgcga ggtcgtccag
     4681 cctcaggcag cagctgaacc aactcgcgag gggatcgagc ccggggtggg cgaagaactc
     4741 cagcatgaga tccccgcgct ggaggatcat ccagccggcg tcccggaaaa cgattccgaa
     4801 gcccaacctt tcatagaagg cggcggtgga atcgaaatct cgtgatggca ggttgggcgt
     4861 cgcttggtcg gtcatttcga actgcgggtc gtggggcggg cgtcaggcac cgggcttgcg
     4921 ggtcatgcac caggtgcgcg gtccttcggg cacctcgacg tcggcggtga cggtgaagcc
     4981 gagccgctcg tagaagggga ggttgcgggg cgcggaggtc tccaggaagg cgggcacccc
     5041 ggcgcgctcg gccgcctcca ctccggggag cacgacggcg ctgcccagac ccttgccctg
     5101 gtggtcgggc gagacgccga cggtggccag gaaccacgcg ggctccttgg gccggtgcgg
     5161 cgccaggagg ccttccatct gttgctgcgc ggccagccgg gaaccgctca actcggccat
     5221 gcgcgggccg atctcggcga acaccgcccc cgcttcgacg ctctccggcg tggtccagac
     5281 cgccacggca gcaccgtcgt ccgcgaccca caccttgccg atgtcgagcc cgacgcgcgt
     5341 gaggaagagt tcttgcagct cggtgacccg ctcgatgtgg cgatcaggat cgacggtgtg
     5401 gcgcgtggcg gggtagtcgg cgaacgcggc ggcgagggtg cgtacggccc tggggacgtc
     5461 gtcgcgggtg gcgaggcgca ccgtgggctt gtactcggtc atggtaagct ttttgcaaaa
     5521 gcctaggcct ccaaaaaagc ctcctcacta cttctggaat agctcagagg ccgaggcggc
     5581 ctcg
//
