LOCUS       sequence-430552-        5585 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a circular RNA containing a Squash aptamer and an IRE (FTL
            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 5585)
  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 5585)
  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-FTL-IRE
FEATURES             Location/Qualifiers
     source          1..5585
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             161..181
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    606..740
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(838..855)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1009..1597)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1089..1108)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1768..2559,2560..2628))
                     /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     2391..2410
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2629..2733)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2760..3215
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2847..2866)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3057..3078
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    3312..3433
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        complement(3905..4153)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(3963..3982)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(4133..4153)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(4181..4201)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4186..4515
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4366..4501
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4525..5124
                     /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(4525..4544)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5021..5041
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     intron          join(5551..5585,1..31)
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
ORIGIN
        1 actactgatt ctaattgttt gtgtatttta gattccaacc tatggaactg atgaatggga
       61 gcagtggtgg aatgccttta atgaggaaaa cctgttttgc tcagaagaaa tgccatctag
      121 tgatgatgag gctactgctg actctcaaca ttctactcct ccaaaaaaga agagaaaggt
      181 agaagacccc aaggactttc cttcagaatt gctaagtttt ttgagtcatg ctgtgtttag
      241 taatagaact cttgcttgct ttgctattta caccacaaag gaaaaagctg cactgctata
      301 caagaaaatt atggaaaaat attctgtaac ctttataagt aggcataaca gttataatca
      361 taacatactg ttttttctta ctccacacag gcatagagtg tctgctatta ataactatgc
      421 tcaaaaattg tgtaccttta gctttttaat ttgtaaaggg gttaataagg aatatttgat
      481 gtatagtgcc ttgactagag atcataatca gccataccac atttgtagag gttttacttg
      541 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
      601 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
      661 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
      721 tatcttatca tgtctggatc gatccaggac ctccctaacc ctatgacgta attcacgtca
      781 cgactccacc cctccaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
      841 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
      901 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
      961 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     1021 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     1081 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     1141 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     1201 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     1261 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     1321 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     1381 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     1441 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     1501 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     1561 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     1621 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     1681 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     1741 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     1801 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     1861 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     1921 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     1981 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     2041 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     2101 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     2161 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     2221 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     2281 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     2341 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     2401 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     2461 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     2521 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     2581 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     2641 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     2701 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     2761 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2821 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2881 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2941 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     3001 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     3061 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     3121 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     3181 cgcgaatttt aacaaaatat taacgcttac aatttccatt cgccattcag gctgcgcaac
     3241 tgttgggaag ggcgatcggt gcgggcctct tcgctataac ccgccatgct acttatctac
     3301 gtaagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     3361 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     3421 caataaacaa gttactagtc ctcggctcga gatcgatgcg gtcgccatat ggatatcggt
     3481 aaccgacgtc aagcttgcat gcctgcaggt cctagtaaaa agcgcaccga agtgcgctct
     3541 agagcgtgga ctgtaccctc cacttttatc cgggcttggg accggcattg gcagtgttct
     3601 acagtccacg ccgaccgcgg ccacttgcca tgaatgatgg gacccgttcc gttcaaacac
     3661 tgttgaagca agaacgggac ccatcatcag agtatcggac cgctaccgct tagagagttt
     3721 acggctctac ttcctatcct aacccaaacc gtattattgg gctcaccttg tagcggaccg
     3781 atacacatgg caagcggccg cactcggcat ggttaggcgg tttcccgccc cctcccattt
     3841 tatccgggct tgggaccggc gagtgcggcc cgtcgactag tatatgtgct gccgaagcga
     3901 gcacggtgtt tcgtcctttc cacaagatat ataaagccaa gaaatcgaaa tactttcaag
     3961 ttacggtaag catatgatag tccattttaa aacataattt taaaactgca aactacccaa
     4021 gaaattatta ctttctacgt cacgtatttt gtactaatat ctttgtgttt acagtcaaat
     4081 taattctaat tatctctcta acagccttgt atcgtatatg caaatatgaa ggaatcatgg
     4141 gaaataggcc ctcttcctgc ccgaccttgg atccggctgt ggaatgtgtg tcagttaggg
     4201 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     4261 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     4321 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     4381 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     4441 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     4501 ctaggctttt gcaaaaagct taccatgacc gagtacaagc ccacggtgcg cctcgccacc
     4561 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     4621 acgcgccaca ccgtcgatcc tgatcgccac atcgagcggg tcaccgagct gcaagaactc
     4681 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggtgctgcc
     4741 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     4801 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc
     4861 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     4921 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag
     4981 cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag
     5041 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     5101 atgacccgca agcccggtgc ctgacgcccg ccccacgacc cgcagttcga aatgaccgac
     5161 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     5221 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     5281 atgctggagt tcttcgccca ccccgggctc gatcccctcg cgagttggtt cagctgctgc
     5341 ctgaggctgg acgacctcgc ggagttctac cggcagtgca aatccgtcgg catccaggaa
     5401 accagcagcg gctatccgcg catccatgcc cccgaactgc aggagtgggg aggcacgatg
     5461 gccgctttgg tccggatctt tgtgaaggaa ccttacttct gtggtgtgac ataattggac
     5521 aaactaccta cagagattta aagctctaag gtaaatataa aatttttaag tgtataatgt
     5581 gttaa
//
