LOCUS       sequence-354604-        6197 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Squash aptamer as circular RNA in mammalian cells using 
            the Tornado expression system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6197)
  AUTHORS   Dey SK, Filonov GS, Olarerin-George AO, Jackson BT, Finley LWS, 
            Jaffrey SR
  TITLE     Repurposing an adenine riboswitch into a fluorogenic imaging and 
            sensing tag.
  JOURNAL   Nat Chem Biol. 2021 Dec 22. pii: 10.1038/s41589-021-00925-0. doi: 
            10.1038/s41589-021-00925-0.
  PUBMED    34937909
REFERENCE   2  (bases 1 to 6197)
  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: "Nat Chem 
            Biol. 2021 Dec 22. pii: 10.1038/s41589-021-00925-0. doi: 
            10.1038/s41589-021-00925-0."
COMMENT     Sequence Label: pAV-U6+27-Tornado-Squash
FEATURES             Location/Qualifiers
     source          1..6197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     42..62
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        90..338
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     90..110
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     261..280
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     polyA_signal    712..833
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   882..1011
                     /label=AAV2 ITR
     rep_origin      complement(1105..1560)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1242..1263)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1454..1473
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1587..1691
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1692..1760,1761..2552)
                     /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(1910..1929)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2723..3311
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3212..3231
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3580..3714
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(4139..4159)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          4289..4354
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             complement(4774..5568)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4885..4904)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5495..5514
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(5925..6197,1..57))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5939..6074
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 ttgcatactt ctgcctgctg gggagcctgg ggactttcca caccctaact gacacacatt
       61 ccacagccgg atccaaggtc gggcaggaag agggcctatt tcccatgatt ccttcatatt
      121 tgcatatacg atacaaggct gttagagaga taattagaat taatttgact gtaaacacaa
      181 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
      241 taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt
      301 tcttggcttt atatatcttg tggaaaggac gaaacaccgt gctcgcttcg gcagcacata
      361 tactagtcga cggccatcag tcgccggtcc caagcccgga taaaatggga gggggcggga
      421 aaccgcctaa ccatgccgac tgatggcagc tacaaggtga gcccaataat acggtttggg
      481 ttaggatagg aagtagagcc gtaaactctc taagcggtag ctgccatcag tcggcgtgga
      541 ctgtagaaca ctgccaatgc cggtcccaag cccggataaa agtggagggt acagtccacg
      601 ctctagagcg gacttcggtc cgctttttac taggacctgc aggcatgcaa gcttgacgtc
      661 ggttaccgat atccatatgg cggccgcatc gatctcgagc cgcggactag taacttgttt
      721 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
      781 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
      841 ttacgtagat aagtagcatg gcgggttaat cattaactac aaggaacccc tagtgatgga
      901 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc
      961 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gagagggaca
     1021 gatctgcaga tctgctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     1081 ttgcgcagcc tgaatggcga atggaaattg taagcgttaa tattttgtta aaattcgcgt
     1141 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     1201 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     1261 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     1321 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     1381 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     1441 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     1501 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     1561 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     1621 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1681 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1741 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1801 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1861 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     1921 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     1981 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     2041 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     2101 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     2161 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     2221 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     2281 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     2341 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     2401 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     2461 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     2521 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     2581 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     2641 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2701 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2761 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2821 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     2881 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2941 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3001 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3061 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3121 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3181 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3241 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3301 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3361 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3421 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3481 agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt
     3541 gacgtgaatt acgtcatagg gttagggagg tcctggatcg atccagacat gataagatac
     3601 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     3661 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     3721 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc
     3781 aagtaaaacc tctacaaatg tggtatggct gattatgatc tctagtcaag gcactataca
     3841 tcaaatattc cttattaacc cctttacaaa ttaaaaagct aaaggtacac aatttttgag
     3901 catagttatt aatagcagac actctatgcc tgtgtggagt aagaaaaaac agtatgttat
     3961 gattataact gttatgccta cttataaagg ttacagaata tttttccata attttcttgt
     4021 atagcagtgc agctttttcc tttgtggtgt aaatagcaaa gcaagcaaga gttctattac
     4081 taaacacagc atgactcaaa aaacttagca attctgaagg aaagtccttg gggtcttcta
     4141 cctttctctt cttttttgga ggagtagaat gttgagagtc agcagtagcc tcatcatcac
     4201 tagatggcat ttcttctgag caaaacaggt tttcctcatt aaaggcattc caccactgct
     4261 cccattcatc agttccatag gttggaatct aaaatacaca aacaattaga atcagtagtt
     4321 taacacatta tacacttaaa aattttatat ttaccttaga gctttaaatc tctgtaggta
     4381 gtttgtccaa ttatgtcaca ccacagaagt aaggttcctt cacaaagatc cggaccaaag
     4441 cggccatcgt gcctccccac tcctgcagtt cgggggcatg gatgcgcgga tagccgctgc
     4501 tggtttcctg gatgccgacg gatttgcact gccggtagaa ctccgcgagg tcgtccagcc
     4561 tcaggcagca gctgaaccaa ctcgcgaggg gatcgagccc ggggtgggcg aagaactcca
     4621 gcatgagatc cccgcgctgg aggatcatcc agccggcgtc ccggaaaacg attccgaagc
     4681 ccaacctttc atagaaggcg gcggtggaat cgaaatctcg tgatggcagg ttgggcgtcg
     4741 cttggtcggt catttcgaac cccagagtcc cgctcagaag aactcgtcaa gaaggcgata
     4801 gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa agcacgagga agcggtcagc
     4861 ccattcgccg ccaagctctt cagcaatatc acgggtagcc aacgctatgt cctgatagcg
     4921 gtccgccaca cccagccggc cacagtcgat gaatccagaa aagcggccat tttccaccat
     4981 gatattcggc aagcaggcat cgccatgggt cacgacgaga tcctcgccgt cgggcatgcg
     5041 cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc tgatgctctt cgtccagatc
     5101 atcctgatcg acaagaccgg cttccatccg agtacgtgct cgctcgatgc gatgtttcgc
     5161 ttggtggtcg aatgggcagg tagccggatc aagcgtatgc agccgccgca ttgcatcagc
     5221 catgatggat actttctcgg caggagcaag gtgagatgac aggagatcct gccccggcac
     5281 ttcgcccaat agcagccagt cccttcccgc ttcagtgaca acgtcgagca cagctgcgca
     5341 aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc tcgtcctgca gttcattcag
     5401 ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc ccctgcgctg acagccggaa
     5461 cacggcggca tcagagcagc cgattgtctg ttgtgcccag tcatagccga atagcctctc
     5521 cacccaagcg gccggagaac ctgcgtgcaa tccatcttgt tcaatcatgc gaaacgatcc
     5581 tcatcctgtc tcttgatcag atcttgatcc cctgcgccat cagatccttg gcggcaagaa
     5641 agccatccag tttactttgc agggcttccc aaccttacca gagggcgccc cagctggcaa
     5701 ttccggttcg cttgctgtcc ataaaaccgc ccagtctagc tatcgccatg taagcccact
     5761 gcaagctacc tgctttctct ttgcgcttgc gttttccctt gtccagatag cccagtagct
     5821 gacattcatc cggggtcagc accgtttctg cggactggct ttctacgtgt tccgcttcct
     5881 ttagcagccc ttgcgccctg agtgcttgcg gcagcgtgaa gctttttgca aaagcctagg
     5941 cctccaaaaa agcctcctca ctacttctgg aatagctcag aggccgaggc ggcctcggcc
     6001 tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga actgggcgga
     6061 gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta attgagatgc
     6121 atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct ggttgctgac
     6181 taattgagat gcatgct
//
