LOCUS       sequence-364014-        6236 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses Squash-SAM sensor 5-1 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 6236)
  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 6236)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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-SAM-sensor 5-1
FEATURES             Location/Qualifiers
     source          1..6236
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(38..58)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(86..106)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        91..420
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      271..406
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             777..1571
                     /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(831..850)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1441..1460
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     intron          1991..2056
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             2186..2206
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2631..2765
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(3034..3622)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3114..3133)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3793..4584,4585..4653))
                     /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     4416..4435
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4654..4758)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4785..5240
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4872..4891)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5082..5103
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     repeat_region   5334..5463
                     /label=AAV2 ITR
     polyA_signal    5512..5633
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        complement(join(6046..6236,1..58))
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(6104..6123)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 ctctaacagc cttgtatcgt atatgcaaat atgaaggaat catgggaaat aggccctctt
       61 cctgcccgac cttggatccg gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
      121 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
      181 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
      241 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
      301 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
      361 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
      421 aagcttcacg ctgccgcaag cactcagggc gcaagggctg ctaaaggaag cggaacacgt
      481 agaaagccag tccgcagaaa cggtgctgac cccggatgaa tgtcagctac tgggctatct
      541 ggacaaggga aaacgcaagc gcaaagagaa agcaggtagc ttgcagtggg cttacatggc
      601 gatagctaga ctgggcggtt ttatggacag caagcgaacc ggaattgcca gctggggcgc
      661 cctctggtaa ggttgggaag ccctgcaaag taaactggat ggctttcttg ccgccaagga
      721 tctgatggcg caggggatca agatctgatc aagagacagg atgaggatcg tttcgcatga
      781 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
      841 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
      901 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
      961 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     1021 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     1081 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     1141 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     1201 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     1261 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     1321 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     1381 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     1441 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     1501 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     1561 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     1621 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     1681 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     1741 ccccgggctc gatcccctcg cgagttggtt cagctgctgc ctgaggctgg acgacctcgc
     1801 ggagttctac cggcagtgca aatccgtcgg catccaggaa accagcagcg gctatccgcg
     1861 catccatgcc cccgaactgc aggagtgggg aggcacgatg gccgctttgg tccggatctt
     1921 tgtgaaggaa ccttacttct gtggtgtgac ataattggac aaactaccta cagagattta
     1981 aagctctaag gtaaatataa aatttttaag tgtataatgt gttaaactac tgattctaat
     2041 tgtttgtgta ttttagattc caacctatgg aactgatgaa tgggagcagt ggtggaatgc
     2101 ctttaatgag gaaaacctgt tttgctcaga agaaatgcca tctagtgatg atgaggctac
     2161 tgctgactct caacattcta ctcctccaaa aaagaagaga aaggtagaag accccaagga
     2221 ctttccttca gaattgctaa gttttttgag tcatgctgtg tttagtaata gaactcttgc
     2281 ttgctttgct atttacacca caaaggaaaa agctgcactg ctatacaaga aaattatgga
     2341 aaaatattct gtaaccttta taagtaggca taacagttat aatcataaca tactgttttt
     2401 tcttactcca cacaggcata gagtgtctgc tattaataac tatgctcaaa aattgtgtac
     2461 ctttagcttt ttaatttgta aaggggttaa taaggaatat ttgatgtata gtgccttgac
     2521 tagagatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     2581 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     2641 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     2701 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     2761 ggatcgatcc aggacctccc taaccctatg acgtaattca cgtcacgact ccacccctcc
     2821 aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctca ctgactcgct
     2881 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     2941 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3001 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3061 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3121 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3181 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3241 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3301 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3361 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3421 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3481 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3541 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3601 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3661 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3721 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3781 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3841 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3901 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     3961 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4021 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     4081 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     4141 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     4201 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4261 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4321 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4381 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4441 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4501 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4561 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4621 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4681 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4741 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg
     4801 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     4861 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     4921 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     4981 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     5041 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     5101 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     5161 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     5221 aatattaacg cttacaattt ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     5281 tcggtgcggg cctcttcgct attacgccag cagatctgca gatctgtccc tctctgcgcg
     5341 ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc tttggtcgcc
     5401 cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc actaggggtt
     5461 ccttgtagtt aatgattaac ccgccatgct acttatctac gtaagacatg ataagataca
     5521 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     5581 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttactagtc
     5641 ctcggctcga gatcgatgcg gtcgccatat ggatatcggt aaccgacgtc aagcttgcat
     5701 gcctgcaggt cctagtaaaa agcggaccga agtccgctct agagcgtgga ctgtaccctc
     5761 cacttttatc cgggcttggg accggcattg gcagtgttct acagtccacg ccgaccgcgg
     5821 ccaccgccat cctttcgaat ccgcttagag agtttacggc tctacttcct atcctaaccc
     5881 aaaccgtatt attgggctca ccttgattcc ctttcggtta caaggcatct ggcggcggcc
     5941 gcactcggca tggttaggcg gtttcccgcc ccctcccatt ttatccgggc ttgggaccgg
     6001 cgagtgcggc ccgtcgacta gtatatgtgc tgccgaagcg agcacggtgt ttcgtccttt
     6061 ccacaagata tataaagcca agaaatcgaa atactttcaa gttacggtaa gcatatgata
     6121 gtccatttta aaacataatt ttaaaactgc aaactaccca agaaattatt actttctacg
     6181 tcacgtattt tgtactaata tctttgtgtt tacagtcaaa ttaattctaa ttatct
//
