LOCUS       sequence-354623-        6156 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ratiometric 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 6156)
  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 6156)
  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-F30-Broccoli-Squash-SAM-sens...
FEATURES             Location/Qualifiers
     source          1..6156
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     288..308
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        336..584
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     336..356
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     507..526
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     polyA_signal    1092..1213
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1310..1765)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1447..1468)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1659..1678
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1792..1896
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1897..1965,1966..2757)
                     /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(2115..2134)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2928..3516
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3417..3436
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3670..3687
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    3785..3919
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(4344..4364)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          4494..4559
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             complement(4979..5773)
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVAARYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5090..5109)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5700..5719
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(6130..6156,1..303))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6144..6156,1..123)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcctcctcac tacttctgga atagctcaga ggccgaggcg gcctcggcct ctgcataaat
       61 aaaaaaaatt agtcagccat ggggcggaga atgggcggaa ctgggcggag ttaggggcgg
      121 gatgggcgga gttaggggcg ggactatggt tgctgactaa ttgagatgca tgctttgcat
      181 acttctgcct gctggggagc ctggggactt tccacacctg gttgctgact aattgagatg
      241 catgctttgc atacttctgc ctgctgggga gcctggggac tttccacacc ctaactgaca
      301 cacattccac agccggatcc aaggtcgggc aggaagaggg cctatttccc atgattcctt
      361 catatttgca tatacgatac aaggctgtta gagagataat tagaattaat ttgactgtaa
      421 acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg
      481 cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt
      541 tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccgtgctc gcttcggcag
      601 cacatatact agtcgacggc catcagtcgc cggtcccaag cccggataaa atgggagggg
      661 gcgggaaacc gcctaaccat gccgactgat ggcagttgcc atgtgtatgt ggccagatgc
      721 cttgtaaccg aaagggaatc aaggtgagcc caataatacg gtttgggtta ggataggaag
      781 tagagccgta aactctctaa gcggattcga aaggatggcc acatactctg atgatcgaga
      841 cggtcgggtc cagatattcg tatctgtcga gtagagtgtg ggctcgatca ttcatggcaa
      901 ctgccatcag tcggcgtgga ctgtagaaca ctgccaatgc cggtcccaag cccggataaa
      961 agtggagggt acagtccacg ctctagagcg cacttcggtg cgctttttac taggacctgc
     1021 aggcatgcaa gcttgacgtc ggttaccgat atccatatgg cgaccgcatc gatctcgagc
     1081 cgaggactag taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     1141 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     1201 tcaatgtatc ttatcatgtc ttacgtagat aagtagcatg gcgggttata gcgaagaggc
     1261 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatgga aattgtaagc
     1321 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1381 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     1441 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     1501 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     1561 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     1621 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     1681 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     1741 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     1801 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     1861 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     1921 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     1981 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     2041 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     2101 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     2161 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     2221 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     2281 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     2341 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     2401 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     2461 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     2521 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     2581 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     2641 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     2701 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     2761 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     2821 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     2881 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     2941 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     3001 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     3061 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     3121 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3181 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     3241 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     3301 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     3361 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     3421 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     3481 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     3541 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     3601 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     3661 accgagcgca gcgagtcagt gagcgagcgc gcagagaggg agtggccaac tccatcacta
     3721 ggggttcctg gaggggtgga gtcgtgacgt gaattacgtc atagggttag ggaggtcctg
     3781 gatcgatcca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     3841 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     3901 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     3961 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta tggctgatta
     4021 tgatctctag tcaaggcact atacatcaaa tattccttat taaccccttt acaaattaaa
     4081 aagctaaagg tacacaattt ttgagcatag ttattaatag cagacactct atgcctgtgt
     4141 ggagtaagaa aaaacagtat gttatgatta taactgttat gcctacttat aaaggttaca
     4201 gaatattttt ccataatttt cttgtatagc agtgcagctt tttcctttgt ggtgtaaata
     4261 gcaaagcaag caagagttct attactaaac acagcatgac tcaaaaaact tagcaattct
     4321 gaaggaaagt ccttggggtc ttctaccttt ctcttctttt ttggaggagt agaatgttga
     4381 gagtcagcag tagcctcatc atcactagat ggcatttctt ctgagcaaaa caggttttcc
     4441 tcattaaagg cattccacca ctgctcccat tcatcagttc cataggttgg aatctaaaat
     4501 acacaaacaa ttagaatcag tagtttaaca cattatacac ttaaaaattt tatatttacc
     4561 ttagagcttt aaatctctgt aggtagtttg tccaattatg tcacaccaca gaagtaaggt
     4621 tccttcacaa agatccggac caaagcggcc atcgtgcctc cccactcctg cagttcgggg
     4681 gcatggatgc gcggatagcc gctgctggtt tcctggatgc cgacggattt gcactgccgg
     4741 tagaactccg cgaggtcgtc cagcctcagg cagcagctga accaactcgc gaggggatcg
     4801 agcccggggt gggcgaagaa ctccagcatg agatccccgc gctggaggat catccagccg
     4861 gcgtcccgga aaacgattcc gaagcccaac ctttcataga aggcggcggt ggaatcgaaa
     4921 tctcgtgatg gcaggttggg cgtcgcttgg tcggtcattt cgaaccccag agtcccgctc
     4981 agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga gcggcgatac
     5041 cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca atatcacggg
     5101 tagccaacgc tatgtcctga tagcgcgccg ccacacccag ccggccacag tcgatgaatc
     5161 cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca tgggtcacga
     5221 cgagatcctc gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg gctggcgcga
     5281 gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc atccgagtac
     5341 gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg
     5401 tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga gcaaggtgag
     5461 atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt cccgcttcag
     5521 tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg
     5581 ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg
     5641 ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt gtctgttgtg
     5701 cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg tgcaatccat
     5761 cttgttcaat catgcgaaac gatcctcatc ctgtctcttg atcagatctt gatcccctgc
     5821 gccatcagat ccttggcggc aagaaagcca tccagtttac tttgcagggc ttcccaacct
     5881 taccagaggg cgccccagct ggcaattccg gttcgcttgc tgtccataaa accgcccagt
     5941 ctagctatcg ccatgtaagc ccactgcaag ctacctgctt tctctttgcg cttgcgtttt
     6001 cccttgtcca gatagcccag tagctgacat tcatccgggg tcagcaccgt ttctgcggac
     6061 tggctttcta cgtgttccgc ttcctttagc agcccttgcg ccctgagtgc ttgcggcagc
     6121 gtgaagcttt ttgcaaaagc ctaggcctcc aaaaaa
//
