LOCUS       sequence-243402-        6336 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Overexpression of bifunctional circular RNA (Broccoli fluorescence 
            and NFkB binding).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6336)
  AUTHORS   Litke JL, Jaffrey SR
  TITLE     Highly efficient expression of circular RNA aptamers in cells using 
            autocatalytic transcripts.
  JOURNAL   Nat Biotechnol. 2019 Apr 8. pii: 10.1038/s41587-019-0090-6. doi: 
            10.1038/s41587-019-0090-6.
  PUBMED    30962542
REFERENCE   2  (bases 1 to 6336)
  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 
            Biotechnol. 2019 Apr 8. pii: 10.1038/s41587-019-0090-6. doi: 
            10.1038/s41587-019-0090-6."
COMMENT     Sequence Label: pAV-U6+27-Tornado-F30-Broccoli-aptNFkB#5
FEATURES             Location/Qualifiers
     source          1..6336
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        25..273
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     25..45
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     196..215
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     polyA_signal    786..907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   956..1085
                     /label=AAV2 ITR
     rep_origin      complement(1179..1634)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1316..1337)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1528..1547
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1661..1765
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1766..1834,1835..2626)
                     /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(1984..2003)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2797..3385
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3286..3305
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3654..3788
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(4213..4233)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          4363..4428
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             complement(4848..5642)
                     /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(4959..4978)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5569..5588
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5999..6328)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6013..6148
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6313..6333
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 gccggatcca aggtcgggca ggaagagggc ctatttccca tgattccttc atatttgcat
       61 atacgataca aggctgttag agagataatt agaattaatt tgactgtaaa cacaaagata
      121 ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa
      181 ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg
      241 gctttatata tcttgtggaa aggacgaaac accgtgctcg cttcggcagc acatatacta
      301 gtcgacgggc cgcactcgcc ggtcccaagc ccggataaaa tgggaggggg cgggaaaccg
      361 cctaaccatg ccgagtgcgg ccgcttgcca tgtgtatcgg tccggagacg gtcgggtcca
      421 gatattcgta tctgtcgagt agagtgtggg ctccggtccg atactctgat gatgggtccc
      481 aaaagaagct tacaagaagg acagcacgaa taaaacctgc gtaaatccgc cccatttgtg
      541 taagggtagt gggtcgaatt ccgctcaaaa aaagggtccc atcattcatg gcaagtggcc
      601 gcggtcggcg tggactgtag aacactgcca atgccggtcc caagcccgga taaaagtgga
      661 gggtacagtc cacgctctag agcgcacttc ggtgcgcttt ttactaggac ctgcaggcat
      721 gcaagcttga cgtcggttac cgatatccat atggcgaccg catcgatctc gagccgagga
      781 ctagtaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
      841 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
      901 tatcttatca tgtcttacgt agataagtag catggcgggt taatcattaa ctacaaggaa
      961 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     1021 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     1081 cgcagagagg gacagatctg cagatctgct ggcgtaatag cgaagaggcc cgcaccgatc
     1141 gcccttccca acagttgcgc agcctgaatg gcgaatggaa attgtaagcg ttaatatttt
     1201 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     1261 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     1321 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     1381 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     1441 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     1501 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     1561 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     1621 gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     1681 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     1741 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     1801 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     1861 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     1921 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     1981 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     2041 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     2101 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     2161 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     2221 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     2281 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     2341 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     2401 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     2461 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     2521 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     2581 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     2641 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     2701 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     2761 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     2821 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     2881 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     2941 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     3001 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3061 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3121 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3181 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3241 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3301 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3361 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3421 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     3481 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     3541 cgagtcagtg agcgagcgcg cagagaggga gtggccaact ccatcactag gggttcctgg
     3601 aggggtggag tcgtgacgtg aattacgtca tagggttagg gaggtcctgg atcgatccag
     3661 acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat
     3721 gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata
     3781 aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg
     3841 aggtttttta aagcaagtaa aacctctaca aatgtggtat ggctgattat gatctctagt
     3901 caaggcacta tacatcaaat attccttatt aaccccttta caaattaaaa agctaaaggt
     3961 acacaatttt tgagcatagt tattaatagc agacactcta tgcctgtgtg gagtaagaaa
     4021 aaacagtatg ttatgattat aactgttatg cctacttata aaggttacag aatatttttc
     4081 cataattttc ttgtatagca gtgcagcttt ttcctttgtg gtgtaaatag caaagcaagc
     4141 aagagttcta ttactaaaca cagcatgact caaaaaactt agcaattctg aaggaaagtc
     4201 cttggggtct tctacctttc tcttcttttt tggaggagta gaatgttgag agtcagcagt
     4261 agcctcatca tcactagatg gcatttcttc tgagcaaaac aggttttcct cattaaaggc
     4321 attccaccac tgctcccatt catcagttcc ataggttgga atctaaaata cacaaacaat
     4381 tagaatcagt agtttaacac attatacact taaaaatttt atatttacct tagagcttta
     4441 aatctctgta ggtagtttgt ccaattatgt cacaccacag aagtaaggtt ccttcacaaa
     4501 gatccggacc aaagcggcca tcgtgcctcc ccactcctgc agttcggggg catggatgcg
     4561 cggatagccg ctgctggttt cctggatgcc gacggatttg cactgccggt agaactccgc
     4621 gaggtcgtcc agcctcaggc agcagctgaa ccaactcgcg aggggatcga gcccggggtg
     4681 ggcgaagaac tccagcatga gatccccgcg ctggaggatc atccagccgg cgtcccggaa
     4741 aacgattccg aagcccaacc tttcatagaa ggcggcggtg gaatcgaaat ctcgtgatgg
     4801 caggttgggc gtcgcttggt cggtcatttc gaaccccaga gtcccgctca gaagaactcg
     4861 tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg
     4921 aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct
     4981 atgtcctgat agcgcgccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg
     5041 ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg
     5101 ccgtcgggca tgcgcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc
     5161 tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg tgctcgctcg
     5221 atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt atgcagccgc
     5281 cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga tgacaggaga
     5341 tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt gacaacgtcg
     5401 agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc tgcctcgtcc
     5461 tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc
     5521 gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc ccagtcatag
     5581 ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc ttgttcaatc
     5641 atgcgaaacg atcctcatcc tgtctcttga tcagatcttg atcccctgcg ccatcagatc
     5701 cttggcggca agaaagccat ccagtttact ttgcagggct tcccaacctt accagagggc
     5761 gccccagctg gcaattccgg ttcgcttgct gtccataaaa ccgcccagtc tagctatcgc
     5821 catgtaagcc cactgcaagc tacctgcttt ctctttgcgc ttgcgttttc ccttgtccag
     5881 atagcccagt agctgacatt catccggggt cagcaccgtt tctgcggact ggctttctac
     5941 gtgttccgct tcctttagca gcccttgcgc cctgagtgct tgcggcagcg tgaagctttt
     6001 tgcaaaagcc taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggccg
     6061 aggcggcctc ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg
     6121 cggaactggg cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg
     6181 actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
     6241 acctggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg
     6301 gggactttcc acaccctaac tgacacacat tccaca
//
