LOCUS       sequence-420921-        7123 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the expression of a split nLuc that produces luminescence only 
            when circularized. To test your IRES of interest, clone into the 
            EcoRI, BsiWI sites..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7123)
  AUTHORS   Unti MJ, Jaffrey SR
  TITLE     Highly efficient cellular expression of circular mRNA enables 
            prolonged protein expression.
  JOURNAL   Cell Chem Biol. 2023 Oct 13:S2451-9456(23)00334-3. doi: 
            10.1016/j.chembiol.2023.09.015.
  PUBMED    37883972
REFERENCE   2  (bases 1 to 7123)
  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. 2023 Oct 13:S2451-9456(23)00334-3. doi: 
            10.1016/j.chembiol.2023.09.015."
COMMENT     Sequence Label: pcDNA3.1+-Tornado-split-nLuc
FEATURES             Location/Qualifiers
     source          1..7123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             901..924
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             937..963
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             937..963
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        988..1006
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1126..1155
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2046..2069
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2676..2694)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2677..2694)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2714..2731)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2720..2944
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2990..3418
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3077..3096)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3287..3308
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3427..3447)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3432..3761
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3612..3747
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3674..3693
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3828..4622
                     /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(3882..3901)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4492..4511
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4796..4917
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4833..4852)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4887..4906
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4966..4982)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4966..4982)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4979..5001)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4990..5006
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5014..5020,5021..5038,5039..5044))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5059..5080
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5197..5214)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5368..5956)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5448..5467)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6127..6918,6919..6987))
                     /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     6750..6769
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6988..7092)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttaccatg
      901 gactacaagg acgacgatga caagctcgat ggaggatacc cctacgacgt gcccgactac
      961 gccagcggat ccagctaaac tcgatgataa tacgactcac tataggccgc actcgccggt
     1021 cccaagcccg gataaaatgg gagggggcgg gaaaccgcct aaccatgccg agtgcggccg
     1081 catatatgga tagtgaccgg ctaccggctg ttcgaggaga ttctggaaca aaaactcatc
     1141 tcagaagagg atctggaaga gtgagaattc aatcatctac aaggtgagcc caataatacg
     1201 gtttgggtta ggataggaag tagagccgta aactctctaa gcggtagaaa acaaaaaaca
     1261 aaaaaaacaa aaaaaaacca aaaaaacaaa acacattaaa acagcctgtg ggttgatccc
     1321 acccacaggc ccattgggcg ctagcactct ggtatcacgg tacctttgtg cgcctgtttt
     1381 ataccccctc ccccaactgt aacttagaag taacacacac cgatcaacag tcagcgtggc
     1441 acaccagcca cgttttgatc aagcacttct gttaccccgg actgagtatc aatagactgc
     1501 tcacgcggtt gaaggagaaa gcgttcgtta tccggccaac tacttcgaaa aacctagtaa
     1561 caccgtggaa gttgcagagt gtttcgctca gcactacccc agtgtagatc aggtcgatga
     1621 gtcaccgcat tccccacggg cgaccgtggc ggtggctgcg ttggcggtct gcccatgggg
     1681 aaacccatgg gacgctctaa tacagacatg gtgcgaagag tctattgagc tagttggtag
     1741 tcctccggcc cctgaatgcg gctaatccta actgcggagc acacaccctc aagccagagg
     1801 gcagtgtgtc gtaacgggca actctgcagc ggaaccgact actttgggtg tccgtgtttc
     1861 attttattcc tatactggct gcttatggtg acaattgaga gatcgttacc atatagctat
     1921 tggattggcc atccggtgac taatagagct attatatatc cctttgttgg gtttatacca
     1981 cttagcttga aagaggttaa aacattacaa ttcattgtta agttgaatac agcaaaatgc
     2041 gtacggacta caaggacgac gatgacaagg tcttcacact cgaagatttc gttggggact
     2101 gggaacagac agccgcctac aacctggacc aagtccttga acagggaggt gtgtccagtt
     2161 tgctgcagaa tctcgccgtg tccgtaactc cgatccaaag gattgtccgg agcggtgaaa
     2221 atgccctgaa gatcgacatc catgtcatca tcccgtatga aggtctgagc gccgaccaaa
     2281 tggcccagat cgaagaggtg tttaaggtgg tgtaccctgt ggatgatcat cactttaagg
     2341 tgatcctgcc ctatggcaca ctggtaatcg acggggttac gccgaacatg ctgaactatt
     2401 tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa aaagatcact gtaacaggga
     2461 ccctgtggaa cggcaacaaa attatcgacg agcgcctgat cacccccgac ggctccatgc
     2521 tgttccgagt aaccatcaac agctctagag ccatgtctat gtggccgcgg tcggcgtgga
     2581 ctgtagaaca ctgccaatgc cggtcccaag cccggataaa agtggagggt acagtccacg
     2641 ccggtccgct cgagcatgca tctagagggc cctattctat agtgtcacct aaatgctaga
     2701 gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     2761 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2821 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2881 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     2941 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     3001 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     3061 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3121 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3181 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     3241 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3301 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     3361 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     3421 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3481 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     3541 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3601 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3661 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3721 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3781 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     3841 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     3901 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     3961 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     4021 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     4081 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     4141 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     4201 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     4261 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     4321 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     4381 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     4441 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     4501 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     4561 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     4621 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     4681 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     4741 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     4801 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4861 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     4921 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     4981 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     5041 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5101 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5161 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5221 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5281 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5341 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5401 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5461 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5521 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5581 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5641 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5701 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5761 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5821 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5881 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5941 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6001 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6061 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6121 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6181 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6241 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6301 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6361 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6421 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6481 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6541 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6601 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6661 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6721 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6781 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6841 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6901 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6961 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     7021 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     7081 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
