LOCUS       sequence-491549-        4366 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Generate scarless circRNA with CVB3 IRES.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4366)
  AUTHORS   Shen Y, Li B, Dong L, Tang W, Ren J, Chen F, Zheng W, Yu Y, Gao L, 
            Wei W
  TITLE     Self-splicing RNA circularization facilitated by intact group I and 
            II introns.
  JOURNAL   Nat Commun. 2025 Aug 10;16(1):7376. doi: 10.1038/s41467-025-62607-y.
  PUBMED    40784880
REFERENCE   2  (bases 1 to 4366)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2025-08-10"; volume: "16(1):7376. doi"; pages: " 
            10.1038/s41467-025-62607-y"
COMMENT     Sequence Label: CIRC-Scarless-Ana-CVB3 IRES split 403-mCherry
FEATURES             Location/Qualifiers
     source          1..4366
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        429..447
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1035..1745
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1180..1198)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1448..1467
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2190..2206)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2190..2206)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2203..2225)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2214..2230
                     /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(2238..2244,2245..2262,2263..2268))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2283..2304
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2421..2438)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2592..3180)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2672..2691)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3351..4166)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3351..4166)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4074..4093
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(4167..4271)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4339..4357
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gacgcgtatt gggatcgagc
      421 tcggtaccta atacgactca ctataggaaa taattggacc ttaaagaaga aattctttaa
      481 gtggatgctc tcaaactcag ggaaacctaa atctagttat agacaaggca atcctgagcc
      541 aagccgaagt agtaattagt aagttaacaa tagatgactt acaactaatc ggaaggtgca
      601 gagactcgac gggagctacc ctaacgtcaa gacgagggta aagagagagt ccaattctca
      661 aagccaatag gcagtagcga aagctgcaag agaatgaata cagacatggt gcgaagagtc
      721 tattgagcta gttggtagtc ctccggcccc tgaatgcggc taatcctaac tgcggagcac
      781 acaccctcaa gccagagggc agtgtgtcgt aacgggcaac tctgcagcgg aaccgactac
      841 tttgggtgtc cgtgtttcat tttattccta tactggctgc ttatggtgac aattgagaga
      901 tcgttaccat atagctattg gattggccat ccggtgacta atagagctat tatatatccc
      961 tttgttgggt ttataccact tagcttgaaa gaggttaaaa cattacaatt cattgttaag
     1021 ttgaatacag caaaatggtg agcaagggcg aggaggataa catggccatc atcaaggagt
     1081 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
     1141 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     1201 gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
     1261 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
     1321 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
     1381 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
     1441 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
     1501 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
     1561 gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
     1621 gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg
     1681 tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca
     1741 agtaattaaa acagcctgtg ggttgatccc acccacaggc ccattgggcg ctagcactct
     1801 ggtatcacgg tacctttgtg cgcctgtttt ataccccctc ccccaactgt aacttagaag
     1861 taacacacac cgatcaacag tcagcgtggc acaccagcca cgttttgatc aagcacttct
     1921 gttaccccgg actgagtatc aatagactgc tcacgcggtt gaaggagaaa gcgttcgtta
     1981 tccggccaac tacttcgaaa aacctagtaa caccgtggaa gttgcagagt gtttcgctca
     2041 gcactacccc agtgtagatc aggtcgatga gtcaccgcat tccccacggg cgaccgtggc
     2101 ggtggctgcg ttggcggcct gcccatgggg aaacccatgg gacgctcttg agacggttta
     2161 aacggcgcgc cgagcttggc tcgagcatgg tcatagctgt ttcctgtgtg aaattgttat
     2221 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     2281 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     2341 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     2401 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     2461 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     2521 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     2581 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     2641 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     2701 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     2761 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     2821 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     2881 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     2941 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     3001 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     3061 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     3121 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     3181 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     3241 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     3301 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttagaaaaac
     3361 tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt
     3421 tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca
     3481 agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc
     3541 ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt
     3601 gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc
     3661 tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg
     3721 agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg
     3781 cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat
     3841 acctggaatg ctgttttccc agggatcgca gtggtgagta accatgcatc atcaggagta
     3901 cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc
     3961 atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc
     4021 gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga
     4081 gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg cctagagcaa
     4141 gacgtttccc gttgaatatg gctcatactc ttcctttttc aatattattg aagcatttat
     4201 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     4261 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc
     4321 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtc
//
