LOCUS       sequence-406177-        5255 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expressing cloned exons as circular RNA. Alternatively, for 
            expressing cDNA (if the EcoRI-XhoI restriction fragment is also 
            removed)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5255)
  AUTHORS   Stringer BW, Gabryelska M, Marri S, Clark L, Lin H, Gantley L, Liu 
            R, Wilusz JE, Conn VM, Conn SJ
  TITLE     Versatile toolkit for highly-efficient and scarless overexpression 
            of circular RNAs.
  JOURNAL   bioRxiv. 2023 Nov 22:2023.11.21.568171. doi: 
            10.1101/2023.11.21.568171. Preprint.
  PUBMED    38045421
REFERENCE   2  (bases 1 to 5255)
  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: "bioRxiv. 
            2023 Nov 22:2023.11.21.568171. doi: 10.1101/2023.11.21.568171. 
            Preprint."
COMMENT     Sequence Label: mc2 hygro EF1a
FEATURES             Location/Qualifiers
     source          1..5255
                     /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"
     promoter        234..445
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     primer_bind     complement(714..731)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    720..944
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      990..1418
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1077..1096)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1287..1308
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1427..1447)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1432..1761
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1612..1747
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1674..1693
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2928..3049
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2965..2984)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3019..3038
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3098..3114)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3098..3114)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3111..3133)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3122..3138
                     /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(3146..3152,3153..3170,3171..3176))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3191..3212
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3329..3346)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3500..4088)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3580..3599)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4259..5050,5051..5119))
                     /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     4882..4901
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5120..5224)
                     /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 cgtgggcaga
      241 gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gatccggtgc
      301 ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt
      361 tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg
      421 caacgggttt gccgccagaa cacaggctag cgtttaaact taagcttggt accgagctcg
      481 gatccactag tccagtgtgg tggaattcaa agtgctgaga ttacaggcgt gagccaccac
      541 ccccggccca ctttttgtaa aggtacgtac taatgacttt ttttttatac ttcagggaga
      601 cggttgtgta cgtctcggta agaagcaagg aaaagaatta ggctcggcac ggtagctcac
      661 acctgtaatc ccagcactcg agtctagagg gcccgtttaa acccgctgat cagcctcgac
      721 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
      781 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
      841 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
      901 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
      961 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     1021 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     1081 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     1141 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     1201 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     1261 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     1321 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     1381 aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag
     1441 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     1501 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     1561 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     1621 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     1681 gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt
     1741 ggaggcctag gcttttgcaa aaagctcccg ggaccggtgc caccatgaaa aagcctgaac
     1801 tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtg tccgacctga
     1861 tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga gggcgtggat
     1921 atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa agatcgttat gtttatcggc
     1981 actttgcatc ggccgcgctc ccgattccgg aagtgcttga cattggggaa ttcagcgaga
     2041 gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa
     2101 ccgaactgcc cgctgttctg cagccggtcg cggaggccat ggatgcgatc gctgcggccg
     2161 atcttagcca gacgagcggg ttcggcccat tcggaccgca aggaatcggt caatacacta
     2221 catggcgtga tttcatatgc gcgattgctg atccccatgt gtatcactgg caaactgtga
     2281 tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg ctttgggccg
     2341 aggactgccc cgaagtccgg cacctcgtgc acgcggattt cggctccaac aatgtcctga
     2401 cggacaatgg ccgcataaca gcggtcattg actggagcga ggcgatgttc ggggattccc
     2461 aatacgaggt cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga
     2521 cgcgctactt cgagcggagg catccggagc ttgcaggatc gccgcggctc cgggcgtata
     2581 tgctccgcat tggtcttgac caactctatc agagcttggt tgacggcaat ttcgatgatg
     2641 cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc
     2701 gtacacaaat cgcccgcaga agcgcggccg tctggaccga tggctgtgta gaagtacttc
     2761 gccgatagtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     2821 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     2881 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     2941 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     3001 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     3061 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     3121 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     3181 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     3241 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     3301 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     3361 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     3421 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     3481 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     3541 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     3601 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3661 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     3721 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3781 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3841 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     3901 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     3961 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     4021 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     4081 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4141 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     4201 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     4261 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     4321 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4381 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     4441 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4501 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4561 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4621 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     4681 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4741 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     4801 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     4861 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     4921 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     4981 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     5041 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     5101 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5161 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     5221 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
