LOCUS       sequence-374481-        5952 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template plasmid to transcribe mRNA encoding mNeonGreen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5952)
  AUTHORS   Chen R, Wang SK, Belk JA, Amaya L, Li Z, Cardenas A, Abe BT, Chen 
            CK, Wender PA, Chang HY
  TITLE     Engineering circular RNA for enhanced protein production.
  JOURNAL   Nat Biotechnol. 2022 Jul 18. pii: 10.1038/s41587-022-01393-0. doi: 
            10.1038/s41587-022-01393-0.
  PUBMED    35851375
REFERENCE   2  (bases 1 to 5952)
  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. 2022 Jul 18. pii: 10.1038/s41587-022-01393-0. doi: 
            10.1038/s41587-022-01393-0."
COMMENT     Sequence Label: mRNA-CleanCap-mNeonGreen
FEATURES             Location/Qualifiers
     source          1..5952
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        655..958
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          1236..2252
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2176..2198
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2260..2279
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2380..2389
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     terminator      3243..3290
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    3304..3528
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3571..3587
                     /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)."
     primer_bind     3576..3598
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3595..3611
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3595..3611
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    3660..3781
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3671..3690)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3725..3744
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3941..4540)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4024..4044)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     4521..4540
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(4551..4879)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4565..4700
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4619..4638)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             complement(4913..5728)
                     /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(4913..5728)
                     /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     5636..5655
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5720..5739
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaac ctgcaggtta
      601 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
      661 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
      721 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
      781 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
      841 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
      901 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
      961 cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat
     1021 tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggnns
     1081 gsgcgccrgg sggggsgggg sggggsgrgg ggsggggsgg ggsgrggcgg agaggtgcgg
     1141 cggcagccaa tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc
     1201 ggcggcccta taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc
     1261 ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac
     1321 tcccacaggt gagcgggcgg gacggccctt ctcctccggg ctgtaattag cgcttggttt
     1381 aatgacggct tgtttctttt ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc
     1441 ctttgtgcgg ggggagcggc tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg
     1501 cgtgcggctc cgcgctgccc ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg
     1561 cgctccgcag tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg
     1621 gctgcgaggg gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg
     1681 tgggcgcgtc ggtcgggctg caaccccccc tgcacccccc tccccgagtt gctgagcacg
     1741 gcccggcttc gggtgcgggg ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg
     1801 gggggtggcg gcaggtgggg gtgccgggcg gggcggggcc gcctcgggcc ggggagggct
     1861 cgggggaggg gcgcggcggc ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca
     1921 gccattgcct tttatggtaa tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct
     1981 gtgcggagcc gaaatctggg aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc
     2041 ggtgcggcgc cggcaggaag gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc
     2101 gtccccttct ccctctccag cctcggggct gtccgcgggg ggacggctgc cttcgggggg
     2161 gacggggcag ggcggggttc ggcttctggc gtgtgaccgg cggctctaga gcctctgcta
     2221 accatgttca tgccttcttc tttttcctac agctcctggg caacgtgctg gttattgtgc
     2281 tgtctcatca ttttggcaaa gcggccgcgg ccagtgaatt gtaatacgac tcactataag
     2341 gaaataagag agaaaagaag agtaagaaga aatataagag ccaccatggg atccgtgtct
     2401 aaaggagagg aagataacat ggcgagcctc ccagccactc atgaactcca catattcgga
     2461 agcatcaacg gggtggactt tgacatggta ggtcagggga ccggcaatcc caacgacggt
     2521 tacgaggaat tgaatctcaa gtccactaag ggcgacctcc agttttctcc gtggattctc
     2581 gtcccacata tcgggtatgg tttccaccaa tacttgcctt acccggatgg aatgtcaccg
     2641 ttccaggctg ccatggttga tggctctggt tatcaggtac acagaacgat gcaatttgag
     2701 gacggagcgt cattgactgt aaactacaga tacacatacg aaggctctca cataaagggc
     2761 gaagcacaag ttaaaggcac gggattcccc gctgatggtc ctgtcatgac aaattctctt
     2821 accgcagccg actggtgtcg atccaagaaa acgtatccca atgacaaaac tatcatatct
     2881 acgttcaaat ggtcatatac aacgggaaac ggtaagagat accggagcac tgccagaacc
     2941 acctatacat ttgccaaacc aatggcggct aactacctga agaaccagcc catgtatgtg
     3001 ttcaggaaaa cggaactgaa acactccaag acagagctga atttcaagga atggcaaaaa
     3061 gccttcacgg atgttatggg gatggatgag ctgtataaat agtaatgagc tggagcctcg
     3121 gtggccatgc ttcttgcccc ttgggcctcc ccccagcccc tcctcccctt cctgcacccg
     3181 tacccccgtg gtctttgaat aaagtctgaa aaaaaaaaaa aaaaaaaaaa aaaagcggcc
     3241 gcctagcata accccttggg gcctctaaac gggtcttgag gggttttttg gctagcggcc
     3301 ggcctgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     3361 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     3421 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     3481 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggcg gcgcgccact
     3541 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     3601 acagctatga ccatgattac gccaagctct agctagaggt cgacggtata cagacatgat
     3661 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     3721 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     3781 tggggtgggc gaagaactcc agcatgagat ccccgcgctg gaggatcatc cagccggcgt
     3841 cccggaaaac gattccgaag cccaaccttt catagaaggc ggcggtggaa tcgaaatctc
     3901 gtgatggcag gttgggcgtc gcttggtcgg tcatttcgaa tcaggcaccg ggcttgcggg
     3961 tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc ggcggtgacg gtgaagccga
     4021 gccgctcgta gaaggggagg ttgcggggcg cggaggtttc caggaaggcg ggcaccccgg
     4081 cgcgctcggc cgcctccact ccggggagca cgacggcgct gcccagaccc ttgccctggt
     4141 ggtcgggcga cacgccgacg gtggccagga accacgcggg ctccttgggc cggtgcggcg
     4201 ccaggaggcc ttccatctgt tgctgcgcgg ccagccggga accgctcaac tcggccatgc
     4261 gcgggccgat ctcggcgaac accgcccccg cttcgacgct ctccggcgtg gtccagaccg
     4321 ccaccgcggc gccgtcgtcc gcgacccaca ccttgccgat gtcgagcccg acgcgcgtga
     4381 ggaagagttc ttgcagctcg gtgacccgct cgatgtggcg gtccggatcg acggtgtggc
     4441 gcgtggcggg gtagtcggcg aacgcggcgg cgagggtgcg tacggccctg gggacgtcgt
     4501 cgcgggtggc gaggcgcacc gtgggcttgt actcggtcat gcgggagctt tttgcaaaag
     4561 cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc agaggcggcc
     4621 tcggcctctg cataaataaa aaaaattagt cagccatggg gcggagaatg ggcggaactg
     4681 ggcggagtta ggggcgggat gggcggagtt aggggcggga ctatggttgc tgactaattg
     4741 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acacctggtt
     4801 gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt
     4861 ccacacccta actgacacag agctcgtgtt acaaccaatt aaccaattct gattagaaaa
     4921 actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt
     4981 tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg
     5041 caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt
     5101 tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg
     5161 gtgagaatgg caaaagctta tgcatttctt tccagacttg ttcaacaggc cagccattac
     5221 gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag
     5281 cgaggcgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc
     5341 ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta
     5401 atacctggaa tgctgttttc ccggggatcg cagtggtgag taaccatgca tcatcaggag
     5461 tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga
     5521 ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg
     5581 gcgcatcggg cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc
     5641 gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctggagc
     5701 aagacgtttc ccgttgaata tggctcataa caccccttgt attactgttt atgtaagcag
     5761 acagttttat tgttcatgat gatatatttt tatcttgtgc aatgtaacat cagagatttt
     5821 gagacacaac gtggctttgt tgaataaatc gaacttttgc tgagttgaag gatcagtcat
     5881 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     5941 caaaggatct tc
//
