LOCUS       sequence-392992-        4910 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid IVT template encoding RFP. NOTE: In the manuscript, 
            ""mCherry"" was listed as the internal control for multiple 
            fluorescence-based experiments, but it was actually RFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4910)
  AUTHORS   Aditham A, Shi H, Guo J, Zeng H, Zhou Y, Wade SD, Huang J, Liu J, 
            Wang X
  TITLE     Chemically Modified mocRNAs for Highly Efficient Protein Expression 
            in Mammalian Cells.
  JOURNAL   ACS Chem Biol. 2022 Dec 16;17(12):3352-3366. doi: 
            10.1021/acschembio.1c00569. Epub 2022 Jan 7.
  PUBMED    34995053
REFERENCE   2  (bases 1 to 4910)
  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"; doi: 
            "10.1021/acschembio.1c00569"; journalName: "ACS Chem Biol"; date: 
            "2022-12-16- 16"; volume: "17"; issue: "12"; pages: "3352-3366"
COMMENT     Sequence Label: pCS2_RFP-60A
FEATURES             Location/Qualifiers
     source          1..4910
                     /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"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1010..1030
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1012..1030
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     polyA_signal    1197..1331
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1234..1253)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1288..1307
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(1468..2142)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSENVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(1527..1550)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(1725..1744)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1994..2012
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     promoter        complement(2186..2204)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2187..2204)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     promoter        complement(2239..3223)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     rep_origin      3294..3749
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3381..3400)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3591..3612
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3775..3879
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3880..3948,3949..4740)
                     /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(4098..4117)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, 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 ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgctccc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     1021 tcactaaagg gaacaaaagc tgggtaccgg gcccaatgca ttggcgccgc ggccgcgaac
     1081 gtctcttttt tttttttttt tttttttttt tttttttttt tttttttttt tttttttttt
     1141 tcctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     1201 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     1261 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     1321 atgtctggat ctacgtaata cgactcacta tagttctaga ggctcgagtt acaaaatgct
     1381 acacttcgtt tttgatttct tcttcttttt gccgtctttg ctcatcttct ctttgtgctt
     1441 gcggatttct cggaccaacg tgaattcggc gccggtggag tggcggccct cggcgcgctc
     1501 gtactgttcc acgatggtgt agtcctcgtt gtgggaggtg atgtccagct tgatgtcggt
     1561 cttgtaggcg ccgggcagct gcacgggctt cttggccatg taggtggtct tgacctcggc
     1621 gtcgtagtgg ccgccgtcct tcagcttcag cctcatcttg atctcgccct tcagggcgcc
     1681 gtcctcgggg tacatccgct cggtggaggc ctcccagccc atggtcttct tctgcattac
     1741 ggggccgtcg gaggggaagt tggtgccgcg cagcttcacc ttgtagatga actcgccgtc
     1801 ctgcagggag gagtcctggg tcacggtcac cacgccgccg tcctcgaagt tcatcacgcg
     1861 ctcccacttg aagccctcgg ggaaggacag cttcaagtag tcggggatgt cggcggggtg
     1921 cttcacgtag gccttggagc cgtactggaa ctgaggggac aggatgtccc aggcgaaggg
     1981 cagggggccg cccttggtca ccttcagctt ggcggtctgg gtgccctcgt aggggcggcc
     2041 ctcgccctcg ccctcgatct cgaactcgtg gccgttcacg gagccctcca tgcgcacctt
     2101 gaagcgcatg aactccttga tgacgttctc ggaggaggcc atttcgaatc gatgggatcc
     2161 tgcaaaaaga acaagtagct tgtattctat agtgtcacct aaatcaagct tgctccgacg
     2221 tccccaggca gaatggcggt tccctaaacg agcattgctt atatagacct cccattaggc
     2281 acgcctaccg cccatttacg tcaatggaac gcccatttgc gtcattgccc ctccccattg
     2341 acgtcaatgg ggatgtactt ggcagccatc gcgggccatt taccgccatt gacgtcaatg
     2401 ggagtactgc caatgtaccc tggcgtactt ccaatagtaa tgtacttgcc aagttactat
     2461 taatagatat tgatgtactg ccaagtgggc catttaccgt cattgacgtc aatagggggc
     2521 gtgagaacgg atatgaatgg gcaatgagcc atcccattga cgtcaatggt gggtggtcct
     2581 attgacgtca atgggcattg agccaggcgg gccatttacc gtaattgacg tcaatggggg
     2641 aggcgccata tacgtcaata ggaccgccca tatgacgtca ataggaaaga ccatatatag
     2701 agaccattga cgtcaatggg ggagtggcta tgggcggtat taggaagccc catatatggt
     2761 atatgggacc gcccattggg aggggctatc tacgtcaata ggaaaaccca tatatggaat
     2821 actatatggc atagggccaa tacatagtat tgaacctggc caatagccat attggcatag
     2881 ggccatattg gatattgcct atatattgat cctggcatat agccaatatg gccgccatta
     2941 ttggcaccat gccaattcaa tatggcggac ttggcacagg gccaagaccc ctcctcaaac
     3001 ttggcaccgt gccaagtaga cccctcccca gttggcacag tgccaggtcc gccatattga
     3061 attggcatgg tgccaagtcc gccatattga attggctggg gtccagatcc accatattga
     3121 attggcacga gtccatatac gccatattga attggcacag gtccagatcc accatattga
     3181 attggcatga gtccatatac gccatattga attggcggtc gactggcgta atagcgaaga
     3241 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc
     3301 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     3361 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     3421 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     3481 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     3541 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     3601 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     3661 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     3721 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
     3781 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     3841 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     3901 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     3961 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     4021 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     4081 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     4141 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     4201 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     4261 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     4321 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     4381 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     4441 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     4501 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     4561 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     4621 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     4681 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     4741 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     4801 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     4861 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
//
