LOCUS       sequence-338181-        7503 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template pDNA to prepare a split CaVT component mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7503)
  AUTHORS   Nakanishi H, Yoshii T, Kawasaki S, Hayashi K, Tsutsui K, Oki C, 
            Tsukiji S, Saito H
  TITLE     Light-controllable RNA-protein devices for translational regulation 
            of synthetic mRNAs in mammalian cells.
  JOURNAL   Cell Chem Biol. 2021 Jan 20. pii: S2451-9456(21)00002-7. doi: 
            10.1016/j.chembiol.2021.01.002.
  PUBMED    33508227
REFERENCE   2  (bases 1 to 7503)
  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. 2021 Jan 20. pii: S2451-9456(21)00002-7. doi: 
            10.1016/j.chembiol.2021.01.002."
COMMENT     Sequence Label: pBCMV-MS2CP-eDHFR
FEATURES             Location/Qualifiers
     source          1..7503
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     57..79
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     71..88
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     72..88
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        95..113
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   180..242
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        486..865
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        866..1069
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1020..1040
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        1114..1132
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1186..1195
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1549..2022
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="ISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRHT
                     WESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFLP
                     KAQKLYLTHIDAEVEGDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     CDS             2041..2064
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2084..2101)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2090..2314
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2360..2788
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2797..2817)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2802..3131
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2982..3117
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3044..3063
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3198..3992
                     /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(3252..3271)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3862..3881
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4166..4287
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4203..4222)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4257..4276
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   4604..4638
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(4787..4805)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4826..4842)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4826..4842)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4839..4861)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4850..4866
                     /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(4874..4880,4881..4898,4899..4904))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4919..4940
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5057..5074)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5228..5816)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5308..5327)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5987..6778,6779..6847))
                     /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     6610..6629
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6848..6952)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6978..7433)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca
       61 gtcacgacgt tgtaaaacga cggccagtga attgtaatac gactcactat agggcgaatt
      121 ggagctcggt attcacgaca gcaggctgaa taataaaaaa attagaaact attatttaac
      181 cctagaaaga taatcatatt gtgacgtacg ttaaagataa tcatgcgtaa aattgacgca
      241 tgtgttttat cggtctgtat atcgaggttt atttattaat ttgaatagat attaagtttt
      301 attatattta cacttacata ctaataataa attcaacaaa caatttattt atgtttattt
      361 atttattaaa aaaaaacaaa aactcaaaat ttcttctata aagtaacaaa acttttaaac
      421 attctctctt ttacaaaaat aaacttattt tgtactttaa aaacagtcat gttgtattac
      481 gcgttgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
      541 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
      601 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
      661 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
      721 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
      781 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
      841 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
      901 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      961 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
     1021 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac
     1081 tagagaaccc actgcttact ggcttatcga aattaatacg actcactata gggagaccca
     1141 agctggctag cgtttaaact taagcttggt accgagctcg gatctgccac catggcttct
     1201 aactttactc agttcgttct cgtcgacaat ggcggaactg gcgacgtgac tgtcgcccca
     1261 agcaacttcg ctaacggggt cgctgaatgg atcagctcta actcgcgatc acaggcttac
     1321 aaagtaacct gtagcgttcg tcagagctct gcgcagaatc gcaaatacac catcaaagtc
     1381 gaggtgccta aaggcgcatg gaggtcttac ttaaatatgg aactaaccat tccaattttc
     1441 gccacgaatt ccgactgcga gcttattgtt aaggcaatgc aaggtctcct aaaagatgga
     1501 aacccgattc cctcggccat cgcggccaac tccggcatct acggatccat cagtctgatt
     1561 gcggcgttag cggtagatcg cgttatcggc atggaaaacg ccatgccgtg gaacctgcct
     1621 gccgatctcg cctggtttaa acgcaacacc ttaaataaac ccgtgattat gggccgccat
     1681 acctgggaat caatcggtcg tccgttgcca ggacgcaaaa atattatcct cagcagtcaa
     1741 ccgggtacgg acgatcgcgt aacgtgggtg aagtcggtgg atgaagccat cgcggcgtgt
     1801 ggtgacgtac cagaaatcat ggtgattggc ggcggtcgcg tttatgaaca gttcttgcca
     1861 aaagcgcaaa aactgtatct gacgcatatc gacgcagaag tggaaggcga cacccatttc
     1921 ccggattacg agccggatga ctgggaatcg gtattcagcg aattccacga tgctgatgcg
     1981 cagaactctc acagctattg ctttgagatt ctggagcggc ggaccggtgt tccaactgtt
     2041 gattacaagg atgacgacga taagtgataa acccgctgat cagcctcgac tgtgccttct
     2101 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     2161 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     2221 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     2281 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     2341 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     2401 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     2461 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     2521 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     2581 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     2641 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     2701 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     2761 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     2821 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2881 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     2941 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3001 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3061 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3121 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag
     3181 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     3241 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     3301 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     3361 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     3421 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     3481 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     3541 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     3601 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     3661 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     3721 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     3781 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     3841 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     3901 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     3961 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     4021 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     4081 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4141 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     4201 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4261 tttgtccaaa ctcatcaatg tatcttatca tgtctgtaag aactacccat tttattatat
     4321 attagtcacg atatctataa caagaaaata tatatataat aagttatcac gtaagtagaa
     4381 catgaaataa caatataatt atcgtatgag ttaaatctta aaagtcacgt aaaagataat
     4441 catgcgtcat tttgactcac gcggtcgtta tagttcaaaa tcagtgacac ttaccgcatt
     4501 gacaagcacg cctcacggga gctccaagcg gcgactgaga tgtcctaaat gcacagcgac
     4561 ggattcgcgc tatttagaaa gagagagcaa tatttcaaga atgcatgcgt caattttacg
     4621 cagactatct ttctagggtt aaaaaagatt tgcgctttac tcgacctaaa ctttaaacac
     4681 gtcatagaat cttcgtttga caaaaaccac attgtggcca agctgtgtga cgcgacgcgc
     4741 gctaaagaat ggcaaaccaa gtcgcgcgag gtaccagctt ttgttccctt tagtgagggt
     4801 taatttcgag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     4861 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     4921 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     4981 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5041 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5101 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5161 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5221 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5281 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5341 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5401 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5461 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     5521 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     5581 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     5641 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     5701 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5761 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5821 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5881 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5941 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     6001 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     6061 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     6121 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     6181 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     6241 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     6301 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     6361 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     6421 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     6481 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     6541 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     6601 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6661 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     6721 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     6781 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     6841 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6901 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6961 cccgaaaagt gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     7021 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     7081 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     7141 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     7201 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     7261 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     7321 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     7381 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc
     7441 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     7501 cca
//
