LOCUS       sequence-12669-e        7878 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7878)
  AUTHORS   Trichas G, Begbie J, Srinivas S
  TITLE     Use of the viral 2A peptide for bicistronic expression in transgenic
            mice.
  JOURNAL   BMC Biol. 2008 Sep 15;6:40. doi: 10.1186/1741-7007-6-40.
  PUBMED    18793381
REFERENCE   2  (bases 1 to 7878)
  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: "BMC Biol.";
            date: "2008-09-15"; volume: "6:40. doi"; pages: " 
            10.1186/1741-7007-6-40"
COMMENT     Sequence Label: pCAG-TAG
FEATURES             Location/Qualifiers
     source          1..7878
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      7..462
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(94..113)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     304..325
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        677..1056
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1059..1334
                     /label=chicken beta-actin promoter
     intron          1335..2343
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2267..2289
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2351..2370
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2418..2427
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2472..2481
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2541..3968
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     CDS             3978..4031
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4050..4427
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAVTKYTSAK"
     CDS             join(4512..4514,4515..5225)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(4554..4575)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4815..4834)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5162..5183
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5510..5644
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5547..5566)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(5693..5711)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5732..5748)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5732..5748)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5745..5767)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5756..5772
                     /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(5780..5786,5787..5804,5805..5810))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5825..5846
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5963..5980)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6134..6722)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6214..6233)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6893..7684,7685..7753))
                     /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     7516..7535
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7754..7858)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
       61 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
      121 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
      181 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
      241 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
      301 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
      361 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
      421 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtaccgg
      661 gccccccctc gaggtcgaca ttgattattg actagttatt aatagtaatc aattacgggg
      721 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      781 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgtccccata
      841 gtaacgccaa tagggacttt ccattgacgt caatgggtgg actatttacg gtaaactgcc
      901 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      961 ggtaaatggc ccgcctggca tttagcccag tacatgacct tatgggactt tcctacttgg
     1021 cagtacatct acgtattagt catcgctatt accatgggtc gaggtgagcc ccacgttctg
     1081 cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat ttatttttta
     1141 attattttgt gcagcgatgg gggcgggggg ggggggggcg cgcgccaggc ggggcggggc
     1201 ggggcgaggg gcggggcggg gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg
     1261 cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg gcggccatat aaaaagcgaa
     1321 gcgcgcggcg ggcgggagtc gctgcgttgc cttcgccccg tgccccgctc cgcgccgcct
     1381 cgcgccgccc gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg
     1441 gcccttctcc tccgggctgt aattagcgct tggtttaatg acggctcgtt tcttttctgt
     1501 ggctgcgtga aagccttaaa gggctccggg agggcccttt gtgcgggggg gagcggctcg
     1561 gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggcccgcg ctgcccggcg
     1621 gctgtgagcg ctgcgggcgc ggcgcggggc tttgtgcgct ccgcgtgtgc gcgaggggag
     1681 cgcggccggg ggcggtgccc cgcggtgcgg gggggctgcg aggggaacaa aggctgcgtg
     1741 cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg cggcggtcgg gctgtaaccc
     1801 ccccctgcac ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg
     1861 tcgcgggcgt ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc
     1921 gggcggggcg gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggccccgg
     1981 agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc
     2041 gagagggcgc agggacttcc tttgtcccaa atctggcgga gccgaaatct gggaggcgcc
     2101 gccgcacccc ctctagcggg cgcgggcgaa gcggtgcggc gccggcagga aggaaatggg
     2161 cggggagggc cttgctgcgt cgccgcgccg ccgtcccctt ctccatctcc agcctcgggg
     2221 ctgccgcagg gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg
     2281 cgtgtgaccg gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta
     2341 cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattagct
     2401 tgattcgagc ctctagagcc accatgggct gcatcaagag caagcgcaag gacaacctga
     2461 acgacgacga ggccgccatg ggctgcatca agagcaagcg caaggacaac ctgaacgacg
     2521 acgaggcacc ggtcgcctcc atggtgagca agggcgagga ggtcatcaaa gagttcatgc
     2581 gcttcaaggt gcgcatggag ggctccatga acggccacga gttcgagatc gagggcgagg
     2641 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggcggcc
     2701 ccctgccctt cgcctgggac atcctgtccc cccagttcat gtacggctcc aaggcgtacg
     2761 tgaagcaccc cgccgacatc cccgattaca agaagctgtc cttccccgag ggcttcaagt
     2821 gggagcgcgt gatgaacttc gaggacggcg gtctggtgac cgtgacccag gactcctccc
     2881 tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg caccaacttc ccccccgacg
     2941 gccccgtaat gcagaagaag accatgggct gggaggcctc caccgagcgc ctgtaccccc
     3001 gcgacggcgt gctgaagggc gagatccacc aggccctgaa gctgaaggac ggcggccact
     3061 acctggtgga gttcaagacc atctacatgg ccaagaagcc cgtgcaactg cccggctact
     3121 actacgtgga caccaagctg gacatcacct cccacaacga ggactacacc atcgtggaac
     3181 agtacgagcg ctccgagggc cgccaccacc tgttcctggg gcatggcacc ggcagcaccg
     3241 gcagcggcag ctccggcacc gcctcctccg aggacaacaa catggccgtc atcaaagagt
     3301 tcatgcgctt caaggtgcgc atggagggct ccatgaacgg ccacgagttc gagatcgagg
     3361 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     3421 gcggccccct gcccttcgcc tgggacatcc tgtcccccca gttcatgtac ggctccaagg
     3481 cgtacgtgaa gcaccccgcc gacatccccg attacaagaa gctgtccttc cccgagggct
     3541 tcaagtggga gcgcgtgatg aacttcgagg acggcggtct ggtgaccgtg acccaggact
     3601 cctccctgca ggacggcacg ctgatctaca aggtgaagat gcgcggcacc aacttccccc
     3661 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctccacc gagcgcctgt
     3721 acccccgcga cggcgtgctg aagggcgaga tccaccaggc cctgaagctg aaggacggcg
     3781 gccactacct ggtggagttc aagaccatct acatggccaa gaagcccgtg caactgcccg
     3841 gctactacta cgtggacacc aagctggaca tcacctccca caacgaggac tacaccatcg
     3901 tggaacagta cgagcgctcc gagggccgcc accacctgtt cctgtacggc atggacgagc
     3961 tgtacaaggg cagtggagag ggcagaggaa gtctgctaac atgcggtgac gtcgaggaga
     4021 atcctggccc aggagatctg aatatcacca tgccagagcc agcgaagtct gctcccgccc
     4081 cgaaaaaggg ctccaagaag gcggtgacta aggcgcagaa gaaaggcggc aagaagcgca
     4141 agcgcagccg caaggagagc tattccatct atgtgtacaa ggttctgaag caggtccacc
     4201 ctgacaccgg catttcgtcc aaggccatgg gcatcatgaa ttcgtttgtg aacgacattt
     4261 tcgagcgcat cgcaggtgag gcttcccgcc tggcgcatta caacaagcgc tcgaccatca
     4321 cctccaggga gatccagacg gccgtgcgcc tgctgctgcc tggggagttg gccaagcacg
     4381 ccgtgtccga gggtactaag gccgtcacca agtacaccag cgctaaggga tctgggggtg
     4441 gaggctccgg cgggggtgga tctggaggtg ggggcagcgg cggaggtggg ctagcgctac
     4501 cgcgggccac catgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     4561 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     4621 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     4681 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     4741 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     4801 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     4861 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     4921 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     4981 agaagaacgg catcaaggcc aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     5041 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     5101 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     5161 acatggtcct gctggagttc gtgaccgccg ccgggatcac tcacggcatg gacgagctgt
     5221 acaagtccgg actcagatca tcaagcttat cggatccggg cggccgtgat aataggtgat
     5281 agtaagtaat agtgagatat ctctagaatt ccagctgagc gccggtcgct accattacca
     5341 gttggtctgg tgtcaaaaat aataataacc gggcaggggg gatctaagct ctagataagt
     5401 aatgatcata atcagccata tcacatctgt agaggtttta cttgctttaa aaaacctccc
     5461 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     5521 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5581 tttttcactg cattctagtt gtggtttgtc caanctcatc aatgtatctt atcatgtctg
     5641 gatccactag ttcagagcgg ccgcccccgc ggtggagctc cagcttttgt tccctttagt
     5701 gagggttaat ttcgagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     5761 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     5821 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     5881 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     5941 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     6001 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     6061 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     6121 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     6181 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     6241 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6301 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6361 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6421 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6481 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6541 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     6601 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6661 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6721 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6781 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     6841 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     6901 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     6961 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     7021 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     7081 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     7141 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     7201 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     7261 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     7321 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     7381 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7441 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     7501 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7561 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7621 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     7681 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7741 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     7801 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     7861 catttccccg aaaagtgc
//
