LOCUS       sequence-371189-        6246 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses blue fluorescent protein Electra2 and EGFP in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6246)
  AUTHORS   Papadaki S, Wang X, Wang Y, Zhang H, Jia S, Liu S, Yang M, Zhang D, 
            Jia JM, Koster RW, Namikawa K, Piatkevich KD
  TITLE     Dual-expression system for blue fluorescent protein optimization.
  JOURNAL   Sci Rep. 2022 Jun 17;12(1):10190. doi: 10.1038/s41598-022-13214-0.
  PUBMED    35715437
REFERENCE   2  (bases 1 to 6246)
  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: "Sci Rep."; 
            date: "2022-06-17"; volume: "12(1):10190. doi"; pages: " 
            10.1038/s41598-022-13214-0"
COMMENT     Sequence Label: pAAV-CAG-Electra2-P2A-EGFP
FEATURES             Location/Qualifiers
     source          1..6246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     837..859
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     921..940
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      979..988
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1720..1776
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(1789..1791,1792..2505)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1789..1791,1792..2502)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1831..1852)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2092..2111)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2439..2460
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2530..3118
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2583..2603)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3001..3012)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3161..3282
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3172..3191)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3226..3245
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3444..3573
                     /label=AAV2 ITR
     rep_origin      complement(3810..4398)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3890..3909)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4569..5360,5361..5429))
                     /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     5192..5211
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5430..5534)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5560..6015)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5697..5718)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5909..5928
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   6087..6216
                     /label=AAV2 ITR
ORIGIN
        1 tatctacgta gccatgctct aggaagagta ccattgacgt caataatgac gtatgttccc
       61 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
      121 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
      181 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
      241 tggcagtaca tctacgtatt agtcatcgct attaccatgg tcgaggtgag ccccacgttc
      301 tgcttcactc tccccatctc ccccccctcc ccacccccaa ttttgtattt atttattttt
      361 taattatttt gtgcagcgat gggggcgggg gggggggggg ggcgcgcgcc rggsggggsg
      421 gggsggggsg rggggsgggg sggggsgrgg cggrgaggkg cggcggcagc caatcagagc
      481 ggcgcgctcc gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc ctataaaaag
      541 cgaagcgcgc ggcgggcggg agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc
      601 gccgcctcgc gccgcccgcc ccggctctga ctgaccgcgt tactcccaca ggtgagcggg
      661 cgggacggcc cttctcctcc gggctgtaat tagcgcttgg tttaatgacg gcttgtttct
      721 tttctgtggc tgcgtgaaag ccttgagggg ctccgggagg gccctttgtg cggggggagc
      781 ggctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca gggcggggtt
      841 cggcttctgg cgtgtgaccg gcggctctag agcctctgct aaccatgttc atgccttctt
      901 ctttttccta cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc attttggcaa
      961 agaattggat ccggtaccgc caccatggtg tctaagggcg aagagctgat cgaggaaaat
     1021 atgcgtatga aggtggtcat ggaaggttcg gtcaacggcc accaattcaa atgcacaggt
     1081 gaaggagaag gcagaccgta cgagggagtg caaaccatga ggatcaaagt catcgaggga
     1141 ggacccctgc catttgcctt tgacattctt gccacgtcgt tcctgttcgg cagcaagacc
     1201 tttatcaagt acccggccga catccctgat ttcttcgaac agtcctttcc tgagggcttt
     1261 acttgggaaa gagttacgag atacgaagat ggtggagtag tcaccgtcac gcaggacacc
     1321 agccttgagg atggtgggct cgtctacaac gtcaaggtca gaggggtaaa ctttcactcc
     1381 aagggtcccg tgatgcagaa gaagaccgag ggttgggagc ctttcacaga gatgatgtat
     1441 ccagcagatg gtggtctgag aggatatacc gacatcgcac tgaaagttga tggtggtggc
     1501 catctgcacg ccaacatcgt gacaacttac aggtcaaaaa agaccgtcgg gaacatcaag
     1561 atgcccggtg tccatgccgt tgattaccgc ctggaaagga tcgaagagag tgacaatgaa
     1621 acctacgtag tgctaagaga agtggcagtt gccaaataca gcaaccttgg tggtggcatg
     1681 gacgagctgt tcaagcgacc ggttcacgtg ggaagtgggg ctacaaactt ctcactgctg
     1741 aaacaggccg gggatgtgga ggaaaaccca ggaccacacg tgactagtgt gagcaagggc
     1801 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     1861 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     1921 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     1981 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     2041 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     2101 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     2161 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     2221 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     2281 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     2341 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
     2401 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     2461 accgccgccg ggatcactct cggcatggac gagctgtaca agtaagaatt cgatatcaag
     2521 cttatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     2581 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     2641 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     2701 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     2761 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     2821 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     2881 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct
     2941 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     3001 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     3061 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat
     3121 cgataccgtc gacccgggcg gccgcttcga gcagacatga taagatacat tgatgagttt
     3181 ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct
     3241 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     3301 cattttatgt ttcaggttca gggggagatg tgggaggttt tttaaagcaa gtaaaacctc
     3361 tacaaatgtg gtaaaatcga taaggatctt cctagagcat ggctacgtag ataagtagca
     3421 tggcgggtta atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct
     3481 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     3541 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcatt aatgaatcgg ccaacgcgcg
     3601 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3661 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3721 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     3781 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3841 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     3901 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     3961 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4021 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4081 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4141 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4201 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4261 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4321 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4381 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4441 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4501 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4561 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4621 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4681 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     4741 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4801 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4861 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     4921 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     4981 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5041 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5101 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5161 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5221 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5281 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5341 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5401 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5461 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     5521 ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt
     5581 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     5641 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     5701 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     5761 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     5821 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     5881 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     5941 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     6001 cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
     6061 tgcgggcctc ttcgctatta cgccagctgc gcgctcgctc gctcactgag gccgcccggg
     6121 caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag cgagcgcgca
     6181 gagagggagt ggccaactcc atcactaggg gttccttgta gttaatgatt aacccgccat
     6241 gctact
//
