LOCUS       sequence-366278-        5845 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express miniGFP1 and FusionRed in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5845)
  AUTHORS   Liang GT, Lai C, Yue Z, Zhang H, Li D, Chen Z, Lu X, Tao L, Subach 
            FV, Piatkevich KD
  TITLE     Enhanced small green fluorescent proteins as a multisensing platform
            for biosensor development.
  JOURNAL   Front Bioeng Biotechnol. 2022 Oct 17;10:1039317. doi: 
            10.3389/fbioe.2022.1039317. eCollection 2022.
  PUBMED    36324888
REFERENCE   2  (bases 1 to 5845)
  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: "Front 
            Bioeng Biotechnol."; date: "2022-10-17"; volume: "10:1039317. doi"; 
            pages: " 10.3389/fbioe.2022.1039317. eCollection 2022"
COMMENT     Sequence Label: pAAV-CAG-miniGFP1-P2A-FusionRed
FEATURES             Location/Qualifiers
     source          1..5845
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        279..556
                     /label=chicken beta-actin promoter
     primer_bind     835..857
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     919..938
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      977..986
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1337..1393
                     /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             1406..2104
                     /codon_start=1
                     /product="monomeric red fluorescent protein optimized for 
                     use as a fusion tag (Shemiakina et al., 2012)"
                     /label=FusionRed
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMPMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFMYGSRTFIKHPPGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKVRGVNFPANGPVMQKKTLGWEASTETMYPADGGLEGACDMAL
                     KLVGGGHLICNLETTYRSKKPATNLKMPGVYNVDHRLERIKEADDETYVEQHEVAVARY
                     STGGAGDGGK"
     misc_feature    2129..2717
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2182..2202)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2600..2611)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2760..2881
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2771..2790)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2825..2844
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3043..3172
                     /label=AAV2 ITR
     rep_origin      complement(3409..3997)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3489..3508)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4168..4959,4960..5028))
                     /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     4791..4810
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5029..5133)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5159..5614)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5296..5317)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5508..5527
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   5686..5815
                     /label=AAV2 ITR
ORIGIN
        1 tctacgtagc catgctctag gaagagtacc attgacgtca ataatgacgt atgttcccat
       61 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      121 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
      181 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
      241 gcagtacatc tacgtattag tcatcgctat taccatggtc gaggtgagcc ccacgttctg
      301 cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat ttatttttta
      361 attattttgt gcagcgatgg gggcgggggg gggggggggg cgcgcgccag gcggggcggg
      421 gcggggcgag gggcggggcg gggcgaggcg gagaggtgcg gcggcagcca atcagagcgg
      481 cgcgctccga aagtttcctt ttatggcgag gcggcggcgg cggcggccct ataaaaagcg
      541 aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc
      601 cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg tgagcgggcg
      661 ggacggccct tctcctccgg gctgtaatta gcgcttggtt taatgacggc ttgtttcttt
      721 tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg gggggagcgg
      781 ctcggggctg tccgcggggg gacggctgcc ttcggggggg acggggcagg gcggggttcg
      841 gcttctggcg tgtgaccggc ggctctagag cctctgctaa ccatgttcat gccttcttct
      901 ttttcctaca gctcctgggc aacgtgctgg ttattgtgct gtctcatcat tttggcaaag
      961 aattggatcc ggtaccgcca ccatggagaa gtcttttgtg attacagacc catggctgcc
     1021 cgactaccct attatttccg ctagtgacgg attcctggag ctgaccgagt attctcggga
     1081 ggaaatcatg ggaaggaacg cacgcttcct gcagggaccc gagactgacc aggccaccgt
     1141 gcagaagatc cgggacgcta ttcgagatcg gagacctacc acagtccagc tgatcaacta
     1201 cacaaagagc ggcaagaagt tctggaatct gttgcacctg cagcctgtgt ttgacggaaa
     1261 aggcgggctg caatatttta ttggagtgca gctggtcggc tccgatcatg tccgaccggt
     1321 tcacgtggga agtggggcta caaacttctc actgctgaaa caggccgggg atgtggagga
     1381 aaacccagga ccacacgtga ctagtatggt gagcgagctg attaaggaga acatgcccat
     1441 gaagctgtac atggagggca ccgtgaacaa ccaccacttc aagtgcacat ccgagggcga
     1501 aggcaagccc tacgagggca cccagaccat gagaatcaag gtcgtcgagg gcggccctct
     1561 ccccttcgcc ttcgacatcc tggctaccag cttcatgtac ggcagcagaa ccttcatcaa
     1621 gcaccctccg ggcatccccg acttctttaa gcagtccttc cctgagggct tcacatggga
     1681 gagagtcacc acatacgaag acgggggcgt gctgaccgct acccaggaca ccagcctcca
     1741 ggacggctgc ctcatctaca acgtcaaggt tagaggggtg aacttcccag ccaacggccc
     1801 tgtgatgcag aagaaaacac tcggctggga ggcctccacc gagacgatgt accctgctga
     1861 cggaggactg gaaggcgcat gtgacatggc tctgaagctc gtgggaggag gacacctgat
     1921 ctgcaacctt gagacaacat acagatccaa gaaacccgct acgaacctca agatgcccgg
     1981 cgtctacaac gtggaccaca gactggaaag aatcaaggag gccgacgatg agacatacgt
     2041 cgagcagcac gaggtggctg tggccagata ctctactggt ggcgctggtg atggaggtaa
     2101 ataagaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     2161 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     2221 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     2281 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     2341 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     2401 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     2461 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     2521 gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     2581 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     2641 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     2701 ttgggccgcc tccccgcatc gataccgtcg acccgggcgg ccgcttcgag cagacatgat
     2761 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     2821 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     2881 taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt
     2941 ttaaagcaag taaaacctct acaaatgtgg taaaatcgat aaggatcttc ctagagcatg
     3001 gctacgtaga taagtagcat ggcgggttaa tcattaacta caaggaaccc ctagtgatgg
     3061 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     3121 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcatta
     3181 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     3241 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3301 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3361 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3421 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3481 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3541 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3601 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3661 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3721 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3781 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3841 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3901 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3961 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     4021 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     4081 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     4141 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     4201 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     4261 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     4321 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     4381 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     4441 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     4501 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4561 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4621 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4681 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     4741 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     4801 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     4861 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     4921 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     4981 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     5041 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     5101 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctaa
     5161 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     5221 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     5281 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     5341 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     5401 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     5461 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     5521 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     5581 cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt cgccattcag gctgcgcaac
     5641 tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctgcg cgctcgctcg
     5701 ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca
     5761 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttccttgtag
     5821 ttaatgatta acccgccatg ctact
//
