LOCUS       sequence-478886-        6106 bp    DNA     circular SYN 17-JUL-2025
DEFINITION  Expresses (n1)QC2-6 FIQ(c4) and mCherry in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6106)
  AUTHORS   Zhang H, Lesnov GD, Subach OM, Zhang W, Kuzmicheva TP, Vlaskina AV, 
            Samygina VR, Chen L, Ye X, Nikolaeva AY, Gabdulkhakov A, Papadaki S,
            Qin W, Borshchevskiy V, Perfilov MM, Gavrikov AS, Drobizhev M, 
            Mishin AS, Piatkevich KD, Subach FV
  TITLE     Bright and stable monomeric green fluorescent protein derived from 
            StayGold.
  JOURNAL   Nat Methods. 2024 Apr;21(4):657-665. doi: 
            10.1038/s41592-024-02203-y. Epub 2024 Feb 26.
  PUBMED    38409224
REFERENCE   2  (bases 1 to 6106)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 17, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41592-024-02203-y"; journalName: "Nat Methods"; date: 
            "2024-04"; volume: "21"; issue: "4"; pages: "657-665"
COMMENT     Sequence Label: pAAV-CAG-(n1)QC2-6 FIQ(c4)-P2A-mCherry
FEATURES             Location/Qualifiers
     source          1..6106
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     847..869
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     931..950
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      993..1002
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             999..1676
                     /codon_start=1
                     /product="bright and highly photostable monomeric green 
                     fluorescent protein (Ando et al., 2023)"
                     /label=mStayGold(J)
                     /note="includes multiple mutations plus an N-terminal 
                     iinker sequence"
                     /translation="MASTGEELFTGVVPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKG
                     KYVCTSGKLPMSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGS
                     IHAKHQHFMKNGTYHNIVEFTGQDFKENSPVLTGDMDVSLPNEVQHIPIDDGVECTVTL
                     QYPLLSDESKCVEAYQNTIIKPLHNQPAPDVPFHWIRKQYTQSKDDTEERDHIIQSETL
                     EAHL"
     CDS             1731..1787
                     /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             1800..2510
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1945..1963)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2213..2232
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2538..3126
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2591..2611)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3009..3020)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3169..3290
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3180..3199)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3234..3253
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3669..4257)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3749..3768)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4428..5219,5220..5288))
                     /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     5051..5070
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5289..5393)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5419..5874)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5556..5577)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5768..5787
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   5946..6075
                     /label=AAV2 ITR
ORIGIN
        1 atctacgtag ccatgctcta ggaagagtac cattgacgtc aataatgacg tatgttccca
       61 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      121 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      181 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      241 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct
      301 gcttcactct ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt
      361 aattattttg tgcagcgatg ggggcggggg gggggggggg gggggggggg gggggggggg
      421 gggggggggg gggggggggg gggggggggg cggggcgagg cggagaggtg cggcggcagc
      481 caatcagagc ggcgcgctcc gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc
      541 ctataaaaag cgaagcgcgc ggcgggcggg agtcgctgcg cgctgccttc gccccgtgcc
      601 ccgctccgcc gccgcctcgc gccgcccgcc ccggctctga ctgaccgcgt tactcccaca
      661 ggtgagcggg cgggacggcc cttctcctcc gggctgtaat tagcgcttgg tttaatgacg
      721 gcttgtttct tttctgtggc tgcgtgaaag ccttgagggg ctccgggagg gccctttgtg
      781 cggggggagc ggctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca
      841 gggcggggtt cggcttctgg cgtgtgaccg gcggctctag agcctctgct aaccatgttc
      901 atgccttctt ctttttccta cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc
      961 attttggcaa agaattggat ccggtaccct aagccaccat ggccagcacc ggggaggagc
     1021 tgttcaccgg ggtggtgccc ttcaagttcc agctgaaggg caccatcaac ggcaagagct
     1081 tcaccgtgga gggcgagggc gagggcaaca gccacgaggg cagccacaag ggcaagtacg
     1141 tgtgcaccag cggcaagctg cccatgagct gggccgccct gggcaccagc ttcggctacg
     1201 gcatgaagta ctacaccaag taccccagcg gcctgaagaa ctggttccac gaggtgatgc
     1261 ccgagggctt cacctacgac aggcacatcc agtacaaggg cgacggcagc atccacgcca
     1321 agcaccagca cttcatgaag aacggcacct accacaacat cgtggagttc accggccagg
     1381 acttcaagga gaacagcccc gtgctgaccg gcgacatgga cgtgagcctg cccaacgagg
     1441 tgcagcacat ccccatcgac gacggcgtgg agtgcaccgt gaccctgcag taccccctgc
     1501 tgagcgacga gagcaagtgc gtggaggcct accagaacac catcatcaag cccctgcaca
     1561 accagcccgc ccccgacgtg cccttccact ggatcaggaa gcagtacacc cagagcaagg
     1621 acgacaccga ggagagggac cacatcatcc agagcgagac cctggaggcc cacctgccat
     1681 ggcacgagcc ttctgcttct gccgtgcgac cggttcacgt gggaagtggg gctacaaact
     1741 tctcactgct gaaacaggcc ggggatgtgg aggaaaaccc aggaccacac gtgactagta
     1801 tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg
     1861 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc
     1921 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct
     1981 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc
     2041 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg
     2101 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg
     2161 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa
     2221 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg
     2281 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg
     2341 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca
     2401 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac
     2461 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtag taagaattcg
     2521 atatcaagct tatcgataat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     2581 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     2641 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     2701 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     2761 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     2821 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     2881 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt
     2941 cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     3001 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     3061 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct
     3121 ccccgcatcg ataccgtcga cccgggcggc cgcttcgagc agacatgata agatacattg
     3181 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     3241 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     3301 attgcattca ttttatgttt caggttcagg gggagatgtg ggaggttttt taaagcaagt
     3361 aaaacctcta caaatgtggt aaaatcgata aggatcttcc tagagcatgg ctacgtagat
     3421 aagtagcatg gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact
     3481 ccctctctgc gcgctcgctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     3541 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     3601 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     3661 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     3721 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     3781 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     3841 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     3901 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     3961 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     4021 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     4081 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     4141 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     4201 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     4261 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     4321 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     4381 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     4441 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     4501 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     4561 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     4621 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     4681 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     4741 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     4801 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     4861 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     4921 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     4981 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     5041 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     5101 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     5161 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     5221 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     5281 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     5341 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     5401 ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     5461 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     5521 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     5581 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     5641 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     5701 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     5761 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     5821 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt
     5881 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     5941 gccagctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga
     6001 cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca
     6061 tcactagggg ttccttgtag ttaatgatta acccgccatg ctactt
//
