LOCUS       sequence-375314-        6355 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual expression vector for bacteria and mammalian cells producing 
            mScarlet-SYFP2 fusion protein for FRET analysis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6355)
  AUTHORS   Gadella TWJ Jr, van Weeren L, Stouthamer J, Hink MA, Wolters AHG, 
            Giepmans BNG, Aumonier S, Dupuy J, Royant A
  TITLE     mScarlet3: a brilliant and fast-maturing red fluorescent protein.
  JOURNAL   Nat Methods. 2023 Apr;20(4):541-545. doi: 
            10.1038/s41592-023-01809-y. Epub 2023 Mar 27.
  PUBMED    36973546
REFERENCE   2  (bases 1 to 6355)
  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"; doi: 
            "10.1038/s41592-023-01809-y"; journalName: "Nat Methods"; date: 
            "2023-04"; volume: "20"; issue: "4"; pages: "541-545"
COMMENT     Sequence Label: pDx_mScarlet-SYFP2
FEATURES             Location/Qualifiers
     source          1..6355
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1..816)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     724..743
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_recomb     1059..1886
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             1059..1183
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(1221..1826)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1954..2257
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2259..2461
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2412..2432
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2664..2686
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2748..2767
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     promoter        2809..2927
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             3002..3019
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3029..3046
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     regulatory      3065..3074
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3071..3766
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(3204..3222)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3472..3491
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             join(3767..3769,3770..3772,3773..4483)
                     /codon_start=1
                     /product="monomeric optimized variant of EYFP, identical to
                     mVenus but with L68V (Kremers et al., 2006)"
                     /label=SYFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3812..3833)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4073..4092)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4420..4441
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4573..4592)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4638..4693
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4692..4711)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      4781..4828
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     4841..5546
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           4929..5301
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(4958..5546)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(5555..5588)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(5756..6343)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5835..5854)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttagaaaaac tcatcgagca tcaagtgaaa ctgcaattta ttcatatcag gattatcaat
       61 accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca
      121 taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc
      181 tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac
      241 tgaatccggt gagaatggca aaagcttatg catttctttc cagacttgtt caacaggcca
      301 gccattacgc tcgtcatcaa aatcactcgc accaaccaaa ccgttattca ttcgtgattg
      361 cgcctgagcg agacgaaata cgcgatcgcc gttaaaagga caattacaaa caggaatcga
      421 atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata
      481 ttcttctaat acctggaatg ctgttttccc tgggatcgca gtggtgagta accatgcatc
      541 atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt
      601 tagcctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa
      661 caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac
      721 attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg
      781 cctcgagcaa gacgtttccc gttgaatatg gctcatactc ttcctttttc aatattattg
      841 aagcatttat cagggttatt gtctcatgtc cgcgcgtttc ctgcatcttt taatcaaatc
      901 ccaagatgtg tataaacgcg ccggtatgta taggaagagg tttatactaa actgttacat
      961 tgcaaacgtg gtttcgtgtg ccaagtgtga aaaccgatgt ttaatcaagg ctctgacgca
     1021 tttctacaac cacgactcca agtgtgtggg tgaagtcatg catcttttaa tcaaatccca
     1081 agatgtgtat aaaccaccaa actgccaaaa aatgaaaact gtcgacaagc tctgtccgtt
     1141 tgctggcaac tgcaagggtc tcaatcctat ttgtaattat tgaataataa aacaattata
     1201 aatgtcaaat ttgtttttta ttaacgatac aaaccaaacg caacaagaac atttgtagta
     1261 ttatctataa ttgaaaacgc gtagttataa tcgctgaggt aatatttaaa atcattttca
     1321 aatgattcac agttaatttg cgacaatata attttatttt cacataaact agacgccttg
     1381 tcgtcttctt cttcgtattc cttctctttt tcatttttct cttcataaaa attaacatag
     1441 ttattatcgt atccatatat gtatctatcg tatagagtaa attttttgtt gtcataaata
     1501 tatatgtctt ttttaatggg gtgtatagta ccgctgcgca tagtttttct gtaatttaca
     1561 acagtgctat tttctggtag ttcttcggag tgtgttgctt taattattaa atttatataa
     1621 tcaatgaatt tgggatcgtc ggttttatac aatatgttgc cggcatagta cgcagcttct
     1681 tctagttcaa ttacaccatt ttttagcagc accggattaa cataactttc caaaatgttg
     1741 tacgaaccgt taaacaaaaa cagttcacct cccttttcta tactattgtc tgcgagcagt
     1801 tgtttgttgt taaaaataac agccattgta atgagacgca caaactaata tcacaaactg
     1861 gaaatgtcta tcaatatata gttgctgatg gccggcctat taatagtaat caattacggg
     1921 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     1981 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     2041 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     2101 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
     2161 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
     2221 gcagtacatc tacgtattag tcatcgctat taccatgctg atgcggtttt ggcagtacac
     2281 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     2341 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
     2401 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagacg
     2461 tcgtttagtg aaccgtcaga tcactagatg ctttattgcg gtagtttatc acagttaaat
     2521 tgctaacgcc agtctcgaac ttaacgtgca gaagttggtc gtgaggcact gggcaggtaa
     2581 gtatcgggcc ctttgtgcgg ggggagcggc tcggggctgt ccgcgggggg acggctgcct
     2641 tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg gctctagagc
     2701 ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca acgtgctggt
     2761 tattgtgctg tctcatcatt ttggcaaaga attggatcgg accgaaatca ccacaattca
     2821 gcaaattgtg aacatcatca cgttcatctt tccctggttg ccaaaggccc attttcctgt
     2881 cagtaacgag aaggtcgcga attcaggcgc tttttagact ggtcgtaggg agaccacaac
     2941 ggtttccctc tagaaataat tttgtttaac tataagaagg agatatacca tgcggggttc
     3001 tcatcatcac caccatcaca gcagcggcct ggtgccgcgc ggcagctcgc tagcgctacc
     3061 ggtcgccacc atggtgagca agggcgaggc agtgatcaag gagttcatgc ggttcaaggt
     3121 gcacatggag ggctccatga acggccacga gttcgagatc gagggcgagg gcgagggccg
     3181 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
     3241 ctcctgggac atcctgtccc ctcagttcat gtacggctcc agggccttca ccaagcaccc
     3301 cgccgacatc cccgactact ataagcagtc cttccccgag ggcttcaagt gggagcgcgt
     3361 gatgaacttc gaggacggcg gcgccgtgac cgtgacccag gacacctccc tggaggacgg
     3421 caccctgatc tacaaggtga agctccgcgg caccaacttc cctcctgacg gccccgtaat
     3481 gcagaagaag acaatgggct gggaagcgtc caccgagcgg ttgtaccccg aggacggcgt
     3541 gctgaagggc gacattaaga tggccctgcg cctgaaggac ggcggccgct acctggcgga
     3601 cttcaagacc acctacaagg ccaagaagcc cgtgcagatg cccggcgcct acaacgtcga
     3661 ccgcaagttg gacatcacct cccacaacga ggactacacc gtggtggaac agtacgaacg
     3721 ctccgagggc cgccactcca ccggcggcat ggacgagctg tacaagatgg tgagcaaggg
     3781 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     3841 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     3901 gaagctgatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     3961 gggctacggc gtgcagtgct tcgcccgcta ccccgaccac atgaagcagc acgacttctt
     4021 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     4081 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     4141 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     4201 ctacaacagc cacaacgtct atatcaccgc cgacaagcag aagaacggca tcaaggccaa
     4261 cttcaagatc cgccacaaca tcgaggacgg cggcgtgcag ctcgccgacc actaccagca
     4321 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagctacca
     4381 gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     4441 gaccgccgcc gggatcactc tcggcatgga cgagctctac aagtaagtga ttaacctcag
     4501 gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac
     4561 cactgagatc gatctttttc cctctgccaa aaattatggg gacatcatga agccccttga
     4621 gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt
     4681 tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac atcagaatga
     4741 gtatttggtt tagagtttgg caacatatgc ccatatgtaa ctagcataac cccttggggc
     4801 ctctaaacgg gtcttgaggg gttttttgct gaaagcatgc ggaggaaatt ctccttgaag
     4861 tttccctggt gttcaaagta aaggagtttg caccagacgc acctctgttc actggtccgg
     4921 cgtattaaaa cacgatacat tgttattagt acatttatta agcgctagat tctgtgcgtt
     4981 gttgatttac agacaattgt tgtacgtatt ttaataattc attaaattta taatctttag
     5041 ggtggtatgt tagagcgaaa atcaaatgat tttcagcgtc tttatatctg aatttaaata
     5101 ttaaatcctc aatagatttg taaaataggt ttcgattagt ttcaaacaag ggttgttttt
     5161 ccgaaccgat ggctggacta tctaatggat tttcgctcaa cgccacaaaa cttgccaaat
     5221 cttgtagcag caatctagct ttgtcgatat tcgtttgtgt tttgttttgt aataaaggtt
     5281 cgacgtcgtt caaaatatta tgcgcttttg tatttctttc atcactgtcg ttagtataca
     5341 attgactcga cgtaaacacg ttaaatagag cttggacata tttaacatcg ggcgtgttag
     5401 ctttattagg ccgattatcg tcgtcgtccc aaccctcgtc gttagaagtt gcttccgaag
     5461 acgattttgc catagccaca cgacgcctat taattgtgtc ggctaacacg tccgcgatca
     5521 aatttgtagt tgagcttttt ggaattgcga tcgcataact tcgtatagca tacattatac
     5581 gaagttataa gctcggaacg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5641 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5701 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5761 ataggctccg ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     5821 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5881 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5941 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     6001 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     6061 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     6121 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     6181 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     6241 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     6301 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttg
//
