LOCUS       sequence-441199-        6230 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Flp-dependent expression of a single-cell discriminating version of 
            oScarlet fluorescent protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6230)
  AUTHORS   Szelenyi ER, Navarrete JS, Murry AD, Zhang Y, Girven KS, Kuo L, 
            Cline MM, Bernstein MX, Burdyniuk M, Bowler B, Goodwin NL, Juarez B,
            Zweifel LS, Golden SA
  TITLE     An arginine-rich nuclear localization signal (ArgiNLS) strategy for 
            streamlined image segmentation of single cells.
  JOURNAL   Proc Natl Acad Sci U S A. 2024 Aug 6;121(32):e2320250121. doi: 
            10.1073/pnas.2320250121. Epub 2024 Jul 29.
  PUBMED    39074275
REFERENCE   2  (bases 1 to 6230)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2024-08-6"; volume: "121(32):e2320250121. 
            doi"; pages: " 10.1073/pnas.2320250121. Epub 2024 Jul 29"
COMMENT     Sequence Label: pAAV-CAG-fDIO-ArgiNLS-oScarlet
FEATURES             Location/Qualifiers
     source          1..6230
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        210..513
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     1136..1158
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1220..1239
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1306..1339
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             1513..2205
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="VSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWDRVMNFEDGGA
                     VTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKM
                     ALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS
                     TGGMDELYK"
     primer_bind     complement(1643..1661)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1911..1930
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2215..2248)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    2357..2945
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2410..2430)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2828..2839)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2977..3453
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3176..3195)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3493..3633
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3493..3622
                     /label=AAV2 ITR
     rep_origin      3708..4163
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3795..3814)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4005..4026
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4180..4199)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4299..4321
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4359..4377)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4445..4549
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4550..4618,4619..5410)
                     /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     complement(4768..4787)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5581..6169
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6070..6089
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaacact agttattaat agtaatcaat tacggggtca
      181 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      241 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      301 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      361 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      421 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      481 tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc
      541 actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta
      601 ttttgtgcag cgatgggggc gggggggggg gggggggggg gggggggggg cgcgcgccag
      661 gcggggcggg gcggggcgag gggcggggcg gggcgaggcg gagaggtgcg gcggcagcca
      721 atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg cggcggccct
      781 ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc
      841 gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg
      901 tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt taatgacggc
      961 ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagcgc cggcaggaag
     1021 gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag
     1081 cctcggggct gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc
     1141 ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc
     1201 tttttcctac agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa
     1261 gaattgcggc ccaacggtac cggatcctct agagtcgact ccggagaagt tcctattctc
     1321 tagaaagtat aggaacttcg cagaatggta gctggattgt agctgctatt agcaatatga
     1381 aacctcttag aagttcctat tcttcaaaag gtataggaac ttcggcgcgc cgccaccatg
     1441 aaaagggtgg ctccccagaa gcagatgagc atagcaaaga aaaagaaggt gagaaggcgg
     1501 cggcgccgtc gagtgagcaa gggcgaggca gtgatcaagg agttcatgcg gttcaaggtg
     1561 cacatggagg gctccatgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
     1621 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc
     1681 tcctgggaca tcctgtcccc tcagttcatg tacggctcca gggccttcac caagcacccc
     1741 gccgacatcc ccgactacta taagcagtcc ttccccgagg gcttcaagtg ggaccgcgtg
     1801 atgaacttcg aggacggcgg cgccgtgacc gtgacccagg acacctccct ggaggacggc
     1861 accctgatct acaaggtgaa gctccgcggc accaacttcc ctcctgacgg ccccgtaatg
     1921 cagaagaaga caatgggctg ggaagcgtcc accgagcggt tgtaccccga ggacggcgtg
     1981 ctgaagggcg acattaagat ggccctgcgc ctgaaggacg gcgggcgcta cctggcggac
     2041 ttcaagacca cctacaaggc caagaagccc gtgcagatgc ccggcgccta caacgtggac
     2101 cgcaagttgg acatcacctc ccacaacgag gactacaccg tggtggaaca gtacgaacgc
     2161 tccgagggcc gccactccac cggcggcatg gacgagctgt acaagtgagc tagcgaagtt
     2221 cctatacttt ctagagaata ggaacttctt gccttaaccc agaaattatc actgttattc
     2281 tttagaatgg tgcaaagaga agttcctata ccttttgaag aataggaact tcgaattcga
     2341 tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat
     2401 tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca
     2461 tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc
     2521 tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc
     2581 tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt
     2641 cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg
     2701 gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc
     2761 ctttccttgg ctgctcgcct atgttgccac ctggattctg cgcgggacgt ccttctgcta
     2821 cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg
     2881 gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc
     2941 cccgcatcga taccgagcgc tgctcgagag atctacgggt ggcatccctg tgacccctcc
     3001 ccagtgcctc tcctggccct ggaagttgcc actccagtgc ccaccagcct tgtcctaata
     3061 aaattaagtt gcatcatttt gtctgactag gtgtccttct ataatattat ggggtggagg
     3121 ggggtggtat ggagcaaggg gcaagttggg aagacaacct gtagggcctg cggggtctat
     3181 tgggaaccaa gctggagtgc agtggcacaa tcttggctca ctgcaatctc cgcctcctgg
     3241 gttcaagcga ttctcctgcc tcagcctccc gagttgttgg gattccaggc atgcatgacc
     3301 aggctcagct aatttttgtt tttttggtag agacggggtt tcaccatatt ggccaggctg
     3361 gtctccaact cctaatctca ggtgatctac ccaccttggc ctcccaaatt gctgggatta
     3421 caggcgtgaa ccactgctcc cttccctgtc cttctgattt tgtaggtaac cacgtgcgga
     3481 ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct
     3541 cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct
     3601 cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta
     3661 cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag
     3721 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     3781 cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     3841 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     3901 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     3961 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4021 aactggaaca acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc
     4081 gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     4141 caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc
     4201 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     4261 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     4321 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     4381 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     4441 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4501 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     4561 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4621 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     4681 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     4741 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     4801 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     4861 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     4921 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     4981 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5041 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5101 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5161 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5221 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     5281 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     5341 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     5401 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     5461 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     5521 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     5581 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     5641 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     5701 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     5761 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     5821 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     5881 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     5941 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6001 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6061 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6121 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6181 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
