LOCUS       sequence-494351-        4735 bp    DNA     circular SYN 02-DEC-2025
DEFINITION  Expression of mScarelt3-S2 fused with Lifeact. This construct is 
            used for imaging..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4735)
  AUTHORS   Ding Y, He W, Wang K, Xue F, Zheng K, Zhao S, Qi D, Li T, Xia P, Pan
            L, Chen L, Chen Z, Yuan L, Xu P
  TITLE     A highly photostable monomeric red fluorescent protein for 
            dual-color 3D STED and time-lapse 3D SIM imaging.
  JOURNAL   Nat Methods. 2025 Nov 26. doi: 10.1038/s41592-025-02962-2.
  PUBMED    41299071
REFERENCE   2  (bases 1 to 4735)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 2, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Methods. 2025 Nov 26. doi: 10.1038/s41592-025-02962-2."
COMMENT     Sequence Label: Lifeact-mScarlet3-S2
FEATURES             Location/Qualifiers
     source          1..4735
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        110..413
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        414..617
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     568..588
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             688..738
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     regulatory      754..763
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             760..1446
                     /codon_start=1
                     /product="bright cysteine-free monomeric red fluorescent 
                     protein with fast and complete maturation (Gadella et al., 
                     2023)"
                     /label=mScarlet3
                     /translation="MDSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LRVTKGGPLPFSWDILSPQFQYGSRAFTKHPADIPDYWKQSFPEGFKWERVMNFEDGGA
                     VSVAQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDVVLKGDIKH
                     ALRLKDGGRYLADFKTTYRAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHS
                     TGGSGGS"
     primer_bind     1158..1177
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1572..1693
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1609..1628)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1663..1682
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1700..2155)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1837..1858)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2049..2068
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2182..2286
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2288..2645
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2288..2479
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2311..2331)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2496..2631
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2680..3474
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2680..3474
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2734..2753)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3344..3363
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3662..3681)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3706..3753
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4082..4670
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4571..4590
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat
       61 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      121 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      181 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      241 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      301 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
      361 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
      421 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
      481 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
      541 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
      601 tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggactcaga
      661 tctcgagctc aagcttcgaa ttccaccatg ggcgtggccg acctgatcaa gaagttcgag
      721 agcatcagca aggaggaggg ggatccaccg gtcgccacca tggatagcac cgaggcagtg
      781 atcaaggagt tcatgcggtt caaggtgcac atggagggct ccatgaacgg ccacgagttc
      841 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgagg
      901 gtgaccaagg gtggccccct gcccttctcc tgggacatcc tgtcccctca gttccagtac
      961 ggctccaggg ccttcacgaa gcaccccgcc gacatccccg actactggaa gcagtccttc
     1021 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgc cgtgtccgtg
     1081 gcccaggaca cctccctgga ggacggcacc ctgatctaca aggtgaagct ccgcggcacc
     1141 aacttccctc ctgacggccc cgtaatgcag aagaagacaa tgggctggga agcatccacc
     1201 gagcggttgt accccgagga cgtcgtgctg aagggcgaca ttaagcacgc cctgcgcctg
     1261 aaggacggcg gccgctacct ggcggacttc aagaccacct acagggccaa gaagcccgtg
     1321 cagatgcccg gcgccttcaa catcgaccgc aagttggaca tcacatccca caacgaggac
     1381 tacaccgtgg tggaacagta cgaacgctcc gtggcccgcc actccaccgg cggctccggt
     1441 ggctcctaaa gcggccgcga ctctagatca taatcagcca taccacattt gtagaggttt
     1501 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     1561 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     1621 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     1681 tcaatgtatc ttaaggcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     1741 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     1801 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     1861 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     1921 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     1981 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     2041 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     2101 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt
     2161 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     2221 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     2281 aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     2341 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2401 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2461 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     2521 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2581 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2641 gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     2701 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     2761 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     2821 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta
     2881 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     2941 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     3001 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     3061 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     3121 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     3181 gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc
     3241 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     3301 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     3361 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     3421 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     3481 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     3541 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     3601 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa
     3661 ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac
     3721 agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct
     3781 ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct
     3841 tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg
     3901 cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc
     3961 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     4021 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     4081 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     4141 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     4201 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     4261 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4321 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4381 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4441 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     4501 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     4561 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     4621 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     4681 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctt
//
