LOCUS       sequence-376487-        4746 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Construct for labelling plasma membranes in mammalian cells using 
            LCK-mScarlet3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4746)
  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 4746)
  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: pLCK-mScarlet3_C1
FEATURES             Location/Qualifiers
     source          1..4746
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             145..939
                     /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             145..939
                     /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(199..218)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     809..828
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1127..1146)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1171..1218
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1547..2135
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2036..2055
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2310..2613
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2614..2817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2768..2788
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2844..2853
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2889..2898
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2895..3581
                     /codon_start=1
                     /product="bright cysteine-free monomeric red fluorescent 
                     protein with fast and complete maturation (Gadella et al., 
                     2023)"
                     /label=mScarlet3
                     /translation="MDSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LRVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYWKQSFPEGFKWERVMNFEDGGA
                     VSVAQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDVVLKGDIKM
                     ALRLKDGGRYLADFKTTYRAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHS
                     TGGSGGS"
     primer_bind     3293..3312
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    3594..3659
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    3783..3904
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3820..3839)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3874..3893
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3911..4366)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4048..4069)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4260..4279
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4393..4497
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(4499..4746,1..110)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4499..4690
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4522..4542)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(4707..4746,1..96)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
       61 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa
      121 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
      181 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
      241 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
      301 ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg
      361 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
      421 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
      481 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
      541 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
      601 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
      661 aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
      721 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
      781 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
      841 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
      901 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
      961 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     1021 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     1081 gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg
     1141 agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg
     1201 gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata
     1261 ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc
     1321 ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca
     1381 tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     1441 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     1501 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     1561 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     1621 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     1681 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     1741 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     1801 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     1861 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     1921 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     1981 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     2041 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     2101 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     2161 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     2221 ctgtggataa ccgtattacc gccatgcatt agttattaat agtaatcaat tacggggtca
     2281 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     2341 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     2401 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     2461 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     2521 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     2581 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
     2641 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     2701 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     2761 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctggt
     2821 ttagtgaacc gtcagatccg ctagccacca tgggctgcgt gtgcagcagc aaccccgagc
     2881 taccggtcgc caccatggat agcaccgagg cagtgatcaa ggagttcatg cggttcaagg
     2941 tgcacatgga gggctccatg aacggccacg agttcgagat cgagggcgag ggcgagggcc
     3001 gcccctacga gggcacccag accgccaagc tgagggtgac caagggtggc cccctgccct
     3061 tctcctggga catcctgtcc cctcagttca tgtacggctc cagggccttc acgaagcacc
     3121 ccgccgacat ccccgactac tggaagcagt ccttccccga gggcttcaag tgggagcgcg
     3181 tgatgaactt cgaggacggc ggcgccgtgt ccgtggccca ggacacctcc ctggaggacg
     3241 gcaccctgat ctacaaggtg aagctccgcg gcaccaactt ccctcctgac ggccccgtaa
     3301 tgcagaagaa gacaatgggc tgggaagcat ccaccgagcg gttgtacccc gaggacgtcg
     3361 tgctgaaggg cgacattaag atggccctgc gcctgaagga cggcggccgc tacctggcgg
     3421 acttcaagac cacctacagg gccaagaagc ccgtgcagat gcccggcgcc ttcaacatcg
     3481 accgcaagtt ggacatcaca tcccacaacg aggactacac cgtggtggaa cagtacgaac
     3541 gctccgtggc ccgccactcc accggcggct ccggtggctc ctaactgtac aagtccggac
     3601 tcagatctcg agctcaagct tcgaattctg cagtcgacgg taccgcgggc ccgggatcca
     3661 ccggatctag ataactgatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
     3721 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     3781 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     3841 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     3901 cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     3961 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     4021 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     4081 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     4141 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     4201 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     4261 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     4321 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt
     4381 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     4441 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct
     4501 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     4561 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     4621 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4681 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4741 ctccgc
//
