LOCUS       sequence-439142-        5554 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express a label of ER-plasma membrane contacts based on 
            GFP-MAPPER (#117721), but with mScarletI3 tag and replacing FRB with
            a protein of similar structure and sequence length (Neoleukin-2/15).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5554)
  AUTHORS   Downie L, Ferrandiz N, Courthold E, Jones M, Royle SJ
  TITLE     Nondisruptive inducible labeling of ER-membrane contact sites using 
            the Lamin B receptor.
  JOURNAL   PLoS Biol. 2025 Jul 10;23(7):e3003249. doi: 
            10.1371/journal.pbio.3003249. eCollection 2025 Jul.
  PUBMED    40638559
REFERENCE   2  (bases 1 to 5554)
  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: "PLoS 
            Biol."; date: "2025-07-10"; volume: "23(7):e3003249. doi"; pages: " 
            10.1371/journal.pbio.3003249. eCollection 2025 Jul"
COMMENT     Sequence Label: mScarletI3-6DG5-MAPPER
FEATURES             Location/Qualifiers
     source          1..5554
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      597..606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             681..1364
                     /codon_start=1
                     /product="bright cysteine-free monomeric red fluorescent 
                     protein with fast and complete maturation (Gadella et al., 
                     2023)"
                     /label=mScarlet3
                     /translation="DSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYWKQSFPEGFKWERVMIFEDGGTV
                     SVTQDTSLEDGTLIYKVKLRGGNFPPDGPVMQKRTMGWEASTERLYPEDVVLKGDIKMA
                     LRLKDGGRYLADFKTTYKAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHST
                     GGSGGS"
     primer_bind     complement(808..826)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     polyA_signal    2342..2463
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2379..2398)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2433..2452
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2470..2925)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2607..2628)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2819..2838
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2952..3056
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3058..3415
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3058..3249
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3081..3101)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3266..3401
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3450..4244
                     /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             3450..4244
                     /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(3504..3523)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4114..4133
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4432..4451)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4476..4523
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4852..5440
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5341..5360
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcca
      601 ccatggatgt atgcgtccgt cttgccctgt ggctcctctg gggactcctc ctgcaccagg
      661 gccagagcct cgcaccggtc gatagcaccg aggcagtgat caaggagttc atgcggttca
      721 aggtgcacat ggagggctcc atgaacggcc acgagttcga gatcgagggc gagggcgagg
      781 gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt ggccccctgc
      841 ccttctcctg ggacatcctg tcccctcagt tcatgtacgg ctccagggcc ttcatcaagc
      901 accccgccga catccccgac tactggaagc agtccttccc cgagggcttc aagtgggagc
      961 gcgtgatgat cttcgaggac ggcggcaccg tgtccgtgac ccaggacacc tccctggagg
     1021 acggcaccct gatctacaag gtgaagctcc gcggcggcaa cttccctcct gacggccccg
     1081 taatgcagaa gaggacaatg ggctgggaag catccaccga gcggttgtac cccgaggacg
     1141 tcgtgctgaa gggcgacatt aagatggccc tgcgcctgaa ggacggcggc cgctacctgg
     1201 cggacttcaa gaccacctac aaggccaaga agcccgtgca gatgcccggc gccttcaaca
     1261 tcgaccgcaa gttggacatc acctcccaca acgaggacta caccgtggtg gaacagtacg
     1321 aacgctccgt ggcccgccac tccaccggcg gctccggtgg ctccttgtac aagtccggat
     1381 caactagtct ggatacagtg ctctttgggc ctcctctctt gactcgccat aatcacctca
     1441 aggacttcat gctggtggtg tctatcgtta ttggtgtggg cggctgctgg tttgcctata
     1501 tccagaaccg ttactccaag gagcacggag cgggcgctgg tgccggagct ggagcaggtg
     1561 ctctcgagga agcagcagca cgagaagctg ctgctagaga agctgctgct agagaagcag
     1621 cagcacgagt cgagggctct cacatgccca agaaaaagat ccagctgcac gccgagcacg
     1681 ctctgtacga cgccctgatg atcctgaaca tcgtgaagac caacagccct cctgctgagg
     1741 aaaagctgga agattacgcc tttaacttcg agctgatcct ggaagaaatc gccagactgt
     1801 tcgagtccgg cgaccaaaag gacgaggccg agaaggccaa gcggatgaag gaatggatga
     1861 aaagaatcaa gacaaccgcc agcgaggatg agcaggagga aatggccaat gccattatca
     1921 ccatcctcca gagctggatc ttcagctcag ctggaggcag tgcaggtggc tctgccggag
     1981 gtagaattct agaagctgct gctagagaag ctgctgctag agaagctgct gctagagaag
     2041 ctgctgctag agaagctgct gctagagaag ctgctgctag agcaattcta atggagaaaa
     2101 aatctaagcc caaaaacagt gtatggaaga ggctaaaatc accattccgg aagaagaaag
     2161 attcagtaac ttgaaagctt cgaattctgc agtcgacggt accgcgggcc cgggatccac
     2221 cggatctaga taactgatca taatcagcca taccacattt gtagaggttt tacttgcttt
     2281 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
     2341 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     2401 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     2461 ttaacgcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     2521 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     2581 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     2641 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     2701 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     2761 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     2821 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     2881 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc
     2941 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     3001 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg
     3061 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     3121 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     3181 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3241 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     3301 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     3361 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg
     3421 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     3481 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     3541 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     3601 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
     3661 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     3721 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     3781 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     3841 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     3901 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     3961 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     4021 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     4081 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     4141 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     4201 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     4261 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     4321 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     4381 gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg
     4441 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
     4501 gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt
     4561 cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc
     4621 ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc
     4681 tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt
     4741 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     4801 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4861 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     4921 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     4981 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     5041 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     5101 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     5161 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     5221 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5281 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5341 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5401 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5461 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5521 tgattctgtg gataaccgta ttaccgccat gcat
//
