LOCUS       sequence-439254-        5824 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express a truncated LBR (encoding amino acids 1-245) with 
            N-terminal FKBP tag accessible to the cytoplasm and C-terminal GFP 
            in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5824)
  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 5824)
  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: FKBP-LBR(1-245)-GFP
FEATURES             Location/Qualifiers
     source          1..5824
                     /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"
     CDS             join(666..668,669..989)
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKLE"
     regulatory      1764..1773
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1770..1772,1773..1775,1776..2489)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1770..1772,1773..1775,1776..2486)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1815..1836)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2076..2095)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2423..2444
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2612..2733
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2649..2668)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2703..2722
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2740..3195)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2877..2898)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3089..3108
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3222..3326
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3328..3685
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3328..3519
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3351..3371)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3536..3671
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3720..4514
                     /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             3720..4514
                     /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(3774..3793)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4384..4403
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4702..4721)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4746..4793
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5122..5710
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5611..5630
                     /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 gctagcgcta
      601 ccggactcag atctcgagag atcgatgtga atgggggccc tgaacccgcc aagaacctgt
      661 acaccatggg agtgcaggtg gaaaccatct ccccaggaga cgggcgcacc ttccccaagc
      721 gcggccagac ctgcgtggtg cactacaccg ggatgcttga agatggaaag aaatttgatt
      781 cctcccggga cagaaacaag ccctttaagt ttatgctagg caagcaggag gtgatccgag
      841 gctgggaaga aggggttgcc cagatgagtg tgggtcagag agccaaactg actatatctc
      901 cagattatgc ctatggtgcc actgggcacc caggcatcat cccaccacat gccactctcg
      961 tcttcgatgt ggagcttcta aaactggaac ggtaccgtga accgtcagat ccgctagcta
     1021 tgccaagtag gaaatttgcc gatggtgaag tggtaagagg tcgatggcct gggagttcac
     1081 tttattatga agtagaaatt ctgagccacg acagcacctc ccagctttac actgtgaagt
     1141 ataaagatgg aacagagctt gaattgaaag agaatgatat taagccttta acttccttta
     1201 ggcaaaggaa aggtggctca acttccagtt ccccttccag acgccgaggg agtcgatcaa
     1261 ggtcacgctc ccgatcccct ggtcgaccac ctaaaagtgc ccgccgatct gcttctgctt
     1321 cccaccaggc cgacattaag gaagcaagga gggaagtgga agttaaattg actccgctga
     1381 ttctgaagcc atttggaaat agcatcagca gatataatgg ggagcctgag catattgaga
     1441 gaaatgacgc acctcataaa aatacacagg aaaaattcag tttgtcacaa gaaagcagtt
     1501 acatagcaac acagtatagc cttcgtccaa gaagagaaga agtcaaatta aaagaaatag
     1561 attctaagga agaaaaatac gttgcaaaag aactggcagt gagaaccttt gaagtgaccc
     1621 ccatccgggc aaaggacttg gagtttggag gagtacctgg tgtgtttctc atcatgtttg
     1681 gcctgcctgt gttcctcttc ctgttgctgt tgatgtgtaa acagaaagat cccagtcttc
     1741 tgaatttccc tcctctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg
     1801 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
     1861 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
     1921 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
     1981 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     2041 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     2101 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     2161 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
     2221 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
     2281 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
     2341 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag
     2401 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
     2461 ctctcggcat ggacgagctg tacaagtaaa gcggccgcga ctctagatca taatcagcca
     2521 taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct
     2581 gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta
     2641 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     2701 ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc gttaatattt
     2761 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     2821 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     2881 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     2941 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     3001 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     3061 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     3121 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     3181 cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3241 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3301 ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg
     3361 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3421 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3481 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     3541 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     3601 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt
     3661 ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga tcgtttcgca
     3721 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     3781 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     3841 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     3901 aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     3961 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     4021 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     4081 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     4141 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     4201 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg
     4261 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     4321 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     4381 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     4441 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     4501 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     4561 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     4621 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     4681 ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc
     4741 tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg ttcataaacg
     4801 cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca
     4861 atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg
     4921 gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact catatatact
     4981 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     5041 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     5101 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     5161 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5221 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5281 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5341 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5401 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     5461 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5521 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5581 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     5641 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     5701 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     5761 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat
     5821 gcat
//
