LOCUS       sequence-189401-        5801 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of GFP-FKBP at the plasma membrane in mammalian cells. 
            Plasma membrane targeting is via CD8 alpha chain..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5801)
  AUTHORS   Clarke NI, Royle SJ
  TITLE     FerriTag is a new genetically-encoded inducible tag for correlative 
            light-electron microscopy.
  JOURNAL   Nat Commun. 2018 Jul 4;9(1):2604. doi: 10.1038/s41467-018-04993-0.
  PUBMED    29973588
REFERENCE   2  (bases 1 to 5801)
  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: "Nat 
            Commun."; date: "2018-07-4"; volume: "9(1):2604. doi"; pages: " 
            10.1038/s41467-018-04993-0"
COMMENT     Sequence Label: CD8-GFP-FKBP
FEATURES             Location/Qualifiers
     source          1..5801
                     /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(1382..1384,1385..2092)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     CDS             join(1382..1384,1385..2092)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     primer_bind     complement(1424..1445)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1685..1704)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2032..2053
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(2141..2143,2144..2467)
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKLE"
     polyA_signal    2589..2710
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2626..2645)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2680..2699
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2717..3172)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2854..2875)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3066..3085
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3199..3303
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3305..3662
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3305..3496
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3328..3348)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3513..3648
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3697..4491
                     /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             3697..4491
                     /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(3751..3770)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4361..4380
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4679..4698)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4723..4770
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5099..5687
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5588..5607
                     /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 atctcgagct caagcttcga attcggcggc tcccgcgcgc ctcccctcgc
      661 gcccgagctt cgagccaagc agcgtcctgg ggagcgcgtc atggccttac cagtgaccgc
      721 cttgctcctg ccgctggcct tgctgctcca cgccgccagg ccgagccagt tccgggtgtc
      781 gccgctggat cggacctgga acctgggcga gacagtggag ctgaagtgcc aggtgctgct
      841 gtccaacccg acgtcgggct gctcgtggct cttccagccg cgcggcgccg ccgccagtcc
      901 caccttcctc ctatacctct cccaaaacaa gcccaaggcg gccgaggggc tggacaccca
      961 gcggttctcg ggcaagaggt tgggggacac cttcgtcctc accctgagcg acttccgccg
     1021 agagaacgag ggctactatt tctgctcggc cctgagcaac tccatcatgt acttcagcca
     1081 cttcgtgccg gtcttcctgc cagcgaagcc caccacgacg ccagcgccgc gaccaccaac
     1141 accggcgccc accatcgcgt cgcagcccct gtccctgcgc ccagaggcgt gccggccagc
     1201 ggcggggggc gcagtgcaca cgagggggct ggacttcgcc tgtgatatct acatctgggc
     1261 gcccttggcc gggacttgtg gagtccttct cctgtcactg gttatcaccc tttactgcaa
     1321 gcggcttaag cgaagaagga tccccgccgc cgccgccgcc gccgccgccg tcagaccggt
     1381 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     1441 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     1501 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     1561 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     1621 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     1681 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     1741 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     1801 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     1861 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     1921 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     1981 cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     2041 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt accgaatcga
     2101 tgtgaatggg ggccctgaac ccgccaagaa cctgtacacc atgggagtgc aggtggaaac
     2161 catctcccca ggagacgggc gcaccttccc caagcgcggc cagacctgcg tggtgcacta
     2221 caccgggatg cttgaagatg gaaagaaatt tgattcctcc cgggacagaa acaagccctt
     2281 taagtttatg ctaggcaagc aggaggtgat ccgaggctgg gaagaagggg ttgcccagat
     2341 gagtgtgggt cagagagcca aactgactat atctccagat tatgcctatg gtgccactgg
     2401 gcacccaggc atcatcccac cacatgccac tctcgtcttc gatgtggagc ttctaaaact
     2461 ggaatgagcg gccgcgactc tagatcataa tcagccatac cacatttgta gaggttttac
     2521 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     2581 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2641 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2701 atgtatctta aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     2761 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     2821 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     2881 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     2941 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     3001 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     3061 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     3121 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc
     3181 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     3241 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3301 agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     3361 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     3421 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3481 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     3541 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     3601 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     3661 aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     3721 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     3781 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     3841 gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg
     3901 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     3961 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4021 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4081 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4141 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4201 gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat
     4261 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     4321 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     4381 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     4441 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     4501 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     4561 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     4621 tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg
     4681 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga
     4741 ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc
     4801 actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt
     4861 ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg
     4921 caggccctgc catagcctca ggttactcat atatacttta gattgattta aaacttcatt
     4981 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     5041 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     5101 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     5161 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     5221 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     5281 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     5341 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     5401 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     5461 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     5521 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     5581 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     5641 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     5701 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     5761 tatcccctga ttctgtggat aaccgtatta ccgccatgca t
//
