LOCUS       sequence-76085-e        5593 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MagFRET-6 in the cytosol of mammalian cells. Weak affinity
            control FRET sensor for MagFRET-1. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5593)
  AUTHORS   Lindenburg LH, Vinkenborg JL, Oortwijn J, Aper SJ, Merkx M
  TITLE     MagFRET: The First Genetically Encoded Fluorescent Mg(2+) Sensor.
  JOURNAL   PLoS One. 2013 Dec 2;8(12):e82009. doi: 
            10.1371/journal.pone.0082009.
  PUBMED    24312622
REFERENCE   2  (bases 1 to 5593)
  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 One.";
            date: "2013-12-2"; volume: "8(12):e82009. doi"; pages: " 
            10.1371/journal.pone.0082009"
COMMENT     Sequence Label: pCMVMagFRET-6
FEATURES             Location/Qualifiers
     source          1..5593
                     /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      600..609
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(615..617,618..620,621..1133)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIE"
     CDS             join(1539..1541,1542..1544,1545..2255)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    2381..2502
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2418..2437)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2472..2491
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2509..2964)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2646..2667)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2858..2877
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2991..3095
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3097..3454
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3097..3288
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3120..3140)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3305..3440
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3489..4283
                     /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             3489..4283
                     /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(3543..3562)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4153..4172
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4471..4490)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4515..4562
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4891..5479
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5380..5399
                     /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 gctagcgccg
      601 ccaccatggg ccatatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
      661 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
      721 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggt aagctgcccg
      781 tgccctggcc caccctcgtg accaccctga cctggggcgt gcagtgcttc gcccgctacc
      841 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
      901 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
      961 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     1021 acatcctggg gcacaagctg gagtacaacg ccatcagcga caacgtctat atcaccgccg
     1081 acaagcagaa gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggca
     1141 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     1201 tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc
     1261 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactagc gaggaacaga
     1321 aacaagaaat taaagatgct tttgaactgt ttgatacaga caaagaagaa gaaatagatt
     1381 atcatgaact gaaggtggca atgagagcct tggggtttga tgtaaaaaaa gctgatgtac
     1441 tgaagattct taaagattat gacagagaag ccacagggaa aatcaccttt gaagatttta
     1501 atgaagttgt gacagactgg atattggaaa gagatgccat ggtgagcaag ggcgaggagc
     1561 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1621 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     1681 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ttcggctacg
     1741 gcctgatgtg cttcgcccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1801 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     1861 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1921 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gcttgagtac aactacaaca
     1981 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     2041 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     2101 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagctac cagtccgccc
     2161 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     2221 ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgcgac tctagatcat
     2281 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     2341 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     2401 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     2461 gcattctagt tgtggtttgt ccaaactcat caatgtatct taaggcgtaa attgtaagcg
     2521 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     2581 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     2641 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     2701 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     2761 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     2821 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     2881 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     2941 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     3001 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     3061 gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg
     3121 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3181 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3241 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3301 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3361 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     3421 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
     3481 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     3541 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     3601 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     3661 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     3721 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3781 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     3841 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     3901 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     3961 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
     4021 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     4081 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     4141 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     4201 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     4261 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     4321 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4381 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4441 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
     4501 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
     4561 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     4621 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     4681 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
     4741 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4801 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     4861 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     4921 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     4981 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     5041 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     5101 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     5161 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     5221 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     5281 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     5341 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     5401 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     5461 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5521 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5581 taccgccatg cat
//
