LOCUS       sequence-164057-        6091 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian vector for the expression of a dual read-out BRET-FRET 
            sensor for intracellular Zn2+ imaging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6091)
  AUTHORS   Aper SJ, Dierickx P, Merkx M
  TITLE     Dual Readout BRET/FRET Sensors for Measuring Intracellular Zinc.
  JOURNAL   ACS Chem Biol. 2016 Oct 21;11(10):2854-2864. Epub 2016 Sep 1.
  PUBMED    27547982
REFERENCE   2  (bases 1 to 6091)
  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: "ACS Chem 
            Biol."; date: "2016-10-21"; volume: "11(10)"; pages: "2854-2864. 
            Epub 2016 Sep 1"
COMMENT     Sequence Label: pCMV-BLZinCh-1
FEATURES             Location/Qualifiers
     source          1..6091
                     /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(1107..1109,1110..1112,1113..1823)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSHLCKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2025..2027,2028..2030,2031..2741)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSHLCKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    2879..3000
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2916..2935)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2970..2989
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3007..3462)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3144..3165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3356..3375
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3489..3593
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3595..3952
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3595..3786
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3618..3638)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3803..3938
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3987..4781
                     /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             3987..4781
                     /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(4041..4060)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4651..4670
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4969..4988)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5013..5060
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5389..5977
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5878..5897
                     /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 gtgctagcca
      601 tggtatttac tcttgaagat tttgtcggtg attggcgcca gaccgccggc tataacctgg
      661 accaagtgct tgaacagggc ggggttagca gcctgtttca aaacctgggg gtgagtgtca
      721 cgccaattca gcgcatcgtt ctgtcgggag agaatggtct gaaaatcgat atccacgtca
      781 ttatcccgta cgaaggtctt tctggtgatc agatggggca gatagaaaaa atattcaaag
      841 tggtgtaccc agtagacgat catcacttca aggttatact gcactatggc accctcgtta
      901 tcgatggcgt tactccgaat atgatcgatt actttgggcg tccttatgaa ggtattgcgg
      961 tgttcgacgg taaaaaaatt acggttaccg ggacgctctg gaatggtaat aaaatcattg
     1021 atgagcgctt gataaaccca gatggcagcc ttctgttcag agttacgata aacggggtta
     1081 cgggttggcg actgtgcgaa agacatatgg tgagcaaggg cgaggagctg ttcaccgggg
     1141 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     1201 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     1261 gtaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctggggc gtgcagtgct
     1321 tcgcccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     1381 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     1441 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     1501 aggaggacgg caacatcctg gggcacaagc tggagtacaa cgccatcagc gacaacgtct
     1561 atatcaccgc cgacaagcag aagaacggca tcaaggccaa cttcaagatc cgccacaaca
     1621 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     1681 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtcccacctg tgcaaagacc
     1741 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     1801 tcggcatgga cgagctgtac aagtccggag gcggcgagct cattcgtggc ggatccggcg
     1861 gaagcggcgg atccggcggt agcggcggat ccggcggctc cggcggatcc ggcggcagcg
     1921 gcggatccgg tggaagcggt ggatccggtg gtagcggtgg atccggtgga agcggtggat
     1981 ccggtggtag cggtggatcc gggggtccgc ggggctcggt acccatggtg agcaagggcg
     2041 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     2101 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     2161 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttcg
     2221 gctacggcct gatgtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
     2281 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     2341 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     2401 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctt gagtacaact
     2461 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     2521 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     2581 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agctaccagt
     2641 cccacctgtg caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     2701 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtacaagcgg ccgcactcga
     2761 gctaagactc tagatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     2821 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     2881 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     2941 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     3001 aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     3061 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     3121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     3181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     3241 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     3301 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     3361 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     3421 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     3481 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3541 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg
     3601 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     3661 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     3721 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     3781 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     3841 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     3901 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc
     3961 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     4021 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     4081 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     4141 acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca
     4201 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     4261 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     4321 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     4381 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     4441 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     4501 ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     4561 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     4621 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     4681 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     4741 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     4801 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     4861 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     4921 cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa
     4981 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     5041 cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     5101 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     5161 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     5221 catagcctca ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5281 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5341 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     5401 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     5461 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     5521 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     5581 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     5641 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     5701 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     5761 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     5821 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     5881 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     5941 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     6001 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     6061 ttctgtggat aaccgtatta ccgccatgca t
//
