LOCUS       sequence-96261-e        6043 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Disintegrator plasmid encoding MET17 promoter driven NanoLuc 
            (Promega) with nuclear localization signal (NLS) .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6043)
  AUTHORS   Mazo-Vargas A, Park H, Aydin M, Buchler NE
  TITLE     Measuring fast gene dynamics in single cells with time-lapse 
            luminescence microscopy
  JOURNAL   Mol Biol Cell. 2014 Sep 17. pii: mbc.E14-07-1187.
  PUBMED    25232010
REFERENCE   2  (bases 1 to 6043)
  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: "Mol Biol 
            Cell. 2014 Sep 17. pii: mbc.E14-07-1187."
COMMENT     Sequence Label: pNB330
FEATURES             Location/Qualifiers
     source          1..6043
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(542..559)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(713..1301)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(793..812)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1472..2263,2264..2332))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2095..2114
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2333..2437)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2505..2523
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2530..2750
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             2751..3554
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     primer_bind     complement(3672..3694)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3794..3813
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4853..5202
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     CDS             5226..5246
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5250..5762
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     terminator      5807..5994
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(5862..5886)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
ORIGIN
        1 cagcatcatt gatgtaatct tctactagtt gatccaattg tccagcagct ctaataacaa
       61 tcaacccacg tggtttagca acacctaagt aaatggtttg tctggctggg ggatatgcag
      121 ggtcgataac tgaaaaggtt gcgccggctt tcaagacacc catcacacat accatcaaat
      181 ccacacccct agaagaatag atcatcacta catcacctct tttgatacct gttttaatca
      241 aataatgggc aactatgtta gaagtgcggt tgatgtcgcg ataagtgaaa gaacgggact
      301 tgtcggaatt tagtgttgga gtctccacaa cacaggttct ctctgggaag gcttcagcat
      361 tgtcctggaa aatgtcgtga atacacccca cgaaatcgca ccagcccaag ttcttagttg
      421 gatcaggtaa actatccttg gatgatgcgg tgatcagaga gaggatctcc cgggtcctgc
      481 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
      541 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
      601 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
      661 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
      721 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
      781 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
      841 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
      901 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
      961 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     1021 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     1081 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     1141 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     1201 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     1261 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     1321 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     1381 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     1441 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     1501 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     1561 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     1621 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     1681 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     1741 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     1801 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     1861 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     1921 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     1981 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     2041 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     2101 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     2161 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     2221 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     2281 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     2341 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     2401 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     2461 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     2521 ggccagcttt tcaattcaat tcatcatttt ttttttattc ttttttttga tttcggtttc
     2581 tttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg aaggagcaca
     2641 gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat tgcccagtat
     2701 tcttaaccca actgcacaga acaaaaacat gcaggaaacg aagataaatc atgtcgaaag
     2761 ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag ctatttaata
     2821 tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc accaaggaat
     2881 tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca catgtggata
     2941 tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta tccgccaagt
     3001 acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca gtcaaattgc
     3061 agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat gcacacggtg
     3121 tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta acaaaggaac
     3181 ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct actggagaat
     3241 atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg
     3301 ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg
     3361 tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg
     3421 tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg
     3481 ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg
     3541 gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta
     3601 gagcttcaat ttaattatat cagttattac ccgccctttc gtctcgcgcg tttcggtgat
     3661 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     3721 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     3781 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     3841 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg
     3901 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca ggcgtcgaga
     3961 agtgacatca cccgaaaaga agctaagtct ttctcctaat tcatatttaa ttattgtaca
     4021 tggacatatc atacgtaatg ctcaacctta agctgctgcg gagcttccac gagcaccagc
     4081 acctgaagca actagactta tttgcgcttg agttagttct atacttggca gtggaagttt
     4141 gtcattatga gttggtggag gtaaaaatcc aaccttgttt aaaaatgcaa tatatatacc
     4201 aacctcttct ggagtaacac ctattccatt agaccaatca acacctgtcc atgcggtgtc
     4261 tttctttaaa gatgccacgg cgttcctatc gtctagttcc ggagctttgg tactttcagg
     4321 taagttgtct aagaccatgt gtagcaaagg atacaacgca ttttcttcat tcctgtcaat
     4381 aacagacgcc tccaatgatt tcttccattt agaatagctt tcgatttcga catcgtaacc
     4441 ataatcgtgt aaagtataca agtagtcttt gaataatatt cttgggtgac ccgttacttg
     4501 agcaacggcc aattcatttt ctttgggagg attcaaagac gtagcaacaa caacacgagc
     4561 aacatgatct actggaacca tattcacgga attttcgata tctggaatct tacctaattg
     4621 gactgaacct ttcaaaaatc tcaataagaa atcatctgtg tttgaagaac cattggcaga
     4681 ggcacctgtt acgtaacctg gtctgacaat acacccacgt aggccccttt cacctgcacg
     4741 tctaatgatg tactcagcag cccatttgga ctgaccatat ccaccagtga gcccgcttgc
     4801 agaggctgga attcgagctc cggatgcaag ggttcgaatc ccttagctct cattattttt
     4861 tgctttttct cttgaggtca catgatcgca aaatggcaaa tggcacgtga agctgtcgat
     4921 attggggaac tgtggtggtt ggcaaatgac taattaagtt agtcaaggcg ccatcctcat
     4981 gaaaactgtg taacataata accgaagtgt cgaaaaggtg gcaccttgtc caattgaaca
     5041 cgctcgatga aaaaaataag atatatataa ggttaagtaa agcgtctgtt agaaaggaag
     5101 tttttccttt ttcttgctct cttgtctttt catctactat ttccttcgtg taatacaggg
     5161 tcgtcagata catagataca attctattac ccccatccat actcttaatt aacaccatgg
     5221 caccaccaaa gaaaaagaga aaagtagcca tggtcttcac actcgaagat ttcgttgggg
     5281 actggcgaca gacagccggc tacaacctgg accaagtcct tgaacaggga ggtgtgtcca
     5341 gtttgtttca gaatctcggg gtgtccgtaa ctccgatcca aaggattgtc ctgagcggtg
     5401 aaaatgggct gaagatcgac atccatgtca tcatcccgta tgaaggtctg agcggcgacc
     5461 aaatgggcca gatcgaaaaa atttttaagg tggtgtaccc tgtggatgat catcacttta
     5521 aggtgatcct gcactatggc acactggtaa tcgacggggt tacgccgaac atgatcgact
     5581 atttcggacg gccgtatgaa ggcatcgccg tgttcgacgg caaaaagatc actgtaacag
     5641 ggaccctgtg gaacggcaac aaaattatcg acgagcgcct gatcaacccc gacggctccc
     5701 tgctgttccg agtaaccatc aacggagtga ccggctggcg gctgtgcgaa cgcattctgg
     5761 cgggtggggg atcattgtac aaataaggcg cgccacttct aaataagcga atttcttatg
     5821 atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca aattttaaag
     5881 tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc tgtaggtcag
     5941 gttgctttct caggtatagt atgaggtcgc tcttattgac cacacctcta ccggcagatc
     6001 cgctagggat aacagggtaa tatagatctc tcgaggcggc cgc
//
