LOCUS       sequence-96214-e        8180 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Firefly luciferase (Promega) and yeast codon-optimized yellow 
            fluorescent protein (yEVenus) fusion with PEST degron driven by 
            MET17 promoter. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8180)
  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 8180)
  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: pNB682
FEATURES             Location/Qualifiers
     source          1..8180
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /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     complement(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1721..1740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2241..2261
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2243..2261
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2312..2661
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     primer_bind     2662..2678
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2662..2678
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             2695..4344
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDVKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(2761..2779)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     terminator      5628..5815
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(5683..5707)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(6185..6204)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6186..6204)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6214..6231)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6214..6230)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6223..6245)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6375..6830
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6462..6481)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6672..6693
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(6961..7764)
                     /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"
     promoter        complement(7765..7985)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(8011..8030)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8130..8152
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttac ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcctat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctcc
     2281 ggatgcaagg gttcgaatcc cttagctctc attatttttt gctttttctc ttgaggtcac
     2341 atgatcgcaa aatggcaaat ggcacgtgaa gctgtcgata ttggggaact gtggtggttg
     2401 gcaaatgact aattaagtta gtcaaggcgc catcctcatg aaaactgtgt aacataataa
     2461 ccgaagtgtc gaaaaggtgg caccttgtcc aattgaacac gctcgatgaa aaaaataaga
     2521 tatatataag gttaagtaaa gcgtctgtta gaaaggaagt ttttcctttt tcttgctctc
     2581 ttgtcttttc atctactatt tccttcgtgt aatacagggt cgtcagatac atagatacaa
     2641 ttctattacc cccatccata ctctagaact agtggatccc ccgggttaat taacatggaa
     2701 gacgtcaaaa acataaagaa aggcccggcg ccattctatc cgctggaaga tggaaccgct
     2761 ggagagcaac tgcataaggc tatgaagaga tacgccctgg ttcctggaac aattgctttt
     2821 acagatgcac atatcgaggt ggacatcact tacgctgagt acttcgaaat gtccgttcgg
     2881 ttggcagaag ctatgaaacg atatgggctg aatacaaatc acagaatcgt cgtatgcagt
     2941 gaaaactctc ttcaattctt tatgccggtg ttgggcgcgt tatttatcgg agttgcagtt
     3001 gcgcccgcga acgacattta taatgaacgt gaattgctca acagtatggg catttcgcag
     3061 cctaccgtgg tgttcgtttc caaaaagggg ttgcaaaaaa ttttgaacgt gcaaaaaaag
     3121 ctcccaatca tccaaaaaat tattatcatg gattctaaaa cggattacca gggatttcag
     3181 tcgatgtaca cgttcgtcac atctcatcta cctcccggtt ttaatgaata cgattttgtg
     3241 ccagagtcct tcgataggga caagacaatt gcactgatca tgaactcctc tggatctact
     3301 ggtctgccta aaggtgtcgc tctgcctcat agaactgcct gcgtgagatt ctcgcatgcc
     3361 agagatccta tttttggcaa tcaaatcatt ccggatactg cgattttaag tgttgttcca
     3421 ttccatcacg gttttggaat gtttactaca ctcggatatt tgatatgtgg atttcgagtc
     3481 gtcttaatgt atagatttga agaagagctg tttctgagga gccttcagga ttacaagatt
     3541 caaagtgcgc tgctggtgcc aaccctattc tccttcttcg ccaaaagcac tctgattgac
     3601 aaatacgatt tatctaattt acacgaaatt gcttctggtg gcgctcccct ctctaaggaa
     3661 gtcggggaag cggttgccaa gaggttccat ctgccaggta tcaggcaagg atatgggctc
     3721 actgagacta catcagctat tctgattaca cccgaggggg atgataaacc gggcgcggtc
     3781 ggtaaagttg ttccattttt tgaagcgaag gttgtggatc tggataccgg gaaaacgctg
     3841 ggcgttaatc aaagaggcga actgtgtgtg agaggtccta tgattatgtc cggttatgta
     3901 aacaatccgg aagcgaccaa cgccttgatt gacaaggatg gatggctaca ttctggagac
     3961 atagcttact gggacgaaga cgaacacttc ttcatcgttg accgcctgaa gtctctgatt
     4021 aagtacaaag gctatcaggt ggctcccgct gaattggaat ccatcttgct ccaacacccc
     4081 aacatcttcg acgcaggtgt cgcaggtctt cccgacgatg acgccggtga acttcccgcc
     4141 gccgttgttg ttttggagca cggaaagacg atgacggaaa aagagatcgt ggattacgtc
     4201 gccagtcaag taacaaccgc gaaaaagttg cgcggaggag ttgtgtttgt ggacgaagta
     4261 ccgaaaggtc ttaccggaaa actcgacgca agaaaaatca gagagatcct cataaaggcc
     4321 aagaagggcg gaaagatcgc cgtgggtggg ggatcaatgt ctaaaggtga agaattattc
     4381 actggtgttg tcccaatttt ggttgaatta gatggtgatg ttaatggtca caaattttct
     4441 gtctccggtg aaggtgaagg tgatgctact tacggtaaat tgaccttaaa attgatttgt
     4501 actactggta aattgccagt tccatggcca accttagtca ctactttagg ttatggtttg
     4561 caatgttttg ctagataccc agatcatatg aaacaacatg actttttcaa gtctgccatg
     4621 ccagaaggtt atgttcaaga aagaactatt tttttcaaag atgacggtaa ctacaagacc
     4681 agagctgaag tcaagtttga aggtgatacc ttagttaata gaatcgaatt aaaaggtatt
     4741 gattttaaag aagatggtaa cattttaggt cacaaattgg aatacaacta taactctcac
     4801 aatgtttaca tcactgctga caaacaaaag aatggtatca aagctaactt caaaattaga
     4861 cacaacattg aagatggtgg tgttcaatta gctgaccatt atcaacaaaa tactccaatt
     4921 ggtgatggtc cagtcttgtt accagacaac cattacttat cctatcaatc tgccttatcc
     4981 aaagatccaa acgaaaagag agaccacatg gtcttgttag aatttgttac tgctgctggt
     5041 attacccatg gtattgatga atctcgagaa gcatccaact tgaacatttc gagaaagctt
     5101 accatatcaa ccccatcatg ctctttcgaa aattcaaata gcacatccat tccttcgccc
     5161 gcttcctcat ctcaaagcca cactccaatg agaaacatga gctcactctc tgataacagc
     5221 gttttcagcc ggaatatgga acaatcatca ccaatcactc caagtatgta ccaatttggt
     5281 cagcagcagt caaacagtat atgtggtagc accgttagtg tgaatagtct ggtgaataca
     5341 aataacaaac aaaggatcta cgaacaaatc acgggtccta acagcaataa cgcaaccaat
     5401 gattatattg atttgctaaa cctaaatgag tctaacaagg aaaaccaaaa tcccgcaacg
     5461 gcgcattacc tcaatggggg cccacccaag acaagcttca ttaaccatgg aatgttcccc
     5521 tcgccaactg ggaccataaa tagcggtaaa tctagcagtg cctcatcttt aatttctttt
     5581 ggtatgggca atacccaagt aatatagggc gcgccacttc taaataagcg aatttcttat
     5641 gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac aaattttaaa
     5701 gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc ctgtaggtca
     5761 ggttgctttc tcaggtatag tatgaggtcg ctcttattga ccacacctct accggcagat
     5821 ccgctaggga taacagggta atatagatct gtttagcttg cctcgtcccc gccgggtcac
     5881 ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca
     5941 ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt
     6001 gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag
     6061 acctgcgagc agggaaacgc tcccctcaca gacgcgtgta cgcatgtaac attatactga
     6121 aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt aatttgcggc cggtacccaa
     6181 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     6241 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     6301 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     6361 tggcgaatgg cgcgacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     6421 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     6481 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     6541 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     6601 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     6661 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     6721 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     6781 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt cctgatgcgg
     6841 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tagggtaata actgatataa
     6901 ttaaattgaa gctctaattt gtgagtttag tatacatgca tttacttata atacagtttt
     6961 ttagttttgc tggccgcatc ttctcaaata tgcttcccag cctgcttttc tgtaacgttc
     7021 accctctacc ttagcatccc ttccctttgc aaatagtcct cttccaacaa taataatgtc
     7081 agatcctgta gagaccacat catccacggt tctatactgt tgacccaatg cgtctccctt
     7141 gtcatctaaa cccacaccgg gtgtcataat caaccaatcg taaccttcat ctcttccacc
     7201 catgtctctt tgagcaataa agccgataac aaaatctttg tcgctcttcg caatgtcaac
     7261 agtaccctta gtatattctc cagtagatag ggagcccttg catgacaatt ctgctaacat
     7321 caaaaggcct ctaggttcct ttgttacttc ttctgccgcc tgcttcaaac cgctaacaat
     7381 acctgggccc accacaccgt gtgcattcgt aatgtctgcc cattctgcta ttctgtatac
     7441 acccgcagag tactgcaatt tgactgtatt accaatgtca gcaaattttc tgtcttcgaa
     7501 gagtaaaaaa ttgtacttgg cggataatgc ctttagcggc ttaactgtgc cctccatgga
     7561 aaaatcagtc aagatatcca catgtgtttt tagtaaacaa attttgggac ctaatgcttc
     7621 aactaactcc agtaattcct tggtggtacg aacatccaat gaagcacaca agtttgtttg
     7681 cttttcgtgc atgatattaa atagcttggc agcaacagga ctaggatgag tagcagcacg
     7741 ttccttatat gtagctttcg acatgattta tcttcgtttc ctgcaggttt ttgttctgtg
     7801 cagttgggtt aagaatactg ggcaatttca tgtttcttca acactacata tgcgtatata
     7861 taccaatcta agtctgtgct ccttccttcg ttcttccttc tgttcggaga ttaccgaatc
     7921 aaaaaaattt caaagaaacc gaaatcaaaa aaaagaataa aaaaaaaatg atgaattgaa
     7981 ttgaaaagct gtggtatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     8041 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     8101 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     8161 gtcatcaccg aaacgcgcga
//
