LOCUS       sequence-99750-e        6906 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Green click beetle luciferase from Promega driven by MET17 
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6906)
  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 6906)
  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: pNB768
FEATURES             Location/Qualifiers
     source          1..6906
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(340..359)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(341..359)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(369..386)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(369..385)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(378..400)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      530..985
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(617..636)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     827..848
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1116..1919)
                     /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(1920..2140)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(2166..2185)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2285..2307
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2345..2363)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2431..2535
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2536..2604,2605..3396)
                     /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(2754..2773)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3567..4155
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4056..4075
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4309..4326
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(4479..4484,4485..4502,4503..4509)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4517..4533
                     /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     4522..4544
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4541..4557
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4541..4557
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4576..4596
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4578..4596
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4647..4996
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     primer_bind     4997..5013
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4997..5013
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             5031..6656
                     /codon_start=1
                     /product="green-emitting variant of click beetle 
                     luciferase"
                     /label=CBG99luc
                     /note="human codon-optimized"
                     /translation="MVKREKNVIYGPEPLHPLEDLTAGEMLFRALRKHSHLPQALVDVV
                     GDESLSYKEFFEATVLLAQSLHNCGYKMNDVVSICAENNTRFFIPVIAAWYIGMIVAPV
                     NESYIPDELCKVMGISKPQIVFTTKNILNKVLEVQSRTNFIKRIIILDTVENIHGCESL
                     PNFISRYSDGNIANFKPLHFDPVEQVAAILCSSGTTGLPKGVMQTHQNICVRLIHALDP
                     RVGTQLIPGVTVLVYLPFFHAFGFSITLGYFMVGLRVIMFRRFDQEAFLKAIQDYEVRS
                     VINVPSVILFLSKSPLVDKYDLSSLRELCCGAAPLAKEVAEVAAKRLNLPGIRCGFGLT
                     ESTSANIHSLGDEFKSGSLGRVTPLMAAKIADRETGKALGPNQVGELCIKGPMVSKGYV
                     NNVEATKEAIDDDGWLHSGDFGYYDEDEHFYVVDRYKELIKYKGSQVAPAELEEILLKN
                     PCIRDVAVVGIPDLEAGELPSAFVVKQPGKEITAKEVYDYLAERVSHTKYLRGGVRFVD
                     SIPRNVTGKITRKELLKQLLEKAGG"
     RBS             5981..5989
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     terminator      6689..6876
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(6744..6768)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
ORIGIN
        1 gatctgttta gcttgcctcg tccccgccgg gtcacccggc cagcgacatg gaggcccaga
       61 ataccctcct tgacagtctt gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt
      121 acatttagcc catacatccc catgtataat catttgcatc catacatttt gatggccgca
      181 cggcgcgaag caaaaattac ggctcctcgc tgcagacctg cgagcaggga aacgctcccc
      241 tcacagacgc gtgtacgcat gtaacattat actgaaaacc ttgcttgaga aggttttggg
      301 acgctcgaag gctttaattt gcggccggta cccaattcgc cctatagtga gtcgtattac
      361 gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
      421 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
      481 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcga cgcgccctgt
      541 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
      601 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
      661 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
      721 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
      781 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
      841 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
      901 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
      961 aacaaaatat taacgtttac aatttcctga tgcggtattt tctccttacg catctgtgcg
     1021 gtatttcaca ccgcataggg taataactga tataattaaa ttgaagctct aatttgtgag
     1081 tttagtatac atgcatttac ttataataca gttttttagt tttgctggcc gcatcttctc
     1141 aaatatgctt cccagcctgc ttttctgtaa cgttcaccct ctaccttagc atcccttccc
     1201 tttgcaaata gtcctcttcc aacaataata atgtcagatc ctgtagagac cacatcatcc
     1261 acggttctat actgttgacc caatgcgtct cccttgtcat ctaaacccac accgggtgtc
     1321 ataatcaacc aatcgtaacc ttcatctctt ccacccatgt ctctttgagc aataaagccg
     1381 ataacaaaat ctttgtcgct cttcgcaatg tcaacagtac ccttagtata ttctccagta
     1441 gatagggagc ccttgcatga caattctgct aacatcaaaa ggcctctagg ttcctttgtt
     1501 acttcttctg ccgcctgctt caaaccgcta acaatacctg ggcccaccac accgtgtgca
     1561 ttcgtaatgt ctgcccattc tgctattctg tatacacccg cagagtactg caatttgact
     1621 gtattaccaa tgtcagcaaa ttttctgtct tcgaagagta aaaaattgta cttggcggat
     1681 aatgccttta gcggcttaac tgtgccctcc atggaaaaat cagtcaagat atccacatgt
     1741 gtttttagta aacaaatttt gggacctaat gcttcaacta actccagtaa ttccttggtg
     1801 gtacgaacat ccaatgaagc acacaagttt gtttgctttt cgtgcatgat attaaatagc
     1861 ttggcagcaa caggactagg atgagtagca gcacgttcct tatatgtagc tttcgacatg
     1921 atttatcttc gtttcctgca ggtttttgtt ctgtgcagtt gggttaagaa tactgggcaa
     1981 tttcatgttt cttcaacact acatatgcgt atatatacca atctaagtct gtgctccttc
     2041 cttcgttctt ccttctgttc ggagattacc gaatcaaaaa aatttcaaag aaaccgaaat
     2101 caaaaaaaag aataaaaaaa aaatgatgaa ttgaattgaa aagctgtggt atggtgcact
     2161 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     2221 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     2281 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     2341 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     2401 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     2461 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     2521 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     2581 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     2641 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     2701 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     2761 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     2821 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     2881 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     2941 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     3001 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     3061 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     3121 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     3181 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     3241 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     3301 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     3361 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     3421 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     3481 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     3541 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     3601 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     3661 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     3721 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     3781 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     3841 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     3901 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     3961 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     4021 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     4081 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     4141 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     4201 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     4261 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     4321 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     4381 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     4441 attaatgtga gttacctcac tcattaggca ccccaggctt tacactttat gcttccggct
     4501 cctatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
     4561 gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctgga gctccggatg
     4621 caagggttcg aatcccttag ctctcattat tttttgcttt ttctcttgag gtcacatgat
     4681 cgcaaaatgg caaatggcac gtgaagctgt cgatattggg gaactgtggt ggttggcaaa
     4741 tgactaatta agttagtcaa ggcgccatcc tcatgaaaac tgtgtaacat aataaccgaa
     4801 gtgtcgaaaa ggtggcacct tgtccaattg aacacgctcg atgaaaaaaa taagatatat
     4861 ataaggttaa gtaaagcgtc tgttagaaag gaagtttttc ctttttcttg ctctcttgtc
     4921 ttttcatcta ctatttcctt cgtgtaatac agggtcgtca gatacataga tacaattcta
     4981 ttacccccat ccatactcta gaactagtgg atcccccggg ttaattaacc atggtgaagc
     5041 gtgagaaaaa tgtcatctat ggccctgagc ctctccatcc tttggaggat ttgactgccg
     5101 gcgaaatgct gtttcgtgct ctccgcaagc actctcattt gcctcaagcc ttggtcgatg
     5161 tggtcggcga tgaatctttg agctacaagg agttttttga ggcaaccgtc ttgctggctc
     5221 agtccctcca caattgtggc tacaagatga acgacgtcgt tagtatctgt gctgaaaaca
     5281 atacccgttt cttcattcca gtcatcgccg catggtatat cggtatgatc gtggctccag
     5341 tcaacgagag ctacattccc gacgaactgt gtaaagtcat gggtatctct aagccacaga
     5401 ttgtcttcac cactaagaat attctgaaca aagtcctgga agtccaaagc cgcaccaact
     5461 ttattaagcg tatcatcatc ttggacactg tggagaatat tcacggttgc gaatctttgc
     5521 ctaatttcat ctctcgctat tcagacggca acatcgcaaa ctttaaacca ctccacttcg
     5581 accctgtgga acaagttgca gccattctgt gtagcagcgg tactactgga ctcccaaagg
     5641 gagtcatgca gacccatcaa aacatttgcg tgcgtctgat ccatgctctc gatccacgcg
     5701 tgggcactca gctgattcct ggtgtcaccg tcttggtcta cttgcctttc ttccatgctt
     5761 tcggctttag cattactttg ggttacttta tggtcggtct ccgcgtgatt atgttccgcc
     5821 gttttgatca ggaggctttc ttgaaagcca tccaagatta tgaagtccgc agtgtcatca
     5881 acgtgcctag cgtgatcctg tttttgtcta agagcccact cgtggacaag tacgacttgt
     5941 cttcactgcg tgaattgtgt tgcggtgccg ctccactggc taaggaggtc gctgaagtgg
     6001 ccgccaaacg cttgaatctt ccagggattc gttgtggctt cggcctcacc gaatctacca
     6061 gcgctaacat tcactctctc ggggatgagt ttaagagcgg ctctttgggc cgtgtcactc
     6121 cactcatggc tgctaagatc gctgatcgcg aaactggtaa ggctttgggc ccgaaccaag
     6181 tgggcgagct gtgtatcaaa ggccctatgg tgagcaaggg ttatgtcaat aacgttgaag
     6241 ctaccaagga ggccatcgac gacgacggct ggttgcattc tggtgatttt ggatattacg
     6301 acgaagatga gcatttttac gtcgtggatc gttacaagga gctgatcaaa tacaagggta
     6361 gccaggttgc tccagctgag ttggaggaga ttctgttgaa aaatccatgc attcgcgatg
     6421 tcgctgtggt cggcattcct gatctggagg ccggcgaact gccttctgct ttcgttgtca
     6481 agcagcctgg taaagaaatt accgccaaag aagtgtatga ttacctggct gaacgtgtga
     6541 gccatactaa gtacttgcgt ggcggcgtgc gttttgttga ctccatccct cgtaacgtaa
     6601 caggcaaaat tacccgcaag gagctgttga aacaattgtt ggagaaggcc ggcggtttgt
     6661 acaaataagg cgcgccactt ctaaataagc gaatttctta tgatttatga tttttattat
     6721 taaataagtt ataaaaaaaa taagtgtata caaattttaa agtgactctt aggttttaaa
     6781 acgaaaattc ttattcttga gtaactcttt cctgtaggtc aggttgcttt ctcaggtata
     6841 gtatgaggtc gctcttattg accacacctc taccggcaga tccgctaggg ataacagggt
     6901 aatata
//
