LOCUS       sequence-387007-        8533 bp    DNA     circular SYN 15-APR-2025
DEFINITION  MoClo Level 1 luciferase reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8533)
  AUTHORS   Cai YM, Witham S, Patron NJ
  TITLE     Tuning Plant Promoters Using a Simple Split Luciferase Method to 
            Assess Transcription Factor-DNA Interactions.
  JOURNAL   ACS Synth Biol. 2023 Nov 17;12(11):3482-3486. doi: 
            10.1021/acssynbio.3c00094. Epub 2023 Oct 19.
  PUBMED    37856867
REFERENCE   2  (bases 1 to 8533)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acssynbio.3c00094"; journalName: "ACS Synth Biol"; date: 
            "2023-11-17- 17"; volume: "12"; issue: "11"; pages: "3482-3486"
COMMENT     Sequence Label: pEPSW1KN0016
FEATURES             Location/Qualifiers
     source          1..8533
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      313..507
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(743..765)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    851..991
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     906..925
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1006..1023)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1177..1765)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1257..1276)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1853..2668)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1853..2668)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2576..2595
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    3101..3125
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     CDS             4436..4948
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             5009..5032
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      5048..5300
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    5324..5348
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             6649..7278
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             join(7707..8533,1..247)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="VSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
ORIGIN
        1 ccgttttctc taccgcctgg cacgccgcgc cgcaggcaag gcagaagcca gatggttgtt
       61 caagacgatc tacgaacgca gtggcagcgc cggagagttc aagaagttct gtttcaccgt
      121 gcgcaagctg atcgggtcaa atgacctgcc ggagtacgat ttgaaggagg aggcggggca
      181 ggctggcccg atcctagtca tgcgctaccg caacctgatc gagggcgaag catccgccgg
      241 ttcctaatgt acggagcaga tgctagggca aattgcccta gcaggggaaa aaggtcgaaa
      301 aggactcttt cctgtggata gcacgtacat tgggaaccca aagccgtaca ttgggaaccg
      361 gaacccgtac attgggaacc caaagccgta cattgggaac cggtcacaca tgtaagtgac
      421 tgatataaaa gagaaaaaag gcgatttttc cgcctaaaac tctttaaaac ttattaaaac
      481 tcttaaaacc cgcctggcct gtgcataact gtctggccag cgcacagccg aagtgctgca
      541 aaaagcgcct acccttcggt cgctgcgctc cctacgcccc gccgcttcgc gtcggcctat
      601 cgcggccgct ggccgctcaa aaatggctgg cctacggcca ggcaatctac cagggcgcgg
      661 acaagccgcg ccgtcgccac tcgaccgccg gcgcccacat caaggcaccc tgcctcgcgc
      721 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggtgacg gtcacagctt
      781 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
      841 ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa
      901 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
      961 cagatgcgta aggagaaaat accgcatcag gcgctcatcc gcttcctcgc tcactgactc
     1021 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     1081 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     1141 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     1201 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     1261 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     1321 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     1381 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     1441 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     1501 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     1561 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac
     1621 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     1681 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     1741 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     1801 tcagtggaac gaaaactcac gttaagggat tttggtcatg cattctaggt gattagaaaa
     1861 actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt
     1921 tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg
     1981 caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt
     2041 tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg
     2101 gtgagaatgg caaaagttta tgcatttctt tccagacttg ttcaacaggc cagccattac
     2161 gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag
     2221 cgagtcgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc
     2281 ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta
     2341 atacctggaa tgctgttttc cctgggatcg cagtggtgag taaccatgca tcatcaggag
     2401 tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga
     2461 ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg
     2521 gcgcatcggg cttcccatac aatcggtaga ttgtcgcacc tgattgcccg acattatcgc
     2581 gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc ggccttgagc
     2641 aagacgtttc ccgttgaata tggctcataa cagaacttat tatttccttc ctcttttcta
     2701 cagtatttaa agatacccca agaagctaat tataacaaga cgaactccaa ttcactgttc
     2761 cttgcattct aaaaccttaa ataccagaaa acagcttttt caaagttgtt ttcaaagttg
     2821 gcgtataaca tagtatcgac ggagccgatt ttgaaaccgc ggtgatcaca ggcagcaacg
     2881 ctctgtcatc gttacaatca acatgctacc ctccgcgaga tcatccgtgt ttcaaacccg
     2941 gcagcttagt tgccgttctt ccgaatagca tcggtaacat gagcaaagtc tgccgcctta
     3001 caacggctct cccgctgacg ccgtcccgga ctgatgggct gcctgtatcg agtggtgatt
     3061 ttgtgccgag ctgccggtcg gggagctgtt ggctggctgg tggcaggata tattgtggtg
     3121 taaacataac gaattgctct tcaatgagga gaaatattta tacatactca atgtaataag
     3181 tgaaataaaa attcattcaa gatcaagaaa aattaaaata tgaatggaga atccaatacc
     3241 aactagatta aagaacaagg ttgtaaaacg attcatgagt agtgtcacat tcatgaaatt
     3301 aatatataaa attttacatt gacagaaaaa atataaaaat taaaaaacaa taaggaattt
     3361 aaaaaatata tactcaacat gtaatatata tatgtaaaat aaaatcacac taaaatggta
     3421 tcctcgattg tatagaaact tccaaatttc taaacggatg caataagaac ttacatattc
     3481 tctttcatta gtcatttatt ggccagattt attaaaaaaa gttttactca atgaccaagg
     3541 attagagtta aagataatat agattattac atatattatt cgaaaaaata tacgcatgtc
     3601 cgacttttta aacctcaaaa atatcaaaac cagaaaagat gataccacac aaaaaaacaa
     3661 taaaataata agtggaagag atatcatcgg acaacagtac aagtacagca ccagctctgc
     3721 caaaagccaa aaccatttgt caattacaga aagatactat tgtttgcaat tactaaatta
     3781 cccctcggac tttacaaaag catctctaac ttatccacgt gtcagtcatc tattgattgt
     3841 ttcaatacca ccttgtatta acgccccacg attcgtggtt gggtacacct gatagtccga
     3901 ggatatttaa atctcacgcg ctcgtgtcta taattcgact gtactcgctt ttcttgtcgt
     3961 gattttagca atttacgaag tcaaatgttt gactcaatca gacttgcgca taagagagcg
     4021 agtataaatg tttactatac tcacgcaagt ggggctttat tgaaactact cttttgtaat
     4081 aaaaccagca gtggttttgt tctgaatccg ctctcttgcc atatatacca caaacagaaa
     4141 ccacagaaga tatcttttga gaaggaaaaa aaaaaagaag cttctcctct tcctctgcct
     4201 tcttctttcc atttattgca aaccctgatc aagtaagtca aatcttcacg aacacatatg
     4261 tatataaatt caatccaaga aactaggaga aatctatgaa agaggacaaa tctaagtcaa
     4321 gtttgaatca ggaagattat ctagatttga tcattttgac atttacgatg tgcttactta
     4381 ttcttgataa actttgatgc agttggtttt ggtgttagtc ttttggggag agagaatggt
     4441 attcacactc gaagatttcg ttggggactg gcgacagaca gccggctaca acctggacca
     4501 agtccttgaa cagggaggtg tgtccagttt gtttcagaat ctcggggtgt ccgtaactcc
     4561 gatccaaagg attgtcctga gcggtgaaaa tgggctgaag atcgacatcc atgtcatcat
     4621 cccgtatgaa ggtctgagcg gcgaccaaat gggccagatc gaaaaaattt ttaaggtggt
     4681 gtaccctgtg gatgatcatc actttaaggt gatcctgcac tatggcacac tggtaatcga
     4741 cggggttacg ccgaacatga tcgactattt cggacggccg tatgaaggca tcgccgtgtt
     4801 cgacggcaaa aagatcactg taacagggac cctgtggaac ggcaacaaaa ttatcgacga
     4861 gcgcctgatc aaccccgacg gctccctgct gttccgagta accatcaacg gagtgaccgg
     4921 ctggcggctg tgcgaacgca ttctggcttc ggattataag gaccatgacg gagactataa
     4981 ggaccatgac ctcgacgctg cagcagcgga ttataaggac gatgacgata agtgagcttg
     5041 tcaagcagat cgttcaaaca tttggcaata aagtttctta agattgaatc ctgttgccgg
     5101 tcttgcgatg attatcatat aatttctgtt gaattacgtt aagcatgtaa taattaacat
     5161 gtaatgcatg acgttattta tgagatgggt ttttatgatt agagtcccgc aattatacat
     5221 ttaatacgcg atagaaaaca aaatatagcg cgcaaactag gataaattat cgcgcgcggt
     5281 gtcatctatg ttactagatc gacgctagca tgaagagcaa ttctgacagg atatattggc
     5341 gggtaaacct aagagaaaag agcgtttatt agaataatcg gatatttaaa agggcgtgaa
     5401 aaggtttatc cgttcgtcca tttgtatgtg catgccaacc acagggttcc cctcgggatc
     5461 aaagtacttt gatccaaccc ctccgctgct atagtgcagt cggcttctga cgttcagtgc
     5521 agccgtcatc tgaaaacgac atgtcgcaca agtcctaagt tacgcgacag gctgccgccc
     5581 tgcccttttc ctggcgtttt cttgtcgcgt gttttagtcg cataaagtag aatacttgcg
     5641 actagaaccg gagacattac gccatgaaca agagcgccgc cgctggcctg ctgggctatg
     5701 cccgcgtcag caccgacgac caggacttga ccaaccaacg ggccgaactg cacgcggccg
     5761 gctgcaccaa gctgttttcc gagaagatca ccggcaccag gcgcgaccgc ccggagctgg
     5821 ccaggatgct tgaccaccta cgccctggcg acgttgtgac agtgaccagg ctagaccgcc
     5881 tggcccgcag cacccgcgac ctactggaca ttgccgagcg catccaggag gccggcgcgg
     5941 gcctgcgtag cctggcagag ccgtgggccg acaccaccac gccggccggc cgcatggtgt
     6001 tgaccgtgtt cgccggcatt gccgagttcg agcgttccct aatcatcgac cgcacccgga
     6061 gcgggcgcga ggccgccaag gcccgaggcg tgaagtttgg cccccgccct accctcaccc
     6121 cggcacagat cgcgcacgcc cgcgagctga tcgaccagga aggccgcacc gtgaaagagg
     6181 cggctgcact gcttggcgtg catcgctcga ccctgtaccg cgcacttgag cgcagcgagg
     6241 aagtgacgcc caccgaggcc aggcggcgcg gtgccttccg tgaggacgca ttgaccgagg
     6301 ccgacgccct ggcggccgcc gagaatgaac gccaagagga acaagcatga aaccgcacca
     6361 ggacggccag gacgaaccgt ttttcattac cgaagagatc gaggcggaga tgatcgcggc
     6421 cgggtacgtg ttcgagccgc ccgcgcacct ctcaaccgtg cggctgcatg aaatcctggc
     6481 cggtttgtct gatgccaagc tggcggcctg gccggccagc ttggccgctg aagaaaccga
     6541 gcgccgccgt ctaaaaaggt gatgtgtatt tgagtaaaac agcttgcgtc atgcggtcgc
     6601 tgcgtatatg atgcgatgag taaataaaca aatacgcaag gggaacgcat gaaggttatc
     6661 gctgtactta accagaaagg cgggtcaggc aagacgacca tcgcaaccca tctagcccgc
     6721 gccctgcaac tcgccggggc cgatgttctg ttagtcgatt ccgatcccca gggcagtgcc
     6781 cgcgattggg cggccgtgcg ggaagatcaa ccgctaaccg ttgtcggcat cgaccgcccg
     6841 acgattgacc gcgacgtgaa ggccatcggc cggcgcgact tcgtagtgat cgacggagcg
     6901 ccccaggcgg cggacttggc tgtgtccgcg atcaaggcag ccgacttcgt gctgattccg
     6961 gtgcagccaa gcccttacga catatgggcc accgccgacc tggtggagct ggttaagcag
     7021 cgcattgagg tcacggatgg aaggctacaa gcggcctttg tcgtgtcgcg ggcgatcaaa
     7081 ggcacgcgca tcggcggtga ggttgccgag gcgctggccg ggtacgagct gcccattctt
     7141 gagtcccgta tcacgcagcg cgtgagctac ccaggcactg ccgccgccgg cacaaccgtt
     7201 cttgaatcag aacccgaggg cgacgctgcc cgcgaggtcc aggcgctggc cgctgaaatt
     7261 aaatcaaaac tcatttgagt taatgaggta aagagaaaat gagcaaaagc acaaacacgc
     7321 taagtgccgg ccgtccgagc gcacgcagca gcaaggctgc aacgttggcc agcctggcag
     7381 acacgccagc catgaagcgg gtcaactttc agttgccggc ggaggatcac accaagctga
     7441 agatgtacgc ggtacgccaa ggcaagacca ttaccgagct gctatctgaa tacatcgcgc
     7501 agctaccaga gtaaatgagc aaatgaataa atgagtagat gaattttagc ggctaaagga
     7561 ggcggcatgg aaaatcaaga acaaccaggc accgacgccg tggaatgccc catgtgtgga
     7621 ggaacgggcg gttggccagg cgtaagcggc tgggttgcct gccggccctg caatggcact
     7681 ggaaccccca agcccgagga atcggcgtga gcggtcgcaa accatccggc ccggtacaaa
     7741 tcggcgcggc gctgggtgat gacctggtgg agaagttgaa ggccgcgcag gccgcccagc
     7801 ggcaacgcat cgaggcagaa gcacgccccg gtgaatcgtg gcaagcggcc gctgatcgaa
     7861 tccgcaaaga atcccggcaa ccgccggcag ccggtgcgcc gtcgattagg aagccgccca
     7921 agggcgacga gcaaccagat tttttcgttc cgatgctcta tgacgtgggc acccgcgata
     7981 gtcgcagcat catggacgtg gccgttttcc gtctgtcgaa gcgtgaccga cgagctggcg
     8041 aggtgatccg ctacgagctt ccagacgggc acgtagaggt ttccgcaggg ccggccggca
     8101 tggccagtgt gtgggattac gacctggtac tgatggcggt ttcccatcta accgaatcca
     8161 tgaaccgata ccgggaaggg aagggagaca agcccggccg cgtgttccgt ccacacgttg
     8221 cggacgtact caagttctgc cggcgagccg atggcggaaa gcagaaagac gacctggtag
     8281 aaacctgcat tcggttaaac accacgcacg ttgccatgca gcgtacgaag aaggccaaga
     8341 acggccgcct ggtgacggta tccgagggtg aagccttgat tagccgctac aagatcgtaa
     8401 agagcgaaac cgggcggccg gagtacatcg agatcgagct agctgattgg atgtaccgcg
     8461 agatcacaga aggcaagaac ccggacgtgc tgacggttca ccccgattac tttttgatcg
     8521 atcccggcat cgg
//
