LOCUS       sequence-386864-        8409 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MoClo Level 1 luciferase reporter: proNLP6::LucN::flag::notT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8409)
  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 8409)
  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"; doi: 
            "10.1021/acssynbio.3c00094"; journalName: "ACS Synth Biol"; date: 
            "2023-11-17- 17"; volume: "12"; issue: "11"; pages: "3482-3486"
COMMENT     Sequence Label: pEPSW1KN0020
FEATURES             Location/Qualifiers
     source          1..8409
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      314..508
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(744..766)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    852..992
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     907..926
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1007..1024)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1178..1766)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1258..1277)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1854..2669)
                     /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(1854..2669)
                     /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     2577..2596
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    3102..3126
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     CDS             4313..4825
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             4886..4909
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      4925..5177
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    5201..5225
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             6526..7155
                     /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(7584..8409,1..248)
                     /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 gccgttttct ctaccgcctg gcacgccgcg ccgcaggcaa ggcagaagcc agatggttgt
       61 tcaagacgat ctacgaacgc agtggcagcg ccggagagtt caagaagttc tgtttcaccg
      121 tgcgcaagct gatcgggtca aatgacctgc cggagtacga tttgaaggag gaggcggggc
      181 aggctggccc gatcctagtc atgcgctacc gcaacctgat cgagggcgaa gcatccgccg
      241 gttcctaatg tacggagcag atgctagggc aaattgccct agcaggggaa aaaggtcgaa
      301 aaggactctt tcctgtggat agcacgtaca ttgggaaccc aaagccgtac attgggaacc
      361 ggaacccgta cattgggaac ccaaagccgt acattgggaa ccggtcacac atgtaagtga
      421 ctgatataaa agagaaaaaa ggcgattttt ccgcctaaaa ctctttaaaa cttattaaaa
      481 ctcttaaaac ccgcctggcc tgtgcataac tgtctggcca gcgcacagcc gaagtgctgc
      541 aaaaagcgcc tacccttcgg tcgctgcgct ccctacgccc cgccgcttcg cgtcggccta
      601 tcgcggccgc tggccgctca aaaatggctg gcctacggcc aggcaatcta ccagggcgcg
      661 gacaagccgc gccgtcgcca ctcgaccgcc ggcgcccaca tcaaggcacc ctgcctcgcg
      721 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggtgac ggtcacagct
      781 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
      841 gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta tactggctta
      901 actatgcggc atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc
      961 acagatgcgt aaggagaaaa taccgcatca ggcgctcatc cgcttcctcg ctcactgact
     1021 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     1081 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     1141 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     1201 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     1261 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     1321 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     1381 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     1441 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     1501 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     1561 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga
     1621 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     1681 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     1741 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     1801 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gcattctagg tgattagaaa
     1861 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     1921 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     1981 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     2041 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     2101 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     2161 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     2221 gcgagtcgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     2281 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     2341 aatacctgga atgctgtttt ccctgggatc gcagtggtga gtaaccatgc atcatcagga
     2401 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     2461 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     2521 ggcgcatcgg gcttcccata caatcggtag attgtcgcac ctgattgccc gacattatcg
     2581 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggccttgag
     2641 caagacgttt cccgttgaat atggctcata acagaactta ttatttcctt cctcttttct
     2701 acagtattta aagatacccc aagaagctaa ttataacaag acgaactcca attcactgtt
     2761 ccttgcattc taaaacctta aataccagaa aacagctttt tcaaagttgt tttcaaagtt
     2821 ggcgtataac atagtatcga cggagccgat tttgaaaccg cggtgatcac aggcagcaac
     2881 gctctgtcat cgttacaatc aacatgctac cctccgcgag atcatccgtg tttcaaaccc
     2941 ggcagcttag ttgccgttct tccgaatagc atcggtaaca tgagcaaagt ctgccgcctt
     3001 acaacggctc tcccgctgac gccgtcccgg actgatgggc tgcctgtatc gagtggtgat
     3061 tttgtgccga gctgccggtc ggggagctgt tggctggctg gtggcaggat atattgtggt
     3121 gtaaacataa cgaattgctc ttcaatgagg agctcatcaa cttgaaacaa gtggggattt
     3181 acgttgcaag catttcttct ccttggggtt aaaatattga ttaacagcta aataacaaat
     3241 tagggaagtt attataacat taaaagaaag aataatcacg attagatgtc cccactagga
     3301 aaacaagtta ctgttaataa aaattttatg tattgcaaaa gggcttaagt aaggatccac
     3361 tttgggtaag tagttgaaaa tggctgagta agaataatca cgattagatg tcctcgcttt
     3421 tttattcatc tcccaaagcg agattcgttt ttttcattgt cgtcaattat tgcattgttt
     3481 atcataatta tttgctaaga acatcattat tattgaataa attatagtag tcgttgttat
     3541 tctactcacc cacaacacgc acttagtgcc taattacgtc ttaatttaat attttttaaa
     3601 gcttttaggc gcataaaata tgctatgaca actttgttca tggtggtgga cgtgtaagaa
     3661 acgtctaaag atctactcta tataacataa tcaattcgtt ttttgaaatg tgataaaaac
     3721 aaacctttac catatattcg cctctggcag aaacaatgca tttggaataa aatagaatcg
     3781 tatcacagca tttaaaaacg attattcaaa tagtctaata gtatttattc acacctttat
     3841 gttttatgta tatcgaattt tgtcaaagcc aatattctag agatggtcaa tcaatatcaa
     3901 tacagttttt ttttttggcc aatgttctaa tattcaagtt tgttagtatt aatagaaaaa
     3961 caaaatttcg atgccttttg tctcataaaa atattaaaga aaataataca aagaataaag
     4021 aatttgcatg aaatgggaga tattctagtg gttggggtaa aatggtacag cgtggaatca
     4081 aatgtcttca acaaaatcaa attattttat ctcaaaaaga tttaaatccc aatttaaata
     4141 atattacaac ttgtcttctc ctcttatttc ttctctctct ctttctgttg aagaagctga
     4201 atcagaagag tttttttttg cttcaaaagg agctccggcg aaacttccgg agatattaac
     4261 acttactctg acgagttttt gtctcgtctt acgtctttgc tgctgctgtt gaatggtatt
     4321 cacactcgaa gatttcgttg gggactggcg acagacagcc ggctacaacc tggaccaagt
     4381 ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc ggggtgtccg taactccgat
     4441 ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc gacatccatg tcatcatccc
     4501 gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa aaaattttta aggtggtgta
     4561 ccctgtggat gatcatcact ttaaggtgat cctgcactat ggcacactgg taatcgacgg
     4621 ggttacgccg aacatgatcg actatttcgg acggccgtat gaaggcatcg ccgtgttcga
     4681 cggcaaaaag atcactgtaa cagggaccct gtggaacggc aacaaaatta tcgacgagcg
     4741 cctgatcaac cccgacggct ccctgctgtt ccgagtaacc atcaacggag tgaccggctg
     4801 gcggctgtgc gaacgcattc tggcttcgga ttataaggac catgacggag actataagga
     4861 ccatgacctc gacgctgcag cagcggatta taaggacgat gacgataagt gagcttgtca
     4921 agcagatcgt tcaaacattt ggcaataaag tttcttaaga ttgaatcctg ttgccggtct
     4981 tgcgatgatt atcatataat ttctgttgaa ttacgttaag catgtaataa ttaacatgta
     5041 atgcatgacg ttatttatga gatgggtttt tatgattaga gtcccgcaat tatacattta
     5101 atacgcgata gaaaacaaaa tatagcgcgc aaactaggat aaattatcgc gcgcggtgtc
     5161 atctatgtta ctagatcgac gctagcatga agagcaattc tgacaggata tattggcggg
     5221 taaacctaag agaaaagagc gtttattaga ataatcggat atttaaaagg gcgtgaaaag
     5281 gtttatccgt tcgtccattt gtatgtgcat gccaaccaca gggttcccct cgggatcaaa
     5341 gtactttgat ccaacccctc cgctgctata gtgcagtcgg cttctgacgt tcagtgcagc
     5401 cgtcatctga aaacgacatg tcgcacaagt cctaagttac gcgacaggct gccgccctgc
     5461 ccttttcctg gcgttttctt gtcgcgtgtt ttagtcgcat aaagtagaat acttgcgact
     5521 agaaccggag acattacgcc atgaacaaga gcgccgccgc tggcctgctg ggctatgccc
     5581 gcgtcagcac cgacgaccag gacttgacca accaacgggc cgaactgcac gcggccggct
     5641 gcaccaagct gttttccgag aagatcaccg gcaccaggcg cgaccgcccg gagctggcca
     5701 ggatgcttga ccacctacgc cctggcgacg ttgtgacagt gaccaggcta gaccgcctgg
     5761 cccgcagcac ccgcgaccta ctggacattg ccgagcgcat ccaggaggcc ggcgcgggcc
     5821 tgcgtagcct ggcagagccg tgggccgaca ccaccacgcc ggccggccgc atggtgttga
     5881 ccgtgttcgc cggcattgcc gagttcgagc gttccctaat catcgaccgc acccggagcg
     5941 ggcgcgaggc cgccaaggcc cgaggcgtga agtttggccc ccgccctacc ctcaccccgg
     6001 cacagatcgc gcacgcccgc gagctgatcg accaggaagg ccgcaccgtg aaagaggcgg
     6061 ctgcactgct tggcgtgcat cgctcgaccc tgtaccgcgc acttgagcgc agcgaggaag
     6121 tgacgcccac cgaggccagg cggcgcggtg ccttccgtga ggacgcattg accgaggccg
     6181 acgccctggc ggccgccgag aatgaacgcc aagaggaaca agcatgaaac cgcaccagga
     6241 cggccaggac gaaccgtttt tcattaccga agagatcgag gcggagatga tcgcggccgg
     6301 gtacgtgttc gagccgcccg cgcacctctc aaccgtgcgg ctgcatgaaa tcctggccgg
     6361 tttgtctgat gccaagctgg cggcctggcc ggccagcttg gccgctgaag aaaccgagcg
     6421 ccgccgtcta aaaaggtgat gtgtatttga gtaaaacagc ttgcgtcatg cggtcgctgc
     6481 gtatatgatg cgatgagtaa ataaacaaat acgcaagggg aacgcatgaa ggttatcgct
     6541 gtacttaacc agaaaggcgg gtcaggcaag acgaccatcg caacccatct agcccgcgcc
     6601 ctgcaactcg ccggggccga tgttctgtta gtcgattccg atccccaggg cagtgcccgc
     6661 gattgggcgg ccgtgcggga agatcaaccg ctaaccgttg tcggcatcga ccgcccgacg
     6721 attgaccgcg acgtgaaggc catcggccgg cgcgacttcg tagtgatcga cggagcgccc
     6781 caggcggcgg acttggctgt gtccgcgatc aaggcagccg acttcgtgct gattccggtg
     6841 cagccaagcc cttacgacat atgggccacc gccgacctgg tggagctggt taagcagcgc
     6901 attgaggtca cggatggaag gctacaagcg gcctttgtcg tgtcgcgggc gatcaaaggc
     6961 acgcgcatcg gcggtgaggt tgccgaggcg ctggccgggt acgagctgcc cattcttgag
     7021 tcccgtatca cgcagcgcgt gagctaccca ggcactgccg ccgccggcac aaccgttctt
     7081 gaatcagaac ccgagggcga cgctgcccgc gaggtccagg cgctggccgc tgaaattaaa
     7141 tcaaaactca tttgagttaa tgaggtaaag agaaaatgag caaaagcaca aacacgctaa
     7201 gtgccggccg tccgagcgca cgcagcagca aggctgcaac gttggccagc ctggcagaca
     7261 cgccagccat gaagcgggtc aactttcagt tgccggcgga ggatcacacc aagctgaaga
     7321 tgtacgcggt acgccaaggc aagaccatta ccgagctgct atctgaatac atcgcgcagc
     7381 taccagagta aatgagcaaa tgaataaatg agtagatgaa ttttagcggc taaaggaggc
     7441 ggcatggaaa atcaagaaca accaggcacc gacgccgtgg aatgccccat gtgtggagga
     7501 acgggcggtt ggccaggcgt aagcggctgg gttgcctgcc ggccctgcaa tggcactgga
     7561 acccccaagc ccgaggaatc ggcgtgagcg gtcgcaaacc atccggcccg gtacaaatcg
     7621 gcgcggcgct gggtgatgac ctggtggaga agttgaaggc cgcgcaggcc gcccagcggc
     7681 aacgcatcga ggcagaagca cgccccggtg aatcgtggca agcggccgct gatcgaatcc
     7741 gcaaagaatc ccggcaaccg ccggcagccg gtgcgccgtc gattaggaag ccgcccaagg
     7801 gcgacgagca accagatttt ttcgttccga tgctctatga cgtgggcacc cgcgatagtc
     7861 gcagcatcat ggacgtggcc gttttccgtc tgtcgaagcg tgaccgacga gctggcgagg
     7921 tgatccgcta cgagcttcca gacgggcacg tagaggtttc cgcagggccg gccggcatgg
     7981 ccagtgtgtg ggattacgac ctggtactga tggcggtttc ccatctaacc gaatccatga
     8041 accgataccg ggaagggaag ggagacaagc ccggccgcgt gttccgtcca cacgttgcgg
     8101 acgtactcaa gttctgccgg cgagccgatg gcggaaagca gaaagacgac ctggtagaaa
     8161 cctgcattcg gttaaacacc acgcacgttg ccatgcagcg tacgaagaag gccaagaacg
     8221 gccgcctggt gacggtatcc gagggtgaag ccttgattag ccgctacaag atcgtaaaga
     8281 gcgaaaccgg gcggccggag tacatcgaga tcgagctagc tgattggatg taccgcgaga
     8341 tcacagaagg caagaacccg gacgtgctga cggttcaccc cgattacttt ttgatcgatc
     8401 ccggcatcg
//
