LOCUS       sequence-387003-        8912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 gene expression cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8912)
  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 8912)
  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: pEPSW1KN0032
FEATURES             Location/Qualifiers
     source          1..8912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    18..42
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             1343..1972
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             2401..3474
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKEXRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      3540..3734
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(3970..3992)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4078..4218
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4133..4152
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4233..4250)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4404..4992)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4484..4503)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5080..5895)
                     /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(5080..5895)
                     /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     5803..5822
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    6328..6352
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     promoter        6456..6801
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     6744..6764
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     polyA_signal    8735..8909
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
ORIGIN
        1 atactgaaga gcaattctga caggatatat tggcgggtaa acctaagaga aaagagcgtt
       61 tattagaata atcggatatt taaaagggcg tgaaaaggtt tatccgttcg tccatttgta
      121 tgtgcatgcc aaccacaggg ttcccctcgg gatcaaagta ctttgatcca acccctccgc
      181 tgctatagtg cagtcggctt ctgacgttca gtgcagccgt catctgaaaa cgacatgtcg
      241 cacaagtcct aagttacgcg acaggctgcc gccctgccct tttcctggcg ttttcttgtc
      301 gcgtgtttta gtcgcataaa gtagaatact tgcgactaga accggagaca ttacgccatg
      361 aacaagagcg ccgccgctgg cctgctgggc tatgcccgcg tcagcaccga cgaccaggac
      421 ttgaccaacc aacgggccga actgcacgcg gccggctgca ccaagctgtt ttccgagaag
      481 atcaccggca ccaggcgcga ccgcccggag ctggccagga tgcttgacca cctacgccct
      541 ggcgacgttg tgacagtgac caggctagac cgcctggccc gcagcacccg cgacctactg
      601 gacattgccg agcgcatcca ggaggccggc gcgggcctgc gtagcctggc agagccgtgg
      661 gccgacacca ccacgccggc cggccgcatg gtgttgaccg tgttcgccgg cattgccgag
      721 ttcgagcgtt ccctaatcat cgaccgcacc cggagcgggc gcgaggccgc caaggcccga
      781 ggcgtgaagt ttggcccccg ccctaccctc accccggcac agatcgcgca cgcccgcgag
      841 ctgatcgacc aggaaggccg caccgtgaaa gaggcggctg cactgcttgg cgtgcatcgc
      901 tcgaccctgt accgcgcact tgagcgcagc gaggaagtga cgcccaccga ggccaggcgg
      961 cgcggtgcct tccgtgagga cgcattgacc gaggccgacg ccctggcggc cgccgagaat
     1021 gaacgccaag aggaacaagc atgaaaccgc accaggacgg ccaggacgaa ccgtttttca
     1081 ttaccgaaga gatcgaggcg gagatgatcg cggccgggta cgtgttcgag ccgcccgcgc
     1141 acctctcaac cgtgcggctg catgaaatcc tggccggttt gtctgatgcc aagctggcgg
     1201 cctggccggc cagcttggcc gctgaagaaa ccgagcgccg ccgtctaaaa aggtgatgtg
     1261 tatttgagta aaacagcttg cgtcatgcgg tcgctgcgta tatgatgcga tgagtaaata
     1321 aacaaatacg caaggggaac gcatgaaggt tatcgctgta cttaaccaga aaggcgggtc
     1381 aggcaagacg accatcgcaa cccatctagc ccgcgccctg caactcgccg gggccgatgt
     1441 tctgttagtc gattccgatc cccagggcag tgcccgcgat tgggcggccg tgcgggaaga
     1501 tcaaccgcta accgttgtcg gcatcgaccg cccgacgatt gaccgcgacg tgaaggccat
     1561 cggccggcgc gacttcgtag tgatcgacgg agcgccccag gcggcggact tggctgtgtc
     1621 cgcgatcaag gcagccgact tcgtgctgat tccggtgcag ccaagccctt acgacatatg
     1681 ggccaccgcc gacctggtgg agctggttaa gcagcgcatt gaggtcacgg atggaaggct
     1741 acaagcggcc tttgtcgtgt cgcgggcgat caaaggcacg cgcatcggcg gtgaggttgc
     1801 cgaggcgctg gccgggtacg agctgcccat tcttgagtcc cgtatcacgc agcgcgtgag
     1861 ctacccaggc actgccgccg ccggcacaac cgttcttgaa tcagaacccg agggcgacgc
     1921 tgcccgcgag gtccaggcgc tggccgctga aattaaatca aaactcattt gagttaatga
     1981 ggtaaagaga aaatgagcaa aagcacaaac acgctaagtg ccggccgtcc gagcgcacgc
     2041 agcagcaagg ctgcaacgtt ggccagcctg gcagacacgc cagccatgaa gcgggtcaac
     2101 tttcagttgc cggcggagga tcacaccaag ctgaagatgt acgcggtacg ccaaggcaag
     2161 accattaccg agctgctatc tgaatacatc gcgcagctac cagagtaaat gagcaaatga
     2221 ataaatgagt agatgaattt tagcggctaa aggaggcggc atggaaaatc aagaacaacc
     2281 aggcaccgac gccgtggaat gccccatgtg tggaggaacg ggcggttggc caggcgtaag
     2341 cggctgggtt gcctgccggc cctgcaatgg cactggaacc cccaagcccg aggaatcggc
     2401 gtgagcggtc gcaaaccatc cggcccggta caaatcggcg cggcgctggg tgatgacctg
     2461 gtggagaagt tgaaggccgc gcaggccgcc cagcggcaac gcatcgaggc agaagcacgc
     2521 cccggtgaat cgtggcaagc ggccgctgat cgaatccgca aagaaycccg gcaaccgccg
     2581 gcagccggtg cgccgtcgat taggaagccg cccaagggcg acgagcaacc agattttttc
     2641 gttccgatgc tctatgacgt gggcacccgc gatagtcgca gcatcatgga cgtggccgtt
     2701 ttccgtctgt cgaagcgtga ccgacgagct ggcgaggtga tccgctacga gcttccagac
     2761 gggcacgtag aggtttccgc agggccggcc ggcatggcca gtgtgtggga ttacgacctg
     2821 gtactgatgg cggtttccca tctaaccgaa tccatgaacc gataccggga agggaaggga
     2881 gacaagcccg gccgcgtgtt ccgtccacac gttgcggacg tactcaagtt ctgccggcga
     2941 gccgatggcg gaaagcagaa agacgacctg gtagaaacct gcattcggtt aaacaccacg
     3001 cacgttgcca tgcagcgtac gaagaaggcc aagaacggcc gcctggtgac ggtatccgag
     3061 ggtgaagcct tgattagccg ctacaagatc gtaaagagcg aaaccgggcg gccggagtac
     3121 atcgagatcg agctagctga ttggatgtac cgcgagatca cagaaggcaa gaacccggac
     3181 gtgctgacgg ttcaccccga ttactttttg atcgatcccg gcatcggccg ttttctctac
     3241 cgcctggcac gccgcgccgc aggcaaggca gaagccagat ggttgttcaa gacgatctac
     3301 gaacgcagtg gcagcgccgg agagttcaag aagttctgtt tcaccgtgcg caagctgatc
     3361 gggtcaaatg acctgccgga gtacgatttg aaggaggagg cggggcaggc tggcccgatc
     3421 ctagtcatgc gctaccgcaa cctgatcgag ggcgaagcat ccgccggttc ctaatgtacg
     3481 gagcagatgc tagggcaaat tgccctagca ggggaaaaag gtcgaaaagg actctttcct
     3541 gtggatagca cgtacattgg gaacccaaag ccgtacattg ggaaccggaa cccgtacatt
     3601 gggaacccaa agccgtacat tgggaaccgg tcacacatgt aagtgactga tataaaagag
     3661 aaaaaaggcg atttttccgc ctaaaactct ttaaaactta ttaaaactct taaaacccgc
     3721 ctggcctgtg cataactgtc tggccagcgc acagccgaag tgctgcaaaa agcgcctacc
     3781 cttcggtcgc tgcgctccct acgccccgcc gcttcgcgtc ggcctatcgc ggccgctggc
     3841 cgctcaaaaa tggctggcct acggccaggc aatctaccag ggcgcggaca agccgcgccg
     3901 tcgccactcg accgccggcg cccacatcaa ggcaccctgc ctcgcgcgtt tcggtgatga
     3961 cggtgaaaac ctctgacaca tgcagctccc ggtgacggtc acagcttgtc tgtaagcgga
     4021 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc
     4081 agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca
     4141 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     4201 agaaaatacc gcatcaggcg ctcatccgct tcctcgctca ctgactcgct gcgctcggtc
     4261 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4321 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4381 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4441 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4501 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     4561 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     4621 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4681 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4741 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     4801 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     4861 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     4921 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4981 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5041 aactcacgtt aagggatttt ggtcatgcat tctaggtgat tagaaaaact catcgagcat
     5101 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
     5161 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
     5221 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
     5281 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
     5341 aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa
     5401 atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gtcgaaatac
     5461 gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac
     5521 tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc
     5581 tgttttccct gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg
     5641 cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt
     5701 aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt
     5761 cccatacaat cggtagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata
     5821 cccatataaa tcagcatcca tgttggaatt taatcgcggc cttgagcaag acgtttcccg
     5881 ttgaatatgg ctcataacag aacttattat ttccttcctc ttttctacag tatttaaaga
     5941 taccccaaga agctaattat aacaagacga actccaattc actgttcctt gcattctaaa
     6001 accttaaata ccagaaaaca gctttttcaa agttgttttc aaagttggcg tataacatag
     6061 tatcgacgga gccgattttg aaaccgcggt gatcacaggc agcaacgctc tgtcatcgtt
     6121 acaatcaaca tgctaccctc cgcgagatca tccgtgtttc aaacccggca gcttagttgc
     6181 cgttcttccg aatagcatcg gtaacatgag caaagtctgc cgccttacaa cggctctccc
     6241 gctgacgccg tcccggactg atgggctgcc tgtatcgagt ggtgattttg tgccgagctg
     6301 ccggtcgggg agctgttggc tggctggtgg caggatatat tgtggtgtaa acataacgaa
     6361 ttgctcttca gcaaggaggt caacatggtg gagcacgaca ctctggtcta ctccaaaaat
     6421 gtcaaagata cagtctcaga agatcaaagg gctattgaga cttttcaaca aaggataatt
     6481 tcgggaaacc tcctcggatt ccattgccca gctatctgtc acttcatcga aaggacagta
     6541 gaaaaggaag gtggctccta caaatgccat cattgcgata aaggaaaggc tatcattcaa
     6601 gatctctctg ccgacagtgg tcccaaagat ggacccccac ccacgaggag catcgtggaa
     6661 aaagaagagg ttccaaccac gtctacaaag caagtggatt gatgtgacat ctccactgac
     6721 gtaagggatg acgcacaatc ccactatcct tcgcaagacc cttcctctat ataaggaagt
     6781 tcatttcatt tggagaggac acgctcgagt ataagagctc atttttacaa caattaccaa
     6841 caacaacaaa caacaaacaa cattacaatt acatttacaa ttatcgatac aatggcgagt
     6901 gttgaaggtg atgatgattt cggaagttct tcgtcaaggt cttatcaaga tcaactatac
     6961 acagagctat ggaaagtttg tgcaggtcca ttagtggaag ttcctcgtgc tcaagagaga
     7021 gttttctact tccctcaggg tcacatggaa caacttgtgg cgtcaactaa tcaaggaatc
     7081 aattcagaag aaatacctgt ttttgatctt cctccaaaga tactttgtcg agttcttgat
     7141 gtcactttaa aggcggagca tgaaacagat gaggtttacg ctcagatcac attacaacca
     7201 gaggaagatc aaagtgaacc aacaagtctt gatccaccta ttgttggacc aactaagcaa
     7261 gagtttcatt cgtttgttaa gattttaacg gcttcagata caagcactca tggtggattc
     7321 tctgttcttc gtaaacacgc cactgaatgc ttgccttctt tggatatgac acaagctact
     7381 cctactcaag aacttgtgac tagagatctt catggctttg aatggaggtt taagcatata
     7441 ttcagaggac aaccacggag gcatttgctt actacgggtt ggagtacatt tgtatcctcg
     7501 aaaagacttg tagctggaga tgcttttgtg ttcttgaggg gtgagaatgg ggatttacgg
     7561 gttggagtga gacgattagc tcggcatcaa agcacaatgc ctacttcggt tatttcaagt
     7621 cagagcatgc atttgggagt tcttgctaca gcttctcatg ctgtgcgtac aacaacaatc
     7681 tttgttgtct tttacaagcc taggataagc caattcatag ttggggtgaa caagtatatg
     7741 gaagctataa agcatggatt ttctctcggt acccgattca gaatgaggtt tgaaggagaa
     7801 gagtctcctg agagaatatt tactggtacg attgtgggaa gtggagatct atcttcacaa
     7861 tggccagctt ctaaatggag gtcattgcag gtacaatggg atgagccaac aacagttcaa
     7921 agaccagata aagtctcacc atgggagata gagcctttct tggcaacttc cccaatttca
     7981 actcctgctc aacaaccaca atcgaaatgc aagcggtcaa gacccatcga gccatcagtt
     8041 aaaacaccag ccccacctag tttcttgtac agcctccctc agagccaaga ttccattaat
     8101 gcatccctta aactgtttca agatccatca cttgagagaa tttcaggtgg atactcctca
     8161 aacaacagct tcaaacccga gactcctcct cctccaacga attgtagcta taggttgttt
     8221 ggatttgatc tcacaagcaa ttctcctgct ccaatccctc aagacaagca accgatggat
     8281 acttgtggag ctgccaagtg tcaagaaccc atcactccaa cctcaatgag tgagcagaag
     8341 aagcaacaaa catcaagaag tcgaactaaa gtgcaaatgc aaggcattgc ggttggtcgt
     8401 gcggttgatt taacactgtt gaaatcttac gatgaactga ttgatgagct tgaggagatg
     8461 tttgagattc aaggacagct tcttgcccga gacaaatgga tcgttgtctt cactgatgat
     8521 gaaggagata tgatgcttgc tggtgatgat ccgtggaatg agttttgcaa gatggcaaag
     8581 aagatattta tatattcgag cgatgaggtt aagaaaatga caacgaaact gaagatttct
     8641 tcgtcgttag agaatgagga atatggtaat gaatcattcg aaaatcgtag tagggggtga
     8701 gcttctctag ctagagtcga tcgacaagct cgagtttctc cataataatg tgtgagtagt
     8761 tcccagataa gggaattagg gttcctatag ggtttcgctc atgtgttgag catataagaa
     8821 acccttagta tgtatttgta tttgtaaaat acttctatca ataaaatttc taattcctaa
     8881 aaccaaaatc cagtactaaa atccagatcg ct
//
