LOCUS       sequence-386980-        8984 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 8984)
  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 8984)
  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: pEPOZ1KN0144
FEATURES             Location/Qualifiers
     source          1..8984
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             477..1550
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      1616..1810
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(2046..2068)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2154..2294
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2209..2228
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2309..2326)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2480..3068)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2560..2579)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3156..3971)
                     /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(3156..3971)
                     /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     3879..3898
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    4404..4428
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     promoter        4859..5204
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     5147..5167
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     polyA_signal    6883..7057
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     misc_feature    7078..7102
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             join(8403..8984,1..48)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
ORIGIN
        1 cgcgaggtcc aggcgctggc cgctgaaatt aaatcaaaac tcatttgagt taatgaggta
       61 aagagaaaat gagcaaaagc acaaacacgc taagtgccgg ccgtccgagc gcacgcagca
      121 gcaaggctgc aacgttggcc agcctggcag acacgccagc catgaagcgg gtcaactttc
      181 agttgccggc ggaggatcac accaagctga agatgtacgc ggtacgccaa ggcaagacca
      241 ttaccgagct gctatctgaa tacatcgcgc agctaccaga gtaaatgagc aaatgaataa
      301 atgagtagat gaattttagc ggctaaagga ggcggcatgg aaaatcaaga acaaccaggc
      361 accgacgccg tggaatgccc catgtgtgga ggaacgggcg gttggccagg cgtaagcggc
      421 tgggttgcct gccggccctg caatggcact ggaaccccca agcccgagga atcggcgtga
      481 gcggtcgcaa accatccggc ccggtacaaa tcggcgcggc gctgggtgat gacctggtgg
      541 agaagttgaa ggccgcgcag gccgcccagc ggcaacgcat cgaggcagaa gcacgccccg
      601 gtgaatcgtg gcaagcggcc gctgatcgaa tccgcaaaga atcccggcaa ccgccggcag
      661 ccggtgcgcc gtcgattagg aagccgccca agggcgacga gcaaccagat tttttcgttc
      721 cgatgctcta tgacgtgggc acccgcgata gtcgcagcat catggacgtg gccgttttcc
      781 gtctgtcgaa gcgtgaccga cgagctggcg aggtgatccg ctacgagctt ccagacgggc
      841 acgtagaggt ttccgcaggg ccggccggca tggccagtgt gtgggattac gacctggtac
      901 tgatggcggt ttcccatcta accgaatcca tgaaccgata ccgggaaggg aagggagaca
      961 agcccggccg cgtgttccgt ccacacgttg cggacgtact caagttctgc cggcgagccg
     1021 atggcggaaa gcagaaagac gacctggtag aaacctgcat tcggttaaac accacgcacg
     1081 ttgccatgca gcgtacgaag aaggccaaga acggccgcct ggtgacggta tccgagggtg
     1141 aagccttgat tagccgctac aagatcgtaa agagcgaaac cgggcggccg gagtacatcg
     1201 agatcgagct agctgattgg atgtaccgcg agatcacaga aggcaagaac ccggacgtgc
     1261 tgacggttca ccccgattac tttttgatcg atcccggcat cggccgtttt ctctaccgcc
     1321 tggcacgccg cgccgcaggc aaggcagaag ccagatggtt gttcaagacg atctacgaac
     1381 gcagtggcag cgccggagag ttcaagaagt tctgtttcac cgtgcgcaag ctgatcgggt
     1441 caaatgacct gccggagtac gatttgaagg aggaggcggg gcaggctggc ccgatcctag
     1501 tcatgcgcta ccgcaacctg atcgagggcg aagcatccgc cggttcctaa tgtacggagc
     1561 agatgctagg gcaaattgcc ctagcagggg aaaaaggtcg aaaaggactc tttcctgtgg
     1621 atagcacgta cattgggaac ccaaagccgt acattgggaa ccggaacccg tacattggga
     1681 acccaaagcc gtacattggg aaccggtcac acatgtaagt gactgatata aaagagaaaa
     1741 aaggcgattt ttccgcctaa aactctttaa aacttattaa aactcttaaa acccgcctgg
     1801 cctgtgcata actgtctggc cagcgcacag ccgaagtgct gcaaaaagcg cctacccttc
     1861 ggtcgctgcg ctccctacgc cccgccgctt cgcgtcggcc tatcgcggcc gctggccgct
     1921 caaaaatggc tggcctacgg ccaggcaatc taccagggcg cggacaagcc gcgccgtcgc
     1981 cactcgaccg ccggcgccca catcaaggca ccctgcctcg cgcgtttcgg tgatgacggt
     2041 gaaaacctct gacacatgca gctcccggtg acggtcacag cttgtctgta agcggatgcc
     2101 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc
     2161 atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg gcatcagagc
     2221 agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa
     2281 aataccgcat caggcgctca tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     2341 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     2401 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     2461 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     2521 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     2581 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     2641 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     2701 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     2761 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     2821 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     2881 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg
     2941 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3001 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3061 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     3121 cacgttaagg gattttggtc atgcattcta ggtgattaga aaaactcatc gagcatcaaa
     3181 tgaaactgca atttattcat atcaggatta tcaataccat atttttgaaa aagccgtttc
     3241 tgtaatgaag gagaaaactc accgaggcag ttccatagga tggcaagatc ctggtatcgg
     3301 tctgcgattc cgactcgtcc aacatcaata caacctatta atttcccctc gtcaaaaata
     3361 aggttatcaa gtgagaaatc accatgagtg acgactgaat ccggtgagaa tggcaaaagt
     3421 ttatgcattt ctttccagac ttgttcaaca ggccagccat tacgctcgtc atcaaaatca
     3481 ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct gagcgagtcg aaatacgcga
     3541 tcgctgttaa aaggacaatt acaaacagga atcgaatgca accggcgcag gaacactgcc
     3601 agcgcatcaa caatattttc acctgaatca ggatattctt ctaatacctg gaatgctgtt
     3661 ttccctggga tcgcagtggt gagtaaccat gcatcatcag gagtacggat aaaatgcttg
     3721 atggtcggaa gaggcataaa ttccgtcagc cagtttagtc tgaccatctc atctgtaaca
     3781 tcattggcaa cgctaccttt gccatgtttc agaaacaact ctggcgcatc gggcttccca
     3841 tacaatcggt agattgtcgc acctgattgc ccgacattat cgcgagccca tttataccca
     3901 tataaatcag catccatgtt ggaatttaat cgcggccttg agcaagacgt ttcccgttga
     3961 atatggctca taacagaact tattatttcc ttcctctttt ctacagtatt taaagatacc
     4021 ccaagaagct aattataaca agacgaactc caattcactg ttccttgcat tctaaaacct
     4081 taaataccag aaaacagctt tttcaaagtt gttttcaaag ttggcgtata acatagtatc
     4141 gacggagccg attttgaaac cgcggtgatc acaggcagca acgctctgtc atcgttacaa
     4201 tcaacatgct accctccgcg agatcatccg tgtttcaaac ccggcagctt agttgccgtt
     4261 cttccgaata gcatcggtaa catgagcaaa gtctgccgcc ttacaacggc tctcccgctg
     4321 acgccgtccc ggactgatgg gctgcctgta tcgagtggtg attttgtgcc gagctgccgg
     4381 tcggggagct gttggctggc tggtggcagg atatattgtg gtgtaaacat aacgaattgc
     4441 tcttcagcaa ggaggtcaac atggtggagc acgacactct ggtctactcc aaaaatgtca
     4501 aagatacagt ctcagaagat caaagggcta ttgagacttt tcaacaaagg ataatttcgg
     4561 gaaacctcct cggattccat tgcccagcta tctgtcactt catcgaaagg acagtagaaa
     4621 aggaaggtgg ctcctacaaa tgccatcatt gcgataaagg aaaggctatc attcaagatc
     4681 tctctgccga cagtggtccc aaagatggac ccccacccac gaggagcatc gtggaaaaag
     4741 aagaggttcc aaccacgtct acaaagcaag tggattgatg tgataacatg gtggagcacg
     4801 acactctggt ctactccaaa aatgtcaaag atacagtctc agaagatcaa agggctattg
     4861 agacttttca acaaaggata atttcgggaa acctcctcgg attccattgc ccagctatct
     4921 gtcacttcat cgaaaggaca gtagaaaagg aaggtggctc ctacaaatgc catcattgcg
     4981 ataaaggaaa ggctatcatt caagatctct ctgccgacag tggtcccaaa gatggacccc
     5041 cacccacgag gagcatcgtg gaaaaagaag aggttccaac cacgtctaca aagcaagtgg
     5101 attgatgtga catctccact gacgtaaggg atgacgcaca atcccactat ccttcgcaag
     5161 acccttcctc tatataagga agttcatttc atttggagag gacacgctcg agtataagag
     5221 ctcattttta caacaattac caacaacaac aaacaacaaa caacattaca attacattta
     5281 caattatcga tacaatggtg aaacaagaac gcaagatcca aaccagcagc acaaaaaagg
     5341 aaatgccttt gtcatcatca ccatcttctt cttcttcttc atcttcttcc tcgtcttcgt
     5401 cttcgtgtaa gaacaagaac aagaagagta agattaagaa gtacaaagga gtgaggatga
     5461 gaagttgggg atcatgggtt tctgagatta gggcaccaaa tcaaaagaca aggatttggt
     5521 taggttctta ctcaacagct gaagcagctg ctagagctta cgatgttgca ctcttatgtc
     5581 tcaaaggccc tcaagccaat ctcaacttcc ctacttcttc ttcttctcat catcttcttg
     5641 ataatctctt agatgaaaat acccttttgt cccccaaatc catccaaaga gtagctgctc
     5701 aagctgccaa ctcatttaac cattttgccc ctacttcatc agccgtctcg tcaccgtccg
     5761 atcatgatca tcaccatgat gatgggatgc aatctttgat gggatctttt gtggacaatc
     5821 atgtgtcttt gatggattca acatcttcat ggtatgatga tcataatggg atgttcttgt
     5881 ttgataatgg agctccattc aattactctc ctcaactaaa ctcgacgacg atgctcgatg
     5941 aatacttcta cgaagatgct gacattccgc tttggagttt caatggttcg ggcgggagcg
     6001 gtggaagcgg cgaaattcag caagccactg caggagtctc acaagacact tcggaaaatc
     6061 ctaacaaaac aatagttcct gcagcattac cacagctcac ccctaccttg gtgtcactgc
     6121 tggaggtgat tgaacccgag gtgttgtatg caggatatga tagctctgtt ccagattcag
     6181 catggagaat tatgaccaca ctcaacatgt taggtgggcg tcaagtgatt gcagcagtga
     6241 aatgggcaaa ggcgatacta ggcttgagaa acttacacct cgatgaccaa atgaccctgc
     6301 tacagtactc atggatgttt ctcatggcat ttgccttggg ttggagatca tacagacaat
     6361 caagcggaaa cctgctctgc tttgctcctg atctgattat taatgagcag agaatgtctc
     6421 taccctgcat gtatgaccaa tgtaaacaca tgctgtttgt ctcctctgaa ttacaaagat
     6481 tgcaggtatc ctatgaagag tatctctgta tgaaaacctt actgcttctc tcctcagttc
     6541 ctaaggaagg tctgaagagt caagagttat ttgatgagat tcgaatgact tatatcaaag
     6601 agctaggaaa agccatcgtc aaaagggaag ggaactccag tcagaactgg caacggtttt
     6661 accaactgac aaagcttctg gactccatgc atgaggtggt tgagaatctc cttacctact
     6721 gcttccagac atttttggat aagaccatga gtattgaatt cccagagatg ttagctgaaa
     6781 tcatcactaa tcagatacca aaatattcaa atggaaatat caaaaagctt ctgtttcatc
     6841 aaaaatgagc ttctctagct agagtcgatc gacaagctcg agtttctcca taataatgtg
     6901 tgagtagttc ccagataagg gaattagggt tcctataggg tttcgctcat gtgttgagca
     6961 tataagaaac ccttagtatg tatttgtatt tgtaaaatac ttctatcaat aaaatttcta
     7021 attcctaaaa ccaaaatcca gtactaaaat ccagatcgct atactgaaga gcaattctga
     7081 caggatatat tggcgggtaa acctaagaga aaagagcgtt tattagaata atcggatatt
     7141 taaaagggcg tgaaaaggtt tatccgttcg tccatttgta tgtgcatgcc aaccacaggg
     7201 ttcccctcgg gatcaaagta ctttgatcca acccctccgc tgctatagtg cagtcggctt
     7261 ctgacgttca gtgcagccgt catctgaaaa cgacatgtcg cacaagtcct aagttacgcg
     7321 acaggctgcc gccctgccct tttcctggcg ttttcttgtc gcgtgtttta gtcgcataaa
     7381 gtagaatact tgcgactaga accggagaca ttacgccatg aacaagagcg ccgccgctgg
     7441 cctgctgggc tatgcccgcg tcagcaccga cgaccaggac ttgaccaacc aacgggccga
     7501 actgcacgcg gccggctgca ccaagctgtt ttccgagaag atcaccggca ccaggcgcga
     7561 ccgcccggag ctggccagga tgcttgacca cctacgccct ggcgacgttg tgacagtgac
     7621 caggctagac cgcctggccc gcagcacccg cgacctactg gacattgccg agcgcatcca
     7681 ggaggccggc gcgggcctgc gtagcctggc agagccgtgg gccgacacca ccacgccggc
     7741 cggccgcatg gtgttgaccg tgttcgccgg cattgccgag ttcgagcgtt ccctaatcat
     7801 cgaccgcacc cggagcgggc gcgaggccgc caaggcccga ggcgtgaagt ttggcccccg
     7861 ccctaccctc accccggcac agatcgcgca cgcccgcgag ctgatcgacc aggaaggccg
     7921 caccgtgaaa gaggcggctg cactgcttgg cgtgcatcgc tcgaccctgt accgcgcact
     7981 tgagcgcagc gaggaagtga cgcccaccga ggccaggcgg cgcggtgcct tccgtgagga
     8041 cgcattgacc gaggccgacg ccctggcggc cgccgagaat gaacgccaag aggaacaagc
     8101 atgaaaccgc accaggacgg ccaggacgaa ccgtttttca ttaccgaaga gatcgaggcg
     8161 gagatgatcg cggccgggta cgtgttcgag ccgcccgcgc acctctcaac cgtgcggctg
     8221 catgaaatcc tggccggttt gtctgatgcc aagctggcgg cctggccggc cagcttggcc
     8281 gctgaagaaa ccgagcgccg ccgtctaaaa aggtgatgtg tatttgagta aaacagcttg
     8341 cgtcatgcgg tcgctgcgta tatgatgcga tgagtaaata aacaaatacg caaggggaac
     8401 gcatgaaggt tatcgctgta cttaaccaga aaggcgggtc aggcaagacg accatcgcaa
     8461 cccatctagc ccgcgccctg caactcgccg gggccgatgt tctgttagtc gattccgatc
     8521 cccagggcag tgcccgcgat tgggcggccg tgcgggaaga tcaaccgcta accgttgtcg
     8581 gcatcgaccg cccgacgatt gaccgcgacg tgaaggccat cggccggcgc gacttcgtag
     8641 tgatcgacgg agcgccccag gcggcggact tggctgtgtc cgcgatcaag gcagccgact
     8701 tcgtgctgat tccggtgcag ccaagccctt acgacatatg ggccaccgcc gacctggtgg
     8761 agctggttaa gcagcgcatt gaggtcacgg atggaaggct acaagcggcc tttgtcgtgt
     8821 cgcgggcgat caaaggcacg cgcatcggcg gtgaggttgc cgaggcgctg gccgggtacg
     8881 agctgcccat tcttgagtcc cgtatcacgc agcgcgtgag ctacccaggc actgccgccg
     8941 ccggcacaac cgttcttgaa tcagaacccg agggcgacgc tgcc
//
