LOCUS       sequence-304709-        5476 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for STARR-seq in tobacco leaves. Enhancer candidates can be 
            inserted about 1.4 kb upstream of the minimal promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5476)
  AUTHORS   Jores T, Tonnies J, Dorrity MW, Cuperus J, Fields S, Queitsch C
  TITLE     Identification of Plant Enhancers and Their Constituent Elements by 
            STARR-seq in Tobacco Leaves.
  JOURNAL   Plant Cell. 2020 May 14. pii: tpc.20.00155. doi: 
            10.1105/tpc.20.00155.
  PUBMED    32409318
REFERENCE   2  (bases 1 to 5476)
  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: "Plant Cell.
            2020 May 14. pii: tpc.20.00155. doi: 10.1105/tpc.20.00155."
COMMENT     Sequence Label: pPSup2
FEATURES             Location/Qualifiers
     source          1..5476
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     27..46
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        27..45
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     70..86
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     71..87
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             join(157..159,160..873)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(199..220)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(460..479)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     807..828
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1194..1214)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1194..1212)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1233..1249)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1233..1249)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1246..1268)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1257..1273
                     /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)."
     promoter        complement(join(1281..1287,1288..1305,1306..1311))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1326..1347
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1464..1481)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1586..1610
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(1701..2289)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1781..1800)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2460..3248)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNGTLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      3539..3974
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
     misc_feature    4102..4124
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     terminator      complement(4186..4438)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     CDS             complement(4458..5009)
                     /codon_start=1
                     /gene="Streptomyces hygroscopicus bar"
                     /product="phosphinothricin acetyltransferase"
                     /label=BlpR
                     /note="confers resistance to bialophos or phosphinothricin"
                     /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
                     WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
                     TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
                     WQLDFSLPVPPRPVLPVTEI"
     promoter        complement(5051..5230)
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     primer_bind     join(5462..5476,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(5476,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtaccg
       61 ggccccccct cgaggtcgac ggtatcgata agcttgatat cgaattccgc ggccgcagga
      121 gctgccgtct tcagtgcctg caggcataga agacccgtga gcaagggcga ggagctgttc
      181 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      241 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
      301 accaccggca agctgcccgt gccctggccc accctcgtga ccaccttcac ctacggcgtg
      361 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
      421 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
      481 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
      541 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
      601 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
      661 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
      721 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
      781 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
      841 atcactcacg gcatggacga gctgtacaag taaagcggcc gcaaattgcc cagaattaat
      901 tccccaactg cacagaacaa atttcccttt gcttttgtgt agacctcaaa actttatccc
      961 ccatctttga tcttatccct tgtttttctg cttttttctt ctttcttggg ttttgatttc
     1021 cggacttaac gtttgttttc cggtttgcga gacatattct atcggattct caactgtctg
     1081 atgaaataaa tatgtaatgt tctataagtc tttcaatttg atgggatgtt ccacttctgt
     1141 ataggagcca attgattgac aacatgcatc aatcggagct ccagcttttg ttccctttag
     1201 tgagggttaa ttgcgcgctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     1261 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     1321 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     1381 ggaaacctgt cgtgccagct gcattaatga atcggccaac gctcggggag aggcggtttg
     1441 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     1501 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     1561 aacgcaggaa agaacatgaa ggccttgaca ggatatattg gcgggtaaac taagtcgctg
     1621 tatgtgtttg tttgagatct catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     1681 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     1741 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     1801 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     1861 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     1921 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     1981 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     2041 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     2101 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     2161 gctctgctga agccagttac cttcggaaga agagttggta gctcttgatc cggcaaacaa
     2221 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     2281 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     2341 tcacgttaag ggattttggt cattagatta tcaaaaagga tcttcaccta gatcctttta
     2401 aattaaaaat gaagttttaa atcaatctaa agtatatatg tgtaacattg gtctagtgat
     2461 tatttgccga ctaccttggt gatctcgcct ttcacgtagt gaacaaattc ttccaactga
     2521 tctgcgcgcg aggccaagcg atcttcttgt ccaagataag cctgcctagc ttcaagtatg
     2581 acgggctgat actgggccgg caggcgctcc attgcccagt cggcagcgac atccttcggc
     2641 gcgattttgc cggttactgc gctgtaccaa atgcgggaca acgtaagcac tacatttcgc
     2701 tcatcgccag cccagtcggg cggcgagttc catagcgtta aggttccatt tagcgcctca
     2761 aatagatcct gttcaggaac cggatcaaag agttcctccg ccgctggacc taccaaggca
     2821 acgctatgtt ctcttgcttt tgtcagcaag atagccagat caatgtcgat cgtggctggc
     2881 tcgaagatac ctgcaagaat gtcattgcgc tgccattctc caaattgcag ttcgcgctta
     2941 gctggataac gccacggaat gatgtcgtcg tgcacaacaa tggtgacttc tacagcgcgg
     3001 agaatctcgc tctctccagg ggaagccgaa gtttccaaaa ggtcgttgat caaagctcgc
     3061 cgcgttgttt catcaagcct tacggtcacc gtaaccagca aatcaatatc actgtgtggc
     3121 ttcaggccgc catccactgc ggagccgtac aaatgtacgg ccagcaacgt cggttcgaga
     3181 tggcgctcga tgacgccaac tacctctgat agttgagtcg atacttcggc gatcaccgct
     3241 tccctcataa caccccttgt attactgttt atgtaagcag acagttttat tgttcatgat
     3301 gatatatttt tatcttgtgc aatgtaacat cagagatttt gagacacaac gtggctttgt
     3361 tgaataaatc gaacttttgc tgagttgaag gatcagatca cgcatcttcc cgacaacgca
     3421 gaccgttccg tggcaaagca aaagttcaaa atcaccaact ggtccaccta caacaaagct
     3481 ctcatcaacc gtggctccct cactttctgg ctggatgatg gggcgattca ggcgatcccc
     3541 atccaacagc ccgccgtcga gcgggctttt ttatccccgg aagcctgtgg atagagggta
     3601 gttatccacg tgaaaccgct aatgccccgc aaagccttga ttcacggggc tttccggccc
     3661 gctccaaaaa ctatccacgt gaaatcgcta atcagggtac gtgaaatcgc taatcggagt
     3721 acgtgaaatc gctaataagg tcacgtgaaa tcgctaatca aaaaggcacg tgagaacgct
     3781 aatagccctt tcagatcaac agcttgcaaa cacccctcgc tccggcaagt agttacagca
     3841 agtagtatgt tcaattagct tttcaattat gaatatatat atcaattatt ggtcgccctt
     3901 ggcttgtgga caatgcgcta cgcgcaccgg ctccgcccgt ggacaaccgc aagcggttgc
     3961 ccaccgtcga gcgcctttgc ccacaacccg gcggccggcc gcaacagatc gttttataaa
     4021 tttttttttt ttgaaaaaga aaaagcccga aaggcggcaa cctctcgggc ttctggattt
     4081 ccgatccccg gaattagatc ttggcaggat atattgtggt gtaacgttat cagcttgcat
     4141 gccgactccg agaccacaag gcgcgcctag tggtctccag gagtcgatct agtaacatag
     4201 atgacaccgc gcgcgataat ttatcctagt ttgcgcgcta tattttgttt tctatcgcgt
     4261 attaaatgta taattgcggg actctaatca taaaaaccca tctcataaat aacgtcatgc
     4321 attacatgtt aattattaca tgcttaacgt aattcaacag aaattatatg ataatcatcg
     4381 caagaccggc aacaggattc aatcttaaga aactttattg ccaaatgttt taacgatctg
     4441 cttgactcta ggggtcatca gatttcggtg acgggcagga ccggacgggg cggcaccggc
     4501 aggctgaagt ccagctgcca gaaacccacg tcatgccagt tcccgtgctt gaagccggcc
     4561 gcccgcagca tgccgcgggg ggcatatccg agcgcctcgt gcatgcgcac gctcgggtcg
     4621 ttgggcagcc cgatgacagc gaccacgctc ttgaagccct gtgcctccag ggacttcagc
     4681 aggtgggtgt agagcgtgga gcccagtccc gtccgctggt ggcgggggga gacgtacacg
     4741 gtggactcgg ccgtccagtc gtaggcgttg cgtgccttcc agggacccgc gtaggcgatg
     4801 ccggcgacct cgccgtccac ctcggcgacg agccagggat agcgctcccg cagacggacg
     4861 aggtcgtccg tccactcctg cggttcctgc ggctcggtac ggaagttgac cgtgcttgtc
     4921 tcgatgtagt ggttgacgat ggtgcagacc gccggcatgt ccgcctcggt ggcacggcgg
     4981 atgtcggccg ggcgtcgttc tgggctcatg gtagatcccc tcgatcgagt tgagagtgaa
     5041 tatgagactc taattggata ccgaggggaa tttatggaac gtcagtggag catttttgac
     5101 aagaaatatt tgctagctta tagtgacctt aggcgacttt tgaacgcgca ataatggttt
     5161 ctgacgtatg tgcttagctc attaaactcc agaaacccgc ggctcagtgg ctccttcaac
     5221 gttgcggttc tgtcagttcc aaacgtaaaa cggcttgtcc cgcgtcatcg gcgggggtca
     5281 taacgtgact cccttaattc tccgctcatg atcgataaca ttaacgctta caatttccat
     5341 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     5401 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
     5461 tcccagtcac gacgtt
//
