LOCUS       sequence-304711-        5476 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for STARR-seq in tobacco leaves. Enhancer candidates can be 
            inserted downstream of the GFP reporter gene..
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: pPSdown2
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    1624..1648
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(1739..2327)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1819..1838)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2498..3286)
                     /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      3577..4012
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
     misc_feature    4140..4162
                     /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 aacgcactcc gagaccacaa ggcgcgccta gtggtctcca ggaaggaaag aacatgaagg
     1621 ccttgacagg atatattggc gggtaaacta agtcgctgta tgtgtttgtt tgagatctca
     1681 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     1741 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     1801 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     1861 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     1921 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     1981 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2041 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2101 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2161 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2221 tcggaagaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2281 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     2341 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     2401 ttagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     2461 caatctaaag tatatatgtg taacattggt ctagtgatta tttgccgact accttggtga
     2521 tctcgccttt cacgtagtga acaaattctt ccaactgatc tgcgcgcgag gccaagcgat
     2581 cttcttgtcc aagataagcc tgcctagctt caagtatgac gggctgatac tgggccggca
     2641 ggcgctccat tgcccagtcg gcagcgacat ccttcggcgc gattttgccg gttactgcgc
     2701 tgtaccaaat gcgggacaac gtaagcacta catttcgctc atcgccagcc cagtcgggcg
     2761 gcgagttcca tagcgttaag gttccattta gcgcctcaaa tagatcctgt tcaggaaccg
     2821 gatcaaagag ttcctccgcc gctggaccta ccaaggcaac gctatgttct cttgcttttg
     2881 tcagcaagat agccagatca atgtcgatcg tggctggctc gaagatacct gcaagaatgt
     2941 cattgcgctg ccattctcca aattgcagtt cgcgcttagc tggataacgc cacggaatga
     3001 tgtcgtcgtg cacaacaatg gtgacttcta cagcgcggag aatctcgctc tctccagggg
     3061 aagccgaagt ttccaaaagg tcgttgatca aagctcgccg cgttgtttca tcaagcctta
     3121 cggtcaccgt aaccagcaaa tcaatatcac tgtgtggctt caggccgcca tccactgcgg
     3181 agccgtacaa atgtacggcc agcaacgtcg gttcgagatg gcgctcgatg acgccaacta
     3241 cctctgatag ttgagtcgat acttcggcga tcaccgcttc cctcataaca ccccttgtat
     3301 tactgtttat gtaagcagac agttttattg ttcatgatga tatattttta tcttgtgcaa
     3361 tgtaacatca gagattttga gacacaacgt ggctttgttg aataaatcga acttttgctg
     3421 agttgaagga tcagatcacg catcttcccg acaacgcaga ccgttccgtg gcaaagcaaa
     3481 agttcaaaat caccaactgg tccacctaca acaaagctct catcaaccgt ggctccctca
     3541 ctttctggct ggatgatggg gcgattcagg cgatccccat ccaacagccc gccgtcgagc
     3601 gggctttttt atccccggaa gcctgtggat agagggtagt tatccacgtg aaaccgctaa
     3661 tgccccgcaa agccttgatt cacggggctt tccggcccgc tccaaaaact atccacgtga
     3721 aatcgctaat cagggtacgt gaaatcgcta atcggagtac gtgaaatcgc taataaggtc
     3781 acgtgaaatc gctaatcaaa aaggcacgtg agaacgctaa tagccctttc agatcaacag
     3841 cttgcaaaca cccctcgctc cggcaagtag ttacagcaag tagtatgttc aattagcttt
     3901 tcaattatga atatatatat caattattgg tcgcccttgg cttgtggaca atgcgctacg
     3961 cgcaccggct ccgcccgtgg acaaccgcaa gcggttgccc accgtcgagc gcctttgccc
     4021 acaacccggc ggccggccgc aacagatcgt tttataaatt tttttttttt gaaaaagaaa
     4081 aagcccgaaa ggcggcaacc tctcgggctt ctggatttcc gatccccgga attagatctt
     4141 ggcaggatat attgtggtgt aacgttatca gcttgcatgc cggtcgatct 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
//
