LOCUS       sequence-239370-        3333 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RBCS terminator with GreenGate C-D flanking sequences.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3333)
  AUTHORS   Lee JE, Neumann M, Duro DI, Schmid M
  TITLE     CRISPR-based tools for targeted transcriptional and epigenetic 
            regulation in plants.
  JOURNAL   PLoS One. 2019 Sep 26;14(9):e0222778. doi: 
            10.1371/journal.pone.0222778. eCollection 2019.
  PUBMED    31557222
REFERENCE   2  (bases 1 to 3333)
  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: "PLoS One.";
            date: "2019-09-26"; volume: "14(9):e0222778. doi"; pages: " 
            10.1371/journal.pone.0222778. eCollection 2019"
COMMENT     Sequence Label: pEF003
FEATURES             Location/Qualifiers
     source          1..3333
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     45..67
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     77..96
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        77..95
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      complement(120..761)
                     /label=E9 terminator
                     /note="terminator and polyadenylation signal from the pea 
                     rbcS-E9 gene"
     promoter        complement(785..803)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(786..803)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    817..833
                     /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(841..847,848..865,866..871))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    886..907
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1024..1041)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1195..1783)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1275..1294)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1954..2745,2746..2814))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2577..2596
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2815..2919)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2987..3005
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3043..3065)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3165..3184
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
       61 taaaacgacg gccaggtaat acgactcact atagggcgag aattcggtct cactgacgat
      121 tgatgcatgt tgtcaatcaa ttggcaagtc ataaaatgca ttaaaaaata ttttcatact
      181 caactacaaa tccatgagta taactataat tataaagcaa tgattagaat ctgacaagga
      241 ttctggaaaa ttacataaag gaaagttcat aaatgtctaa aacacaagag gacatacttg
      301 tattcagtaa catttgcagc ttttctaggt ctgaaaatat atttgttgcc tagtgaataa
      361 gcataatggt acaactacaa gtgttttact cctcatatta acttcggtca ttagaggcca
      421 cgatttgaca catttttact caaaacaaaa tgtttgcata tctcttataa tttcaaattc
      481 aacacacaac aaataagaga aaaaacaaat aatattaatt tgagaatgaa caaaaggacc
      541 atatcattca ttaactcttc tccatccatt tccatttcac agttcgatag cgaaaaccga
      601 ataaaaaaca cagtaaatta caagcacaac aaatggtaca agaaaaacag ttttcccaat
      661 gccataatac tcaaactcag taggattctg gtgtgtgcgc aatgaaactg atgcattgaa
      721 cttgacgaac gttgtcgaaa ccgatgatac gaacgaaagc tttagcctga gaccaagctt
      781 cacttctata gtgtcaccta aatagctcgt gtgaaattgt tatccgctca caattccaca
      841 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
      901 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
      961 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     1021 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     1081 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     1141 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     1201 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     1261 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     1321 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     1381 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     1441 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     1501 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     1561 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     1621 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     1681 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     1741 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     1801 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     1861 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     1921 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     1981 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     2041 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac ctcgagagcc
     2101 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     2161 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     2221 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     2281 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     2341 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     2401 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     2461 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     2521 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     2581 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     2641 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     2701 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     2761 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     2821 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     2881 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     2941 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     3001 gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     3061 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     3121 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
     3181 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
     3241 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     3301 ggcctcttcg ctattacgcc agctggcgaa agg
//
