LOCUS       40924_22568        7086 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pGrDL_SP, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7086)
  AUTHORS   Moyle RL, Carvalhais L, Nowak E, Pretorius L-S., Subramaniam G, 
            Dalton-Morgan J, Schenk PM.
  TITLE     Quantifying MicroRNA Directed Repression of Targeted Transcripts 
            using an Optimised Transient Dual Luciferase Assay
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7086)
  AUTHORS   Moyle RL, Carvalhais L, Nowak E, Pretorius L-S., Subramaniam G, 
            Dalton-Morgan J, Schenk PM.
  TITLE     Direct Submission
  JOURNAL   Submitted (20-AUG-2016) School of Agriculture and Food Science, 
            University of Queensland, St Lucia, Brisbane, QLD 4072, Australia
REFERENCE   3  (bases 1 to 7086)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7086)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (20-AUG-2016) School of Agriculture and Food Science, University of 
            Queensland, St Lucia, Brisbane, QLD 4072, Australia"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7086
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    542..564
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA
                     (truncated)"
     terminator      complement(587..839)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     CDS             complement(857..1789)
                     /label=Rluc
                     /note="luciferase from the anthozoan coelenterate Renilla 
                     reniformis (sea pansy)"
     promoter        complement(1827..2006)
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     primer_bind     2253..2269
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2279..2297
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2372..2715
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     CDS             2722..4371
                     /label=luciferase
                     /note="firefly luciferase"
     regulatory      4484..4649
                     /label=35S terminator
                     /note="35S terminator"
                     /regulatory_class="terminator"
     promoter        complement(4685..4703)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4724..4740)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4748..4764)
                     /label=lac operator
                     /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(4772..4803)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4818..4839)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    5078..5102
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(5193..5781)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5955..6767)
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     rep_origin      7058..7086
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 tttttatccc cggaagcctg tggatagagg gtagttatcc acgtgaaacc gctaatgccc
       61 cgcaaagcct tgattcacgg ggctttccgg cccgctccaa aaactatcca cgtgaaatcg
      121 ctaatcaggg tacgtgaaat cgctaatcgg agtacgtgaa atcgctaata aggtcacgtg
      181 aaatcgctaa tcaaaaaggc acgtgagaac gctaatagcc ctttcagatc aacagcttgc
      241 aaacacccct cgctccggca agtagttaca gcaagtagta tgttcaatta gcttttcaat
      301 tatgaatata tatatcaatt attggtcgcc cttggcttgt ggacaatgcg ctacgcgcac
      361 cggctccgcc cgtggacaac cgcaagcggt tgcccaccgt cgagcgccag cgcctttgcc
      421 cacaacccgg cggccggccg caacagatcg ttttataaat tttttttttt gaaaaagaaa
      481 aagcccgaaa ggcggcaacc tctcgggctt ctggatttcc gatccccgga attagagatc
      541 ttggcaggat atattgtggt gtaacgtatc agcttgcatg ccggtcgatc tagtaacata
      601 gatgacaccg cgcgcgataa tttatcctag tttgcgcgct atattttgtt ttctatcgcg
      661 tattaaatgt ataattgcgg gactctaatc ataaaaaccc atctcataaa taacgtcatg
      721 cattacatgt taattattac atgcttaacg taattcaaca gaaattatat gataatcatc
      781 gcaagaccgg caacaggatt caatcttaag aaactttatt gccaaatgtt tgaacgatct
      841 gcttgactct agattattgt tcatttttga gaactcgctc aacgaacgat ttgatatatt
      901 ttcccatttc atcaggtgca tcttcttgcg aaaaatgaag accttttact ttgacaaatt
      961 cagtattagg aaacttcttg gcaccttcaa caatagcatt ggaaaagaat cctgggtccg
     1021 attcaataaa catttttggt aaatcatcac ttgcacgtag ataagcatta taattcctaa
     1081 caatttgtac aacgtcaggt ttaccacctt ttactaacgg gatttcacga ggccatgata
     1141 atgttggacg acgaacttca cctttctctt tgaatggttc aagatatgct gcaaattctt
     1201 ctggttctaa ctttctcatg atttttgatg gcaacatggt ttccacgaag aagttattct
     1261 ccaaaaccat tttttctcct tcttcagatt tgatcaacgc aatatcttct tcaatatcag
     1321 gccattcatc ccatgattca atcacatcta ctacactttc agcgtgaact attgctttga
     1381 tcttatcttg atgctcatag ctataatgaa atgccaaaca agcaccccaa tcatggccga
     1441 caaaaatgat cttctttggt aaattaagaa gttcaaacca tgcagtaaga tatttgtaat
     1501 gatcaagtaa cctataagaa ccattaccag atttgcctga tttgcccata ccaataaggt
     1561 ctggtataat acaccgcgct actggctcaa tatgtggcac aacatgtcgc cataaataag
     1621 aagaggccgc gttaccatgt aaaaaaataa cagcattttc tgcatgtttt tctgaatcat
     1681 aataattaat aaatgaatca agaacattca tttgtttaca tctggcccac cactgcggac
     1741 cagttatcat ccgtttcctt tgttctggat cataaacttt cgaagtcatg tcgatcgacc
     1801 tcgagttgag agtgaatatg agactctaat tggataccga ggggaattta tggaacgtca
     1861 gtggagcatt tttgacaaga aatatttgct agctgatagt gaccttaggc gacttttgaa
     1921 cgcgcaataa tggtttctga cgtatgtgct tagctcatta aactccagaa acccgcggct
     1981 cagtggctcc ttcaacgttg cggttctgtc agttccaaac gtaaaacggc ttgtcccgcg
     2041 tcatcggcgg gggtcataac gtgactccct taattctccg ctcatgatcg ataacattaa
     2101 cgcttacaat ttccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     2161 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     2221 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgcgta
     2281 atacgactca ctatagggcg aattgggtac cgggcccacc cccctactcc aaaaatgtca
     2341 aagatacagt ctcagaagac caaagggcta ttgagacttt tcaacaaagg gtaatttcgg
     2401 gaaacctcct cggattccat tgcccagcta tctgtcactt catcgaaagg acagtagaaa
     2461 aggaaggtgg ctcctacaaa tgccatcatt gcgataaagg aaaggctatc attcaagatg
     2521 cctctgccga cagtggtccc aaagatggac ccccacccac gaggagcatc gtggaaaaag
     2581 aagacgttcc aaccacgtct tcaaagcaag tggattgatg tgacatctcc actgacgtaa
     2641 gggatgacgc acaatcccac tatccttcgc aagacccttc ctctatataa ggaagtcatt
     2701 tcatttggag aggacctcga gatggaagac gccaaaaaca taaagaaagg cccggcgcca
     2761 ttctatccgc tggaagatgg aaccgctgga gagcaactgc ataaggctat gaagagatac
     2821 gccctggttc ctggaacaat tgcttttaca gatgcacata tcgaggtgga catcacttac
     2881 gctgagtact tcgaaatgtc cgttcggttg gcagaagcta tgaaacgata tgggctgaat
     2941 acaaatcaca gaatcgtcgt atgcagtgaa aactctcttc aattctttat gccggtgttg
     3001 ggcgcgttat ttatcggagt tgcagttgcg cccgcgaacg acatttataa tgaacgtgaa
     3061 ttgctcaaca gtatgggcat ttcgcagcct accgtggtgt tcgtttccaa aaaggggttg
     3121 caaaaaattt tgaacgtgca aaaaaagctc ccaatcatcc aaaaaattat tatcatggat
     3181 tctaaaacgg attaccaggg atttcagtcg atgtacacgt tcgtcacatc tcatctacct
     3241 cccggtttta atgaatacga ttttgtgcca gagtccttcg atagggacaa gacaattgca
     3301 ctgatcatga actcctctgg atctactggt ctgcctaaag gtgtcgctct gcctcataga
     3361 actgcctgcg tgagattctc gcatgccaga gatcctattt ttggcaatca aatcattccg
     3421 gatactgcga ttttaagtgt tgttccattc catcacggtt ttggaatgtt tactacactc
     3481 ggatatttga tatgtggatt tcgagtcgtc ttaatgtata gatttgaaga agagctgttt
     3541 ctgaggagcc ttcaggatta caagattcaa agtgcgctgc tggtgccaac cctattctcc
     3601 ttcttcgcca aaagcactct gattgacaaa tacgatttat ctaatttaca cgaaattgct
     3661 tctggtggcg ctcccctctc taaggaagtc ggggaagcgg ttgccaagag gttccatctg
     3721 ccaggtatca ggcaaggata tgggctcact gagactacat cagctattct gattacaccc
     3781 gagggggatg ataaaccggg cgcggtcggt aaagttgttc cattttttga agcgaaggtt
     3841 gtggatctgg ataccgggaa aacgctgggc gttaatcaaa gaggcgaact gtgtgtgaga
     3901 ggtcctatga ttatgtccgg ttatgtaaac aatccggaag cgaccaacgc cttgattgac
     3961 aaggatggat ggctacattc tggagacata gcttactggg acgaagacga acacttcttc
     4021 atcgttgacc gcctgaagtc tctgattaag tacaaaggct atcaggtggc tcccgctgaa
     4081 ttggaatcca tcttgctcca acaccccaac atcttcgacg caggtgtcgc aggtcttccc
     4141 gacgatgacg ccggtgaact tcccgccgcc gttgttgttt tggagcacgg aaagacgatg
     4201 acggaaaaag agatcgtgga ttacgtcgcc agtcaagtaa caaccgcgaa aaagttgcgc
     4261 ggaggagttg tgtttgtgga cgaagtaccg aaaggtctta ccggaaaact cgacgcaaga
     4321 aaaatcagag agatcctcat aaaggccaag aagggcggaa agatcgccgt gtaagaattc
     4381 cttgtcgact tccctgcagt cgccatgcgg tcattacact atccctagga tcggtatcca
     4441 gctagtacaa accggtttta ctatctttta ctcttgaact agttaatgtg tgagtagttt
     4501 cccgataagg gaaattaggg ttcttatagg gtttcgctca tgtgttgagc atataagaaa
     4561 cccttagtat gtatttgtat ttgtaaaata cttctatcaa taaaatttct aattcctaaa
     4621 accaaaatcc agtactaaaa tccagatcgc ggccgccacc gcggtggagc tccagctttt
     4681 gttcccttta gtgagggtta attccgagct tggcgtaatc atggtcatag ctgtttcctg
     4741 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagh cataaagtgt
     4801 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     4861 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4921 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4981 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5041 gaatcagggg ataacgcagg aaagaacatg aaggccttga caggatatat tggcgggtaa
     5101 actaagtcgc tgtatgtgtt tgtttgagat ctcatgtgag caaaaggcca gcaaaaggcc
     5161 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     5221 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     5281 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     5341 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     5401 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     5461 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     5521 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     5581 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     5641 tttggtatct gcgctctgct gaagccagtt accttcggaa gaagagttgg tagctcttga
     5701 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     5761 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     5821 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     5881 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgtgtaacat
     5941 tggtctagtg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca
     6001 ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg
     6061 aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca
     6121 tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca
     6181 tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt
     6241 tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc
     6301 attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa
     6361 acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct
     6421 gaatcaggat attcttctaa tacctggaat gctgttttcc ctgggatcgc agtggtgagt
     6481 aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc
     6541 gtcagccagt ttagtctgac catctcatct gtaacaacat tggcaacgct acctttgcca
     6601 tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcggtagat tgtcgcacct
     6661 gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa
     6721 tttaatcgcg gccttgagca agacgtttcc cgttgaatat ggctcataac accccttgta
     6781 ttactgttta tgtaagcaga cagttttatt gttcatgatg atatattttt atcttgtgca
     6841 atgtaacatc agagattttg agacacaacg tggctttgtt gaataaatcg aacttttgct
     6901 gagttgaagg atcagatcac gcatcttccc gacaacgcag accgttccgt ggcaaagcaa
     6961 aagttcaaaa tcaccaactg gtccacctac aacaaagctc tcatcaaccg tggctccctc
     7021 actttctggc tggatgatgg ggcgattcag gcgatcccca tccaacagcc cgccgtcgag
     7081 cgggct
//
