LOCUS       40924_36628        8191 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway recycling vector pRED56-MD8-Pnos-18 DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8191)
  AUTHORS   Saisho T, Shibahara K, Nakao A, Tanaka Y, Nishimura K, Kimura T, 
            Nakagawa T.
  TITLE     Improved Gateway Recycling Cloning System for Multigene Construction
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8191)
  AUTHORS   Saisho T, Shibahara K, Nakao A, Tanaka Y, Nishimura K, Kimura T, 
            Nakagawa T.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-NOV-2015) Contact:Tsuyoshi Nakagawa Shimane 
            University, Interdisciplinary Center for Science Research; 1060 
            Nishikawatsu, Matsue, Shimane 690-8504, Japan
REFERENCE   3  (bases 1 to 8191)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8191)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (02-NOV-2015) Contact:Tsuyoshi Nakagawa Shimane University, 
            Interdisciplinary Center for Science Research; 1060 Nishikawatsu, 
            Matsue, Shimane 690-8504, Japan"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     constructed using pDONR201.
FEATURES             Location/Qualifiers
     source          1..8191
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    28..127
                     /label=attL1
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    674..1071
                     /label=MD8
                     /note="MD8"
     promoter        1192..1375
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     CDS             1408..1437
                     /codon_start=1
                     /label=Myc
                     /note="Myc (human c-Myc proto-oncogene) epitope tag"
                     /translation="EQKLISEEDL"
     CDS             1444..1473
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1489..1518
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1525..1554
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    1573..1697
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1734..1764
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             1818..2474
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             2819..3121
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(3165..3289)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      3315..3567
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    3776..3900
                     /label=attR4
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3925..3955
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     misc_feature    4227..4267
                     /label=rare cutter sites
                     /note="rare cutter sites"
     CDS             complement(4359..5216)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5217..5321)
                     /label=AmpR promoter
     protein_bind    complement(5618..5741)
                     /label=attR3
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(6067..6162)
                     /label=attL6
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             6286..7092
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      7238..7826
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(7961..7988)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(8080..8166)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
ORIGIN
        1 gttaacgcta gcatggatct cgggccccaa ataatgattt tattttgact gatagtgacc
       61 tgttcgttgc aacaaattga tgagcaatgc ttttttataa tgccaacttt gtatacaaaa
      121 gtaggctgga tgaatcgaat aacttcttct tcaaatgcgc aaacgaagaa atttcctcag
      181 tctcagccac agccgccgga gaaaaatgag aaaaaactga aaagggagag atttccggtg
      241 aaccctttga gagatttcag tacaagaagc tctggaagca gcagttgtag caatgagagt
      301 gcttctggtt caacttccgg tgaagcatcc aacggttgtc acaggttctt gctctctcat
      361 tctttctcat cgtcttcttc atcatcttct tcttcgttgg gagtttttcc tcgtagacca
      421 gtgagatcag tagctaaaaa ccccaaatct gctccggtag ttagtaaacc cttaattaga
      481 aagaaaccaa gctctcttga ggaggttaag ttgaaatcta ctcttactga gaaacctaat
      541 ttgctgaagt ctcagagatg caagacaaat cctgtatctg gaaagagacc aacttgtaag
      601 attaccatga aaccggagaa agtttcggtt ttgaagaaac aatcttctgt ttctaggtaa
      661 gctgcagtgt tttcatcaca cgtaatatca tccattttat attatgatcc acaaacagca
      721 aaggcccatt aatctcatta tattttagat taattgattg caattgccca acttctccac
      781 tcagcaatgt tagttttaaa gacacatcaa gttagtgtat tttattttta atctttctat
      841 aaaaaacaac actttgtttt tgtctatttc ttattctgtg aaaaagagaa atgaaaaagt
      901 aaaaaagata acaaaaaggc tggtttttca cggtgagaat ttttcatggt cacgcaataa
      961 cttaaatatg aactattaaa ataactaaga ttgaaaaatt agcatgatca aaagaagaag
     1021 aaaaaaaaga caacatgaag cggaaaggaa aaaaaagaaa gctgaaaatc taagcttatc
     1081 atgagcggag aattaaggga gtcacgttat gacccccgcc gatgacgcgg gacaagccgt
     1141 tttacgtttg gaactgacag aaccgcaacg ttgaaggagc cactcagccg cgggtttctg
     1201 gagtttaatg agctaagcac atacgtcaga aaccattatt gcgcgttcaa aagtcgccta
     1261 aggtcactat cagctagcaa atatttcttg tcaaaaatgc tccactgacg ttccataaat
     1321 tcccctcggt atccaattag agtctcatat tcactctcaa tccaaataat ctgcaccgaa
     1381 tctagaatga gcgggttaat taacggtgaa caaaagctaa tctccgagga agacttgaac
     1441 ggtgaacaaa aattaatctc agaagaagac ttgaacggac tcgacggtga acaaaagttg
     1501 atttctgaag aagatttgaa cggtgaacaa aagctaatct ccgaggaaga cttgaacggt
     1561 agcgctgtta tcacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa tgatataaat
     1621 atcaatatat taaattagat tttgcataaa aaacagacta cataatactg taaaacacaa
     1681 catatccagt cactatggcg gcccctcgag gggccgcatt aggcacccca ggctttacac
     1741 tttatgcttc cggctcgtat aatgtgtgga ttttgagtta ggatccggcg agattttcag
     1801 gagctaagga agctaaaatg gagaaaaaaa tcactggata taccaccgtt gatatatccc
     1861 aatggcatcg taaagaacat tttgaggcat ttcagtcagt tgctcaatgt acctataacc
     1921 agaccgttca gctggatatt acggcctttt taaagaccgt aaagaaaaat aagcacaagt
     1981 tttatccggc ctttattcac attcttgccc gcctgatgaa tgctcatccg gaattccgta
     2041 tggcaatgaa agacggtgag ctggtgatat gggatagtgt tcacccttgt tacaccgttt
     2101 tccatgagca aactgaaacg ttttcatcgc tctggagtga ataccacgac gatttccggc
     2161 agtttctaca catatattcg caagatgtgg cgtgttacgg tgaaaacctg gcctatttcc
     2221 ctaaagggtt tattgagaat atgtttttcg tctcagccaa tccctgggtg agtttcacca
     2281 gttttgattt aaacgtggcc aatatggaca acttcttcgc ccccgttttc accatgggca
     2341 aatattatac gcaaggcgac aaggtgctga tgccgctggc gattcaggtt catcatgccg
     2401 tctgtgatgg cttccatgtc ggcagaatgc ttaatgaatt acaacagtac tgcgatgagt
     2461 ggcagggcgg ggcgtaaacg cgtggatccg gcttactaaa agccagataa cagtatgcgt
     2521 atttgcgcgc tgatttttgc ggtataagaa tatatactga tatgtatacc cgaagtatgt
     2581 caaaaagagg tgtgctatga agcagcgtat tacagtgaca gttgacagcg acagctatca
     2641 gttgctcaag gcatatatga tgtcaatatc tccggtctgg taagcacaac catgcagaat
     2701 gaagcccgtc gtctgcgtgc cgaacgctgg aaagcggaaa atcaggaagg gatggctgag
     2761 gtcgcccggt ttattgaaat gaacggctct tttgctgacg agaacaggga ctggtgaaat
     2821 gcagtttaag gtttacacct ataaaagaga gagccgttat cgtctgtttg tggatgtaca
     2881 gagtgatatt attgacacgc ccgggcgacg gatggtgatc cccctggcca gtgcacgtct
     2941 gctgtcagat aaagtctccc gtgaacttta cccggtggtg catatcgggg atgaaagctg
     3001 gcgcatgatg accaccgata tggccagtgt gccggtctcc gttatcgggg aagaagtggc
     3061 tgatctcagc caccgcgaaa atgacatcaa aaacgccatt aacctgatgt tctggggaat
     3121 ataaatgtca ggctccctta tacacagcca gtctgcaggt cgaccatagt gactggatat
     3181 gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt aatatattga
     3241 tatttatatc attttacgtt tctcgttcag ctttcttgta caaagtggtg ataacctaga
     3301 gctcgaattt ccccgatcgt tcaaacattt ggcaataaag tttcttaaga ttgaatcctg
     3361 ttgccggtct tgcgatgatt atcatataat ttctgttgaa ttacgttaag catgtaataa
     3421 ttaacatgta atgcatgacg ttatttatga gatgggtttt tatgattaga gtcccgcaat
     3481 tatacattta atacgcgata gaaaacaaaa tatagcgcgc aaactaggat aaattatcgc
     3541 gcgcggtgtc atctatgtta ctagatcggg aattctagct tcgacaatta ttttggactc
     3601 tcatgttctc aagctgaaat ccaaaagaaa cgtgatgttg gttcagattt ggaatcactg
     3661 acaatgaagt ttcagtcagc cggtttgtca ccaaggatcc aggcttcatc atgggagccg
     3721 agccgttcaa gtttcaactt cgactatttg gatccgtcct agtggatccc ccatcacaac
     3781 tttgtataga aaagttgaac gagaaacgta aaatgatata aatatcaata tattaaatta
     3841 gattttgcat aaaaaacaga ctacataata ctgtaaaaca caacatatcc agtcactatg
     3901 gcggccgcat taggcacccc aggctttaca ctttatgctt ccggctcgta taatgtgtgg
     3961 attttgagtt aggatccggc gagattttca ggagctaagg aagctaaaat ggagaaaaaa
     4021 atcactggat ataccaccgt tgatatatcc caatggcatc gtaaagaaca ttttgaggca
     4081 tttcagtcag ttgctcaatg tacctataac cagaccgttc agctggatat tacggccttt
     4141 ttaaagaccg taaagaaaaa taagcacaag ttttatccgg cctttattca cattcttgcc
     4201 cgcctgatga atgctcatcc ggaattgcga tcgcatttaa atggccggcc cttaattaag
     4261 gcgcgccgtt taaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     4321 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     4381 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     4441 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     4501 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     4561 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     4621 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tgcaggcatc
     4681 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     4741 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     4801 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     4861 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     4921 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aacacgggat
     4981 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     5041 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     5101 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     5161 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     5221 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     5281 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     5341 ccacctgaaa cgggcgacgg atggtgatcc ccctggccag tgcacgtctg ctgtcagata
     5401 aagtctcccg tgaactttac ccggtggtgc atatcgggga tgaaagctgg cgcatgatga
     5461 ccaccgatat ggccagtgtg ccggtctccg ttatcgggga agaagtggct gatctcagcc
     5521 accgcgaaaa tgacatcaaa aacgccatta acctgatgtt ctggggaata taaatgtcag
     5581 gctcccttat acacagccag tctgcaggtc gatcgaccat agtgactgga tatgttgtgt
     5641 tttacagtat tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat
     5701 atcattttac gtttctcgtt caactttatt atacaaagtt gtgatccaca tcttctgatt
     5761 ctattgatct ctcacagttt cagagaggtt tagttgacag gagttcttgt catcctcatg
     5821 taacactgga taaagtgtca aggacacatt tgagagtaga gcaaacaaat agccacatgc
     5881 ctgaaatgaa atcccaacag attacagata ctgaatttga tattcaaaat cacaaagaaa
     5941 gtgctgcttc tctgggaaaa ggaagagaat tgcttccatg ctctgctgca gagtcaataa
     6001 gcactgatgg aggtggtctg atttgttctg aggattcaaa ttggatggca tgttacaaga
     6061 attacccaac tttttaatac aaagttggca ttataagaaa gcattgctta tcaatttgtt
     6121 gcaacgaaca ggtcactatc agtcaaaata aaatcattat ttgccatcca gctgcagctc
     6181 tggcccgtgt ctcaaaatct ctgatgttac attgcacaag ataaaaatat atcatcatga
     6241 acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa
     6301 cgggaaacgt cgaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa
     6361 tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgcttgta tgggaagccc
     6421 gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat
     6481 gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt
     6541 atccgtactc ctgatgatgc atggttactc accactgcga tccccggaaa aacagcattc
     6601 caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc
     6661 ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt
     6721 cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat
     6781 gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa acttttgcca
     6841 ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac
     6901 gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag
     6961 gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt
     7021 tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc
     7081 gatgagtttt tctaatcaga attggttaat tggttgtaac actggcagag cattacgctg
     7141 acttgacggg acggcgcaag ctcatgacca aaatccctta acgtgagttt tcgttccact
     7201 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     7261 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     7321 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     7381 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     7441 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     7501 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     7561 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     7621 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     7681 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     7741 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     7801 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     7861 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     7921 accgtattac cgctagccag gaagagtttg tagaaacgca aaaaggccat ccgtcaggat
     7981 ggccttctgc ttagtttgat gcctggcagt ttatggcggg cgtcctgccc gccaccctcc
     8041 gggccgttgc ttcacaacgt tcaaatccgc tcccggcgga tttgtcctac tcaggagagc
     8101 gttcaccgac aaacaacaga taaaacgaaa ggcccagtct tccgactgag cctttcgttt
     8161 tatttgatgc ctggcagttc cctactctcg c
//
