LOCUS       40924_36623        8189 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway recycling vector pRED56-MD8-Pnos-17 DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8189)
  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  JOURNAL   Unpublished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 8189)
  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 8189)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8189)
  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            "Unpublished"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..8189
                     /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"
     protein_bind    1394..1518
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1555..1585
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             1639..2295
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             2640..2942
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(2986..3110)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             3136..3165
                     /codon_start=1
                     /label=Myc
                     /note="Myc (human c-Myc proto-oncogene) epitope tag"
                     /translation="EQKLISEEDL"
     CDS             3172..3201
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3217..3246
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3253..3282
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      3313..3565
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    3774..3898
                     /label=attR4
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3923..3953
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     misc_feature    4225..4265
                     /label=rare cutter sites
                     /note="rare cutter sites"
     CDS             complement(4357..5214)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5215..5319)
                     /label=AmpR promoter
     protein_bind    complement(5616..5739)
                     /label=attR3
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(6065..6160)
                     /label=attL6
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             6284..7090
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      7236..7824
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(7959..7986)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(8078..8164)
                     /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 tctagagtta tcaacaagtt tgtacaaaaa agctgaacga gaaacgtaaa atgatataaa
     1441 tatcaatata ttaaattaga ttttgcataa aaaacagact acataatact gtaaaacaca
     1501 acatatccag tcactatggc ggcccctcga ggggccgcat taggcacccc aggctttaca
     1561 ctttatgctt ccggctcgta taatgtgtgg attttgagtt aggatccggc gagattttca
     1621 ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt tgatatatcc
     1681 caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg tacctataac
     1741 cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa taagcacaag
     1801 ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc ggaattccgt
     1861 atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg ttacaccgtt
     1921 ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga cgatttccgg
     1981 cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc
     2041 cctaaagggt ttattgagaa tatgtttttc gtctcagcca atccctgggt gagtttcacc
     2101 agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt caccatgggc
     2161 aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt tcatcatgcc
     2221 gtctgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta ctgcgatgag
     2281 tggcagggcg gggcgtaaag atctggatcc ggcttactaa aagccagata acagtatgcg
     2341 tatttgcgcg ctgatttttg cggtataaga atatatactg atatgtatac ccgaagtatg
     2401 tcaaaaagag gtgtgctatg aagcagcgta ttacagtgac agttgacagc gacagctatc
     2461 agttgctcaa ggcatatatg atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa
     2521 tgaagcccgt cgtctgcgtg ccgaacgctg gaaagcggaa aatcaggaag ggatggctga
     2581 ggtcgcccgg tttattgaaa tgaacggctc ttttgctgac gagaacaggg actggtgaaa
     2641 tgcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     2701 agagtgatat tattgacacg cccgggcgac ggatggtgat ccccctggcc agtgcacgtc
     2761 tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     2821 ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg
     2881 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     2941 tataaatgtc aggctccctt atacacagcc agtctgcagg tcgaccatag tgactggata
     3001 tgttgtgttt tacagtatta tgtagtctgt tttttatgca aaatctaatt taatatattg
     3061 atatttatat cattttacgt ttctcgttca gctttcttgt acaaagtggt tgataacagc
     3121 gggttaatta acggtgaaca aaagctaatc tccgaggaag acttgaacgg tgaacaaaaa
     3181 ttaatctcag aagaagactt gaacggactc gacggtgaac aaaagttgat ttctgaagaa
     3241 gatttgaacg gtgaacaaaa gctaatctcc gaggaagact tgaacggtag cgcttagagc
     3301 tcgaatttcc ccgatcgttc aaacatttgg caataaagtt tcttaagatt gaatcctgtt
     3361 gccggtcttg cgatgattat catataattt ctgttgaatt acgttaagca tgtaataatt
     3421 aacatgtaat gcatgacgtt atttatgaga tgggttttta tgattagagt cccgcaatta
     3481 tacatttaat acgcgataga aaacaaaata tagcgcgcaa actaggataa attatcgcgc
     3541 gcggtgtcat ctatgttact agatcgggaa ttctagcttc gacaattatt ttggactctc
     3601 atgttctcaa gctgaaatcc aaaagaaacg tgatgttggt tcagatttgg aatcactgac
     3661 aatgaagttt cagtcagccg gtttgtcacc aaggatccag gcttcatcat gggagccgag
     3721 ccgttcaagt ttcaacttcg actatttgga tccgtcctag tggatccccc atcacaactt
     3781 tgtatagaaa agttgaacga gaaacgtaaa atgatataaa tatcaatata ttaaattaga
     3841 ttttgcataa aaaacagact acataatact gtaaaacaca acatatccag tcactatggc
     3901 ggccgcatta ggcaccccag gctttacact ttatgcttcc ggctcgtata atgtgtggat
     3961 tttgagttag gatccggcga gattttcagg agctaaggaa gctaaaatgg agaaaaaaat
     4021 cactggatat accaccgttg atatatccca atggcatcgt aaagaacatt ttgaggcatt
     4081 tcagtcagtt gctcaatgta cctataacca gaccgttcag ctggatatta cggccttttt
     4141 aaagaccgta aagaaaaata agcacaagtt ttatccggcc tttattcaca ttcttgcccg
     4201 cctgatgaat gctcatccgg aattgcgatc gcatttaaat ggccggccct taattaaggc
     4261 gcgccgttta aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     4321 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     4381 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     4441 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     4501 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     4561 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     4621 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgctg caggcatcgt
     4681 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     4741 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     4801 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     4861 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     4921 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa
     4981 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     5041 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     5101 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     5161 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     5221 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     5281 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     5341 acctgaaacg ggcgacggat ggtgatcccc ctggccagtg cacgtctgct gtcagataaa
     5401 gtctcccgtg aactttaccc ggtggtgcat atcggggatg aaagctggcg catgatgacc
     5461 accgatatgg ccagtgtgcc ggtctccgtt atcggggaag aagtggctga tctcagccac
     5521 cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct ggggaatata aatgtcaggc
     5581 tcccttatac acagccagtc tgcaggtcga tcgaccatag tgactggata tgttgtgttt
     5641 tacagtatta tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat
     5701 cattttacgt ttctcgttca actttattat acaaagttgt gatccacatc ttctgattct
     5761 attgatctct cacagtttca gagaggttta gttgacagga gttcttgtca tcctcatgta
     5821 acactggata aagtgtcaag gacacatttg agagtagagc aaacaaatag ccacatgcct
     5881 gaaatgaaat cccaacagat tacagatact gaatttgata ttcaaaatca caaagaaagt
     5941 gctgcttctc tgggaaaagg aagagaattg cttccatgct ctgctgcaga gtcaataagc
     6001 actgatggag gtggtctgat ttgttctgag gattcaaatt ggatggcatg ttacaagaat
     6061 tacccaactt tttaatacaa agttggcatt ataagaaagc attgcttatc aatttgttgc
     6121 aacgaacagg tcactatcag tcaaaataaa atcattattt gccatccagc tgcagctctg
     6181 gcccgtgtct caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac
     6241 aataaaactg tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg
     6301 ggaaacgtcg aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg
     6361 ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg ggaagcccga
     6421 tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga
     6481 gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat
     6541 ccgtactcct gatgatgcat ggttactcac cactgcgatc cccggaaaaa cagcattcca
     6601 ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct
     6661 gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg
     6721 tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga
     6781 cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt
     6841 ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga
     6901 ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga
     6961 tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt
     7021 tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga
     7081 tgagtttttc taatcagaat tggttaattg gttgtaacac tggcagagca ttacgctgac
     7141 ttgacgggac ggcgcaagct catgaccaaa atcccttaac gtgagttttc gttccactga
     7201 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     7261 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     7321 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     7381 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     7441 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     7501 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     7561 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     7621 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     7681 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     7741 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     7801 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     7861 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     7921 cgtattaccg ctagccagga agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg
     7981 ccttctgctt agtttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg
     8041 gccgttgctt cacaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt
     8101 tcaccgacaa acaacagata aaacgaaagg cccagtcttc cgactgagcc tttcgtttta
     8161 tttgatgcct ggcagttccc tactctcgc
//
