LOCUS       40924_36563        8258 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway recycling vector pRED56-MD8-NY2 DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8258)
  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  JOURNAL   Unpublished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 8258)
  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 8258)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8258)
  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            "Unpublished"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..8258
                     /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"
     protein_bind    1109..1233
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1270..1300
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             1354..2010
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             2355..2657
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(2701..2825)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2836..3354
                     /codon_start=1
                     /label=VN173
                     /note="N-terminal fragment of mVenus for use in bimolecular
                     fluorescence complementation (BiFC) (Kodama and Hu, 2010)"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     terminator      3382..3634
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    3843..3967
                     /label=attR4
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3992..4022
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     misc_feature    4294..4334
                     /label=rare cutter sites
                     /note="rare cutter sites"
     CDS             complement(4426..5283)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5284..5388)
                     /label=AmpR promoter
     protein_bind    complement(5685..5808)
                     /label=attR3
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(6134..6229)
                     /label=attL6
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             6353..7159
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      7305..7893
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(8028..8055)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(8147..8233)
                     /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 taagcttgca
     1081 tgcctgcagg tcgactctag agttatcaac aagtttgtac aaaaaagctg aacgagaaac
     1141 gtaaaatgat ataaatatca atatattaaa ttagattttg cataaaaaac agactacata
     1201 atactgtaaa acacaacata tccagtcact atggcggccc ctcgaggggc cgcattaggc
     1261 accccaggct ttacacttta tgcttccggc tcgtataatg tgtggatttt gagttaggat
     1321 ccgtcgagat tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc
     1381 accgttgata tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct
     1441 caatgtacct ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag
     1501 aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct
     1561 catccggaat tccgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac
     1621 ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac
     1681 cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa
     1741 aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc
     1801 tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc
     1861 gttttcacca tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt
     1921 caggttcatc atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa
     1981 cagtactgcg atgagtggca gggcggggcg taaagatctg gatccggctt actaaaagcc
     2041 agataacagt atgcgtattt gcgcgctgat ttttgcggta taagaatata tactgatatg
     2101 tatacccgaa gtatgtcaaa aagaggtatg ctatgaagca gcgtattaca gtgacagttg
     2161 acagcgacag ctatcagttg ctcaaggcat atatgatgtc aatatctccg gtctggtaag
     2221 cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa cgctggaaag cggaaaatca
     2281 ggaagggatg gctgaggtcg cccggtttat tgaaatgaac ggctcttttg ctgacgagaa
     2341 caggggctgg tgaaatgcag tttaaggttt acacctataa aagagagagc cgttatcgtc
     2401 tgtttgtgga tgtacagagt gatattattg acacgcccgg gcgacggatg gtgatccccc
     2461 tggccagtgc acgtctgctg tcagataaag tctcccgtga actttacccg gtggtgcata
     2521 tcggggatga aagctggcgc atgatgacca ccgatatggc cagtgtgccg gtctccgtta
     2581 tcggggaaga agtggctgat ctcagccacc gcgaaaatga catcaaaaac gccattaacc
     2641 tgatgttctg gggaatataa atgtcaggct cccttataca cagccagtct gcaggtcgac
     2701 catagtgact ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc
     2761 taatttaata tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa
     2821 gtggttgata acagcatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     2881 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
     2941 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     3001 gtgccctggc ccaccctcgt gaccaccttc ggctacggcc tgcagtgctt cgcccgctac
     3061 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
     3121 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     3181 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     3241 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc
     3301 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggactag
     3361 gcttagagct cgaatttccc cgatcgttca aacatttggc aataaagttt cttaagattg
     3421 aatcctgttg ccggtcttgc gatgattatc atataatttc tgttgaatta cgttaagcat
     3481 gtaataatta acatgtaatg catgacgtta tttatgagat gggtttttat gattagagtc
     3541 ccgcaattat acatttaata cgcgatagaa aacaaaatat agcgcgcaaa ctaggataaa
     3601 ttatcgcgcg cggtgtcatc tatgttacta gatcgggaat tctagcttcg acaattattt
     3661 tggactctca tgttctcaag ctgaaatcca aaagaaacgt gatgttggtt cagatttgga
     3721 atcactgaca atgaagtttc agtcagccgg tttgtcacca aggatccagg cttcatcatg
     3781 ggagccgagc cgttcaagtt tcaacttcga ctatttggat ccgtcctagt ggatccccca
     3841 tcacaacttt gtatagaaaa gttgaacgag aaacgtaaaa tgatataaat atcaatatat
     3901 taaattagat tttgcataaa aaacagacta cataatactg taaaacacaa catatccagt
     3961 cactatggcg gccgcattag gcaccccagg ctttacactt tatgcttccg gctcgtataa
     4021 tgtgtggatt ttgagttagg atccggcgag attttcagga gctaaggaag ctaaaatgga
     4081 gaaaaaaatc actggatata ccaccgttga tatatcccaa tggcatcgta aagaacattt
     4141 tgaggcattt cagtcagttg ctcaatgtac ctataaccag accgttcagc tggatattac
     4201 ggccttttta aagaccgtaa agaaaaataa gcacaagttt tatccggcct ttattcacat
     4261 tcttgcccgc ctgatgaatg ctcatccgga attgcgatcg catttaaatg gccggccctt
     4321 aattaaggcg cgccgtttaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4381 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4441 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     4501 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     4561 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     4621 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     4681 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctgc
     4741 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     4801 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     4861 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     4921 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     4981 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaac
     5041 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5101 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5161 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     5221 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     5281 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     5341 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     5401 aaaagtgcca cctgaaacgg gcgacggatg gtgatccccc tggccagtgc acgtctgctg
     5461 tcagataaag tctcccgtga actttacccg gtggtgcata tcggggatga aagctggcgc
     5521 atgatgacca ccgatatggc cagtgtgccg gtctccgtta tcggggaaga agtggctgat
     5581 ctcagccacc gcgaaaatga catcaaaaac gccattaacc tgatgttctg gggaatataa
     5641 atgtcaggct cccttataca cagccagtct gcaggtcgat cgaccatagt gactggatat
     5701 gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt aatatattga
     5761 tatttatatc attttacgtt tctcgttcaa ctttattata caaagttgtg atccacatct
     5821 tctgattcta ttgatctctc acagtttcag agaggtttag ttgacaggag ttcttgtcat
     5881 cctcatgtaa cactggataa agtgtcaagg acacatttga gagtagagca aacaaatagc
     5941 cacatgcctg aaatgaaatc ccaacagatt acagatactg aatttgatat tcaaaatcac
     6001 aaagaaagtg ctgcttctct gggaaaagga agagaattgc ttccatgctc tgctgcagag
     6061 tcaataagca ctgatggagg tggtctgatt tgttctgagg attcaaattg gatggcatgt
     6121 tacaagaatt acccaacttt ttaatacaaa gttggcatta taagaaagca ttgcttatca
     6181 atttgttgca acgaacaggt cactatcagt caaaataaaa tcattatttg ccatccagct
     6241 gcagctctgg cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc
     6301 atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     6361 tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg
     6421 gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg
     6481 gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt
     6541 tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa
     6601 gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac
     6661 agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc
     6721 agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg
     6781 cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga
     6841 ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact
     6901 tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat
     6961 ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg
     7021 ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa
     7081 acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt
     7141 gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat
     7201 tacgctgact tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg tgagttttcg
     7261 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     7321 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     7381 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     7441 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     7501 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     7561 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     7621 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     7681 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     7741 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     7801 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     7861 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     7921 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     7981 gtggataacc gtattaccgc tagccaggaa gagtttgtag aaacgcaaaa aggccatccg
     8041 tcaggatggc cttctgctta gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc
     8101 accctccggg ccgttgcttc acaacgttca aatccgctcc cggcggattt gtcctactca
     8161 ggagagcgtt caccgacaaa caacagataa aacgaaaggc ccagtcttcc gactgagcct
     8221 ttcgttttat ttgatgcctg gcagttccct actctcgc
//
