LOCUS       40924_36653        8051 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway recycling vector pRED56-MD8-Pnos-26 DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8051)
  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  JOURNAL   Unpublished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 8051)
  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 8051)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8051)
  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            "Unpublished"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..8051
                     /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             3121..3153
                     /codon_start=1
                     /label=T7 tag (gene 10 leader)
                     /note="leader peptide from bacteriophage T7 gene 10"
                     /translation="MASMTGGQQMG"
     terminator      3175..3427
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    3636..3760
                     /label=attR4
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3785..3815
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     misc_feature    4087..4127
                     /label=rare cutter sites
                     /note="rare cutter sites"
     CDS             complement(4219..5076)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5077..5181)
                     /label=AmpR promoter
     protein_bind    complement(5478..5601)
                     /label=attR3
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(5927..6022)
                     /label=attL6
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             6146..6952
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      7098..7686
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(7821..7848)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(7940..8026)
                     /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 atggctagca tgactggtgg acagcaaatg ggtgcttaga gctcgaattt ccccgatcgt
     3181 tcaaacattt ggcaataaag tttcttaaga ttgaatcctg ttgccggtct tgcgatgatt
     3241 atcatataat ttctgttgaa ttacgttaag catgtaataa ttaacatgta atgcatgacg
     3301 ttatttatga gatgggtttt tatgattaga gtcccgcaat tatacattta atacgcgata
     3361 gaaaacaaaa tatagcgcgc aaactaggat aaattatcgc gcgcggtgtc atctatgtta
     3421 ctagatcggg aattctagct tcgacaatta ttttggactc tcatgttctc aagctgaaat
     3481 ccaaaagaaa cgtgatgttg gttcagattt ggaatcactg acaatgaagt ttcagtcagc
     3541 cggtttgtca ccaaggatcc aggcttcatc atgggagccg agccgttcaa gtttcaactt
     3601 cgactatttg gatccgtcct agtggatccc ccatcacaac tttgtataga aaagttgaac
     3661 gagaaacgta aaatgatata aatatcaata tattaaatta gattttgcat aaaaaacaga
     3721 ctacataata ctgtaaaaca caacatatcc agtcactatg gcggccgcat taggcacccc
     3781 aggctttaca ctttatgctt ccggctcgta taatgtgtgg attttgagtt aggatccggc
     3841 gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt
     3901 tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg
     3961 tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa
     4021 taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc
     4081 ggaattgcga tcgcatttaa atggccggcc cttaattaag gcgcgccgtt taaggatctt
     4141 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     4201 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     4261 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     4321 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     4381 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     4441 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     4501 taatagtttg cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac gctcgtcgtt
     4561 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     4621 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     4681 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     4741 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     4801 gcggcgaccg agttgctctt gcccggcgtc aacacgggat aataccgcgc cacatagcag
     4861 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     4921 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     4981 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     5041 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     5101 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     5161 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgaaa cgggcgacgg
     5221 atggtgatcc ccctggccag tgcacgtctg ctgtcagata aagtctcccg tgaactttac
     5281 ccggtggtgc atatcgggga tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg
     5341 ccggtctccg ttatcgggga agaagtggct gatctcagcc accgcgaaaa tgacatcaaa
     5401 aacgccatta acctgatgtt ctggggaata taaatgtcag gctcccttat acacagccag
     5461 tctgcaggtc gatcgaccat agtgactgga tatgttgtgt tttacagtat tatgtagtct
     5521 gttttttatg caaaatctaa tttaatatat tgatatttat atcattttac gtttctcgtt
     5581 caactttatt atacaaagtt gtgatccaca tcttctgatt ctattgatct ctcacagttt
     5641 cagagaggtt tagttgacag gagttcttgt catcctcatg taacactgga taaagtgtca
     5701 aggacacatt tgagagtaga gcaaacaaat agccacatgc ctgaaatgaa atcccaacag
     5761 attacagata ctgaatttga tattcaaaat cacaaagaaa gtgctgcttc tctgggaaaa
     5821 ggaagagaat tgcttccatg ctctgctgca gagtcaataa gcactgatgg aggtggtctg
     5881 atttgttctg aggattcaaa ttggatggca tgttacaaga attacccaac tttttaatac
     5941 aaagttggca ttataagaaa gcattgctta tcaatttgtt gcaacgaaca ggtcactatc
     6001 agtcaaaata aaatcattat ttgccatcca gctgcagctc tggcccgtgt ctcaaaatct
     6061 ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta
     6121 cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cgaggccgcg
     6181 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
     6241 gcaatcaggt gcgacaatct atcgcttgta tgggaagccc gatgcgccag agttgtttct
     6301 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg
     6361 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
     6421 atggttactc accactgcga tccccggaaa aacagcattc caggtattag aagaatatcc
     6481 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
     6541 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc
     6601 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
     6661 tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg attcagtcgt
     6721 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
     6781 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     6841 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     6901 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaatcaga
     6961 attggttaat tggttgtaac actggcagag cattacgctg acttgacggg acggcgcaag
     7021 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     7081 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     7141 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     7201 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     7261 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     7321 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     7381 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     7441 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     7501 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     7561 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     7621 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     7681 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     7741 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgctagccag
     7801 gaagagtttg tagaaacgca aaaaggccat ccgtcaggat ggccttctgc ttagtttgat
     7861 gcctggcagt ttatggcggg cgtcctgccc gccaccctcc gggccgttgc ttcacaacgt
     7921 tcaaatccgc tcccggcgga tttgtcctac tcaggagagc gttcaccgac aaacaacaga
     7981 taaaacgaaa ggcccagtct tccgactgag cctttcgttt tatttgatgc ctggcagttc
     8041 cctactctcg c
//
