LOCUS       40924_36638        8425 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway recycling vector pRED56-MD8-Pnos-21 DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8425)
  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  JOURNAL   Unpublished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 8425)
  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 8425)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8425)
  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            "Unpublished"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..8425
                     /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"
     CDS             1561..1590
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1606..1635
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1642..1671
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1678..1707
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1723..1752
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1759..1788
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    1807..1931
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1968..1998
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             2052..2708
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             3053..3355
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(3399..3523)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      3549..3801
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    4010..4134
                     /label=attR4
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        4159..4189
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     misc_feature    4461..4501
                     /label=rare cutter sites
                     /note="rare cutter sites"
     CDS             complement(4593..5450)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5451..5555)
                     /label=AmpR promoter
     protein_bind    complement(5852..5975)
                     /label=attR3
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(6301..6396)
                     /label=attL6
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             6520..7326
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      7472..8060
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(8195..8222)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(8314..8400)
                     /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 gaacaaaaat taatctcaga agaagacttg aacggactcg acggtgaaca aaagttgatt
     1621 tctgaagaag atttgaacgg tgaacaaaag ctaatctccg aggaagactt gaacggtgaa
     1681 caaaaattaa tctcagaaga agacttgaac ggactcgacg gtgaacaaaa gttgatttct
     1741 gaagaagatt tgaacggtga acaaaagcta atctccgagg aagacttgaa cggtagcgct
     1801 gttatcacaa gtttgtacaa aaaagctgaa cgagaaacgt aaaatgatat aaatatcaat
     1861 atattaaatt agattttgca taaaaaacag actacataat actgtaaaac acaacatatc
     1921 cagtcactat ggcggcccct cgaggggccg cattaggcac cccaggcttt acactttatg
     1981 cttccggctc gtataatgtg tggattttga gttaggatcc ggcgagattt tcaggagcta
     2041 aggaagctaa aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc
     2101 atcgtaaaga acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg
     2161 ttcagctgga tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc
     2221 cggcctttat tcacattctt gcccgcctga tgaatgctca tccggaattc cgtatggcaa
     2281 tgaaagacgg tgagctggtg atatgggata gtgttcaccc ttgttacacc gttttccatg
     2341 agcaaactga aacgttttca tcgctctgga gtgaatacca cgacgatttc cggcagtttc
     2401 tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag
     2461 ggtttattga gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc accagttttg
     2521 atttaaacgt ggccaatatg gacaacttct tcgcccccgt tttcaccatg ggcaaatatt
     2581 atacgcaagg cgacaaggtg ctgatgccgc tggcgattca ggttcatcat gccgtctgtg
     2641 atggcttcca tgtcggcaga atgcttaatg aattacaaca gtactgcgat gagtggcagg
     2701 gcggggcgta aacgcgtgga tccggcttac taaaagccag ataacagtat gcgtatttgc
     2761 gcgctgattt ttgcggtata agaatatata ctgatatgta tacccgaagt atgtcaaaaa
     2821 gaggtgtgct atgaagcagc gtattacagt gacagttgac agcgacagct atcagttgct
     2881 caaggcatat atgatgtcaa tatctccggt ctggtaagca caaccatgca gaatgaagcc
     2941 cgtcgtctgc gtgccgaacg ctggaaagcg gaaaatcagg aagggatggc tgaggtcgcc
     3001 cggtttattg aaatgaacgg ctcttttgct gacgagaaca gggactggtg aaatgcagtt
     3061 taaggtttac acctataaaa gagagagccg ttatcgtctg tttgtggatg tacagagtga
     3121 tattattgac acgcccgggc gacggatggt gatccccctg gccagtgcac gtctgctgtc
     3181 agataaagtc tcccgtgaac tttacccggt ggtgcatatc ggggatgaaa gctggcgcat
     3241 gatgaccacc gatatggcca gtgtgccggt ctccgttatc ggggaagaag tggctgatct
     3301 cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg atgttctggg gaatataaat
     3361 gtcaggctcc cttatacaca gccagtctgc aggtcgacca tagtgactgg atatgttgtg
     3421 ttttacagta ttatgtagtc tgttttttat gcaaaatcta atttaatata ttgatattta
     3481 tatcatttta cgtttctcgt tcagctttct tgtacaaagt ggtgataacc tagagctcga
     3541 atttccccga tcgttcaaac atttggcaat aaagtttctt aagattgaat cctgttgccg
     3601 gtcttgcgat gattatcata taatttctgt tgaattacgt taagcatgta ataattaaca
     3661 tgtaatgcat gacgttattt atgagatggg tttttatgat tagagtcccg caattataca
     3721 tttaatacgc gatagaaaac aaaatatagc gcgcaaacta ggataaatta tcgcgcgcgg
     3781 tgtcatctat gttactagat cgggaattct agcttcgaca attattttgg actctcatgt
     3841 tctcaagctg aaatccaaaa gaaacgtgat gttggttcag atttggaatc actgacaatg
     3901 aagtttcagt cagccggttt gtcaccaagg atccaggctt catcatggga gccgagccgt
     3961 tcaagtttca acttcgacta tttggatccg tcctagtgga tcccccatca caactttgta
     4021 tagaaaagtt gaacgagaaa cgtaaaatga tataaatatc aatatattaa attagatttt
     4081 gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac tatggcggcc
     4141 gcattaggca ccccaggctt tacactttat gcttccggct cgtataatgt gtggattttg
     4201 agttaggatc cggcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact
     4261 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag
     4321 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag
     4381 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg
     4441 atgaatgctc atccggaatt gcgatcgcat ttaaatggcc ggcccttaat taaggcgcgc
     4501 cgtttaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4561 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4621 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4681 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4741 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4801 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4861 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg
     4921 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4981 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5041 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5101 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5161 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg ggataatacc
     5221 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5281 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5341 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5401 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5461 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5521 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5581 gaaacgggcg acggatggtg atccccctgg ccagtgcacg tctgctgtca gataaagtct
     5641 cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg
     5701 atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg
     5761 aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggctccc
     5821 ttatacacag ccagtctgca ggtcgatcga ccatagtgac tggatatgtt gtgttttaca
     5881 gtattatgta gtctgttttt tatgcaaaat ctaatttaat atattgatat ttatatcatt
     5941 ttacgtttct cgttcaactt tattatacaa agttgtgatc cacatcttct gattctattg
     6001 atctctcaca gtttcagaga ggtttagttg acaggagttc ttgtcatcct catgtaacac
     6061 tggataaagt gtcaaggaca catttgagag tagagcaaac aaatagccac atgcctgaaa
     6121 tgaaatccca acagattaca gatactgaat ttgatattca aaatcacaaa gaaagtgctg
     6181 cttctctggg aaaaggaaga gaattgcttc catgctctgc tgcagagtca ataagcactg
     6241 atggaggtgg tctgatttgt tctgaggatt caaattggat ggcatgttac aagaattacc
     6301 caacttttta atacaaagtt ggcattataa gaaagcattg cttatcaatt tgttgcaacg
     6361 aacaggtcac tatcagtcaa aataaaatca ttatttgcca tccagctgca gctctggccc
     6421 gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata
     6481 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa
     6541 acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct
     6601 cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg
     6661 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg
     6721 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt
     6781 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta
     6841 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc
     6901 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc
     6961 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag
     7021 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca
     7081 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg
     7141 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt
     7201 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa
     7261 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag
     7321 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga
     7381 cgggacggcg caagctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     7441 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     7501 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     7561 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     7621 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     7681 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     7741 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     7801 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     7861 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7921 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7981 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     8041 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     8101 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     8161 ttaccgctag ccaggaagag tttgtagaaa cgcaaaaagg ccatccgtca ggatggcctt
     8221 ctgcttagtt tgatgcctgg cagtttatgg cgggcgtcct gcccgccacc ctccgggccg
     8281 ttgcttcaca acgttcaaat ccgctcccgg cggatttgtc ctactcagga gagcgttcac
     8341 cgacaaacaa cagataaaac gaaaggccca gtcttccgac tgagcctttc gttttatttg
     8401 atgcctggca gttccctact ctcgc
//
