LOCUS       40924_31270        6126 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway entry vector pMpGE_En01 DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6126)
  AUTHORS   Sugano SS, Nishihama R, Shirakawa M, Matsuda Y, Takagi T, 
            Hara-Nishimura I, Osakabe K, Kohchi T.
  TITLE     Efficient CRISPR/Cas9 vectors in Marchantia polymorpha
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6126)
  AUTHORS   Sugano SS, Kohchi T, Nishihama R.
  TITLE     Direct Submission
  JOURNAL   Submitted (14-OCT-2015) Contact:Shigeo S Sugano Tokushima 
            University, Center for Collaboration among Agriculture, Industry, 
            and Commerse; Kuramoto cho 3-18-15, Tokushima, Tokushima 770-8503, 
            Japan URL :http://www.tokushima-u.ac.jp/ccaic/
REFERENCE   3  (bases 1 to 6126)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6126)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (14-OCT-2015) Contact:Shigeo S Sugano Tokushima University, Center 
            for Collaboration among Agriculture, Industry, and Commerse"; 
            volume: " Kuramoto cho 3-18-15, Tokushima, Tokushima 770-8503, Japan
            URL :http"; pages: "//www.tokushima-u.ac.jp/ccaic"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6126
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(387..473)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    569..668
                     /label=attL1
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    689..2699
                     /label=MpU6-1pro
                     /note="MpU6-1pro"
     misc_feature    2700..2705
                     /label=SacI
                     /note="SacI"
     promoter        2722..2752
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             2806..3483
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     CDS             3806..4108
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     misc_feature    4144..4148
                     /label=PstI
                     /note="PstI"
     misc_RNA        4149..4224
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(4251..4350)
                     /label=attL2
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(4368..4386)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4391..4407)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4520..5326
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      5476..6064
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctcc gcggccgccc ccttcaccta ttcattcaaa agagattttt aaagatcaat
      721 ttttctttct cataagtcta atacgtcatg ataataactt taaaaaaaat tgaactcgtt
      781 aaaaaatttg atcaaatttt aatttattat tagatcatat attacatatc agtttcataa
      841 tgttatacta agatctttca tatatatttt aacccacgag acaaaatatg tttatatctt
      901 tatttattta aatatttttt agctcaaaac atcgcaaact cagctatcat aatgcttttt
      961 aatgaaaaaa acaaattttc ctatcatact ctcattgttt ataatttttt agaaaaaaaa
     1021 cttgaagata aaataacaac aataatagtt tcaaaagcac atttttgttg taaatttcaa
     1081 gaaaaataca atcatattac tgattgacta caaaaatttg taaattaatc caggctacat
     1141 gtttaagatg tgtcaaaact gcatcatcta catctcattt taatttttct aaactacata
     1201 catagtatca ataacaattg aacaatgcta gttaatgtgt tatactcatt agcttgtaat
     1261 atatcttcaa cttgatgaat cggattgatc ataaaatttc acaaacttat gtgctgatta
     1321 ataaaaaata taatcagatt tttcatgaat aattaagata aataacacat gcaagtctca
     1381 attagaattt aacaatctga ctataaaaag tcggtttatc ttgattttca caaaaattta
     1441 acatagtttc aaggtgaatg ggcttttatt ttaatccaat caactatttg acacatacat
     1501 cttgcaaatt tatttatttt ttatggaaaa aaacatgatt tatgtttttc tttgagaaag
     1561 tatattgtgg attgtcaaaa aaaacctatg gaagatttat atgtaagagt taaaaattta
     1621 ttttcaaata tgtcaacaat ccacaatatc gaagaaaatt ttcaaaaaat catcacatat
     1681 gatgtaaata taatatgaat atgtgcaatc ttatttcatt ttactctgaa aacttaattt
     1741 ctttttatag agcattctgt acttactggt ctcaaatata cagataaaat gcagtccata
     1801 acgtatcact ctataagcaa aatcataaac ttcgtgatct ctcacagttc aaacgagaac
     1861 tatagaaacc cttcaactag caaaaaactc acaactttct attccaatat ctttaattgt
     1921 ctgacgattt acaaatagaa acgaaaacca gacacacaca cacacgcaaa ctacaaagta
     1981 aaaacacaac cacctgacat tagacagtga gattattaaa catcctaaac taaaaaacaa
     2041 acaaattaat tctctatgta caagtcccca tcgtgatcaa tttttgcatc ccaatatagt
     2101 ggcggttagt ataagaggac cttcgtctat tcactctcca tgtttgtcaa actatagtac
     2161 agtacatata tgtatgggaa caaattccga acactctagt acactcctca acgcaagttc
     2221 tatctcatat ctctgcaaga gcttattcca agaaactttc ttgtgcttct atgcatgccc
     2281 aggtacctta agtcgcgcgc atagtcactt cacagccttt tcagttctcc ctttgatggc
     2341 tcagagtgac gcaactgcac aaaaataaga aaaaagtcga agcgtcacga gagatgtatc
     2401 cggctagttt ccttggagtt cggattgctc tctttctctg tctccccgaa ccatatggaa
     2461 tttatgaact gctgttcact agtcagtcac ttgtttcaga acttttgagt ggtcgattta
     2521 tccaccttct tgtttcttcc tgggtccaga atctgacttc ttgagacaac aaacagtgca
     2581 cagtggatcg aagaatgacc acagttgaga cacgtggcgc tggcctcatt tattgcacca
     2641 tacgtttagt gaagaactga ataatggtgg atataaagca gagttgcacc cagcctctcg
     2701 agctcattag gcaccccagg ctttacactt tatgcttccg gctcgtataa tgtgtggatt
     2761 ttgagttagg atccgtcgag attttcagga gctaaggaag ctaaaatgga gaaaaaaatc
     2821 actggatata ccaccgttga tatatcccaa tggcatcgta aagaacattt tgaggcattt
     2881 cagtcagttg ctcaatgtac ctataaccag accgttcagc tggatattac ggccttttta
     2941 aagaccgtaa agaaaaataa gcacaagttt tatccggcct ttattcacat tcttgcccgc
     3001 ctgatgaatg ctcatccgga attccgtatg gcaatgaaag acggtgagct ggtgatatgg
     3061 gatagtgttc acccttgtta caccgttttc catgagcaaa ctgaaacgtt ttcatcgctc
     3121 tggagtgaat accacgacga tttccggcag tttctacaca tatattcgca agatgtggcg
     3181 tgttacggtg aaaacctggc ctatttccct aaagggttta ttgagaatat gtttttcgtc
     3241 tcagccaatc cctgggtgag tttcaccagt tttgatttaa acgtggccaa tatggacaac
     3301 ttcttcgccc ccgttttcac catgggcaaa tattatacgc aaggcgacaa ggtgctgatg
     3361 ccgctggcga ttcaggttca tcatgccgtt tgtgatggct tccatgtcgg cagaatgctt
     3421 aatgaattac aacagtactg cgatgagtgg caggcggggc gtaatctaga ggatccggct
     3481 tactaaaagc cagataacag tatgcgtatt tgcgcgctga tttttgcggt ataagaatat
     3541 atactgatat gtatacccga agtatgtcaa aaagaggtat gctatgaagc agcgtattac
     3601 agtgacagtt gacagcgaca gctatcagtt gctcaaggca tatatgatgt caatatctcc
     3661 ggtctggtaa gcacaaccat gcagaatgaa gcccgtcgtc tgcgtgccga acgctggaaa
     3721 gcggaaaatc aggaagggat ggctgaggtc gcccggttta ttgaaatgaa cggctctttt
     3781 gctgacgaga acaggggctg gtgaaatgca gtttaaggtt tacacctata aaagagagag
     3841 ccgttatcgt ctgtttgtgg atgtacagag tgatattatt gacacgcccg ggcgacggat
     3901 ggtgatcccc ctggccagtg cacgtctgct gtcagataaa gtcccccgtg aactttaccc
     3961 ggtggtgcat atcggggatg aaagctggcg catgatgacc accgatatgg ccagtgtgcc
     4021 ggtctccgtt atcggggaag aagtggctga tctcagccac cgcgaaaatg acatcaaaaa
     4081 cgccattaac ctgatgttct ggggaatata aatgtcaggc tcccttatac acagccagtc
     4141 tgcctgcagt tttagagcta gaaatagcaa gttaaaataa ggctagtccg ttatcaactt
     4201 gaaaaagtgg caccgagtcg gtgctttttt tctaaagggt gggcgcgccg acccagcttt
     4261 cttgtacaaa gttggcatta taagaaagca ttgcttatca atttgttgca acgaacaggt
     4321 cactatcagt caaaataaaa tcattatttg ccatccagct gatatcccct atagtgagtc
     4381 gtattacatg gtcatagctg tttcctggca gctctggccc gtgtctcaaa atctctgatg
     4441 ttacattgca caagataaaa atatatcatc atgaacaata aaactgtctg cttacataaa
     4501 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcgaggc cgcgattaaa
     4561 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     4621 aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca
     4681 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     4741 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     4801 actcaccact gcgatccccg gaaaaacagc attccaggta ttagaagaat atcctgattc
     4861 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     4921 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     4981 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     5041 acaagtctgg aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca
     5101 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     5161 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     5221 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     5281 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagaattggt
     5341 taattggttg taacactggc agagcattac gctgacttga cgggacggcg caagctcatg
     5401 accaaaatcc cttaacgtga gttacgcgtc gttccactga gcgtcagacc ccgtagaaaa
     5461 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     5521 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     5581 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     5641 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     5701 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     5761 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     5821 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagcatt gagaaagcgc
     5881 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     5941 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     6001 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     6061 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     6121 catgtt
//
