LOCUS       40924_27369        5694 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pL0D_4-R, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5694)
  AUTHORS   Hochrein L, Machens F, Gremmels J, Schulz K, Messerschmidt K, 
            Mueller-Roeber B.
  TITLE     AssemblX: a user-friendly toolkit for rapid and reliable multi-gene 
            assemblies
  JOURNAL   Nucleic Acids Res. (2017) In press
  PUBMED    28130422
REFERENCE   2  (bases 1 to 5694)
  AUTHORS   Hochrein L, Machens F, Gremmels J, Schulz K, Messerschmidt K, 
            Mueller-Roeber B.
  TITLE     Direct Submission
  JOURNAL   Submitted (11-NOV-2016) Department of Molecular Biology - Cell2Fab 
            research unit, University of Potsdam, Karl-Liebknecht-Str. 24-25, 
            Potsdam, Brandenburg 14476, Germany
REFERENCE   3  (bases 1 to 5694)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5694)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. (2017) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (11-NOV-2016) Department of Molecular Biology - Cell2Fab research 
            unit, University of Potsdam, Karl-Liebknecht-Str. 24-25, Potsdam, 
            Brandenburg 14476, Germany"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(575..1375)
                     /codon_start=1
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(1376..1596)
                     /label=URA3 promoter
     rep_origin      complement(2195..3075)
                     /direction=LEFT
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(3424..4215)
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHDDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        complement(4216..4320)
                     /label=AmpR promoter
     misc_feature    4389..4428
                     /label=8 cutter set: AscI, SbfI, SwaI, FseI, PmeI
                     /note="8 cutter set: AscI, SbfI, SwaI, FseI, PmeI"
     misc_feature    4429..4464
                     /label=homology region D4*
                     /note="homology region D4*"
     CDS             4800..5102
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     promoter        5194..5433
                     /label=LYS2 promoter
     misc_feature    5434..5484
                     /label=homology region DR*
                     /note="homology region DR*"
     misc_feature    5488..5527
                     /label=8 cutter set: SwaI, AscI, PmeI, SbfI, FseI
                     /note="8 cutter set: SwaI, AscI, PmeI, SbfI, FseI"
     rep_origin      5553..5694
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
       61 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
      121 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
      181 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
      241 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
      301 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
      361 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
      421 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
      481 gccttttgct ggccttttgc tcatgatata attaaattga agctctaatt tgtgagttta
      541 gtatacatgc atttacttat aatacagttt tttagttttg ctggccgcat cttctcaaat
      601 atgcttccca gcctgctttt ctgtaacgtt caccctctac cttagcatcc cttccctttg
      661 caaatagtcc tcttccaaca ataataatgt cagatcctgt agagaccaca tcatccacgg
      721 ttctatactg ttgacccaat gcgtctccct tgtcatctaa acccacaccg ggtgtcataa
      781 tcaaccaatc gtaaccttca tctcttccac ccatgtctct ttgagcaata aagccgataa
      841 caaaatcttt gtcgctcttc gcaatgtcaa cagtaccctt agtatattct ccagtagata
      901 gggagccctt gcatgacaat tctgctaaca tcaaaaggcc tctaggttcc tttgttactt
      961 cttctgccgc ctgcttcaaa ccgctaacaa tacctgggcc caccacaccg tgtgcattcg
     1021 taatgtctgc ccattctgct attctgtata cacccgcaga gtactgcaat ttgactgtat
     1081 taccaatgtc agcaaatttt ctgtcttcga agagtaaaaa attgtacttg gcggataatg
     1141 cctttagcgg cttaactgtg ccctccatgg aaaaatcagt caagatatcc acatgtgttt
     1201 ttagtaaaca aattttggga cctaatgctt caactaactc cagtaattcc ttggtggtac
     1261 gaacatccaa tgaagcacac aagtttgttt gcttttcgtg catgatatta aatagcttgg
     1321 cagcaacagg actaggatga gtagcagcac gttccttata tgtagctttc gacatgattt
     1381 atcttcgttt cctgcaggtt tttgttctgt gcagttgggt taagaatact gggcaatttc
     1441 atgtttcttc aacactacat atgcgtatat ataccaatct aagtctgtgc tccttccttc
     1501 gttcttcctt ctgttcggag attaccgaat caaaaaaatt tcaaagaaac cgaaatcaaa
     1561 aaaaagaata aaaaaaaaat gatgaattga attgaaaagc tagcttatcg atgataagct
     1621 gtcaaagatg agaattaatt ccacggacta tagactatac tagatactcc gtctactgta
     1681 cgatacactt ccgctcaggt ccttgtcctt taacgaggcc ttaccactct tttgttactc
     1741 tattgatcca gctcagcaaa ggcagtgtga tctaagattc tatcttcgcg atgtagtaaa
     1801 actagctaga ccgagaaaga gactagaaat gcaaaaggca cttctacaat ggctgccatc
     1861 attattatcc gatgtgacgc tgcagcttct caatgatatt cgaatacgct ttgaggagat
     1921 acagcctaat atccgacaaa ctgttttaca gatttacgat cgtacttgtt acccatcatt
     1981 gaattttgaa catccgaacc tgggagtttt ccctgaaaca gatagtatat ttgaacctgt
     2041 ataataatat atagtctagc gctttacgga agacaatgta tgtatttcgg ttcctggaga
     2101 aactattgca tctattgcat aggtaatctt gcacgtcgca tccccggttc attttctgcg
     2161 tttccatctt gcacttcaat agcatatctt tgttaacgaa gcatctgtgc ttcattttgt
     2221 agaacaaaaa tgcaacgcga gagcgctaat ttttcaaaca aagaatctga gctgcatttt
     2281 tacagaacag aaatgcaacg cgaaagcgct attttaccaa cgaagaatct gtgcttcatt
     2341 tttgtaaaac aaaaatgcaa cgcgacgaga gcgctaattt ttcaaacaaa gaatctgagc
     2401 tgcattttta cagaacagaa atgcaacgcg agagcgctat tttaccaaca aagaatctat
     2461 acttcttttt tgttctacaa aaatgcatcc cgagagcgct atttttctaa caaagcatct
     2521 tagattactt tttttctcct ttgtgcgctc tataatgcag tctcttgata actttttgca
     2581 ctgtaggtcc gttaaggtta gaagaaggct actttggtgt ctattttctc ttccataaaa
     2641 aaagcctgac tccacttccc gcgtttactg attactagcg aagctgcggg tgcatttttt
     2701 caagataaag gcatccccga ttatattcta taccgatgtg gattgcgcat actttgtgaa
     2761 cagaaagtga tagcgttgat gattcttcat tggtcagaaa attatgaacg gtttcttcta
     2821 ttttgtctct atatactacg tataggaaat gtttacattt tcgtattgtt ttcgattcac
     2881 tctatgaata gttcttacta caattttttt gtctaaagag taatactaga gataaacata
     2941 aaaaatgtag aggtcgagtt tagatgcaag ttcaaggagc gaaaggtgga tgggtaggtt
     3001 atatagggat atagcacaga gatatatagc aaagagatac ttttgagcaa tgtttgtgga
     3061 agcggtattc gcaatgggaa gctccacccc ggttgataat cagaaaagcc ccaaaaacag
     3121 gaagattgta taagcaaata tttaaattgt aattctttcc tgcgttatcc cctgattctg
     3181 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     3241 agcgcagcga gtcagtgagc gaggaagcgg aagagggtct gacgctcagt ggaacgaaaa
     3301 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3361 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3421 tcagaagaac tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat
     3481 accgtaaagc acgaggaagc ggtcagccca ttcgccgcca agctcttcag caatatcacg
     3541 ggtagccaac gctatgtcct gatagcggtc cgccacaccc agccggccac agtcgatgaa
     3601 tccagaaaag cggccatttt ccaccatgat attcggcaag caggcatcgt catgggtcac
     3661 gacgagatcc tcgccgtcgg gcatgctcgc cttgagcctg gcgaacagtt cggctggcgc
     3721 gagcccctga tgctcttcgt ccagatcatc ctgatcgaca agaccggctt ccatccgagt
     3781 acgtgctcgc tcgatgcgat gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag
     3841 cgtatgcagc cgccgcattg catcagccat gatggatact ttctcggcag gagcaaggtg
     3901 agatgacagg agatcctgcc ccggcacttc gcccaatagc agccagtccc ttcccgcttc
     3961 agtgacaacg tcgagcacag ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg
     4021 cgctgcctcg tcttgcagtt cattcagggc accggacagg tcggtcttga caaaaagaac
     4081 cgggcgcccc tgcgctgaca gccggaacac ggcggcatca gagcagccga ttgtctgttg
     4141 tgcccagtca tagccgaata gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc
     4201 atcttgttca atcatactct tcctttttca atattattga agcatttatc agggttattg
     4261 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     4321 cacatttccc cgaaaagtgc cacctgacgt cggacggatc gcttgcctgt aacttacacg
     4381 cgcctcgtgg cgcgcccctg caggatttaa atggccggcc gtttaaacat ttatcggaga
     4441 aaacctcgtc gttgaaccag atttaagctt ggcttactaa aagccagata acagtatgcg
     4501 tatttgcgcg ctgatttttg cggtataaga atatatactg atatgtatac ccgaagtatg
     4561 tcaaaaagag gtatgctatg aagcagcgta ttacagtgac agttgacagc gacagctatc
     4621 agttgctcaa ggcatatatg atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa
     4681 tgaagcccgt cgtctgcgtg ccgaacgctg gaaagcggaa aatcaggaag ggatggctga
     4741 ggtcgcccgg tttattgaaa tgaacggctc ttttgctgac gagaacaggg gctggtgaaa
     4801 tgcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     4861 agagtgatat tattgacacg cccgggcgac ggatggtgat ccccctggcc agtgcacgtc
     4921 tgctgtcaga taaagtcccc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     4981 ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg
     5041 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     5101 tataaatgtc aggctccctt atacacagcc agtaagcttt agaggcatcg cacagtttta
     5161 gcgaggaaaa ctcttcaata gttttgccag cggaattcca cttgcaatta cataaaaaat
     5221 tccggcggtt tttcgcgtgt gactcaatgt cgaaatacct gcctaatgaa catgaacatc
     5281 gcccaaatgt atttgaagac ccgctgggag aagttcaaga tatataagta acaagcagcc
     5341 aatagtataa aaaaaaatct gagtttatta cctttcctgg aatttcagtg aaaaactgct
     5401 aattatagag agatatcaca gagttactca ctaatgacta acgaaaaggt ctggatagag
     5461 aagttggata atccaactct ttcatgaatt taaatggcgc gccgtttaaa ccctgcaggg
     5521 gccggccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5581 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5641 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaat
//
