LOCUS       40924_31970        5900 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pMQ117, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5900)
  AUTHORS   Shanks RM, Kadouri DE, MacEachran DP, O'Toole GA.
  TITLE     New yeast recombineering tools for bacteria
  JOURNAL   Plasmid 62 (2), 88-97 (2009)
  PUBMED    19477196
REFERENCE   2  (bases 1 to 5900)
  AUTHORS   Shanks RM, MacEachran DP, Powers SZ, Kadouri DE, O'Toole GA.
  TITLE     In vivo recombination vectors for modification and expression of 
            genes in Gram-negative and Gram-positive bacteria
  JOURNAL   Unpublished
REFERENCE   3  (bases 1 to 5900)
  AUTHORS   Shanks RM, MacEachran DP, Powers SZ, Kadouri DE, O'Toole GA.
  TITLE     Direct Submission
  JOURNAL   Submitted (03-MAR-2008) Ophthalmology, University of Pittsburgh, 203
            Lothrop St., Pittsburgh, PA 15213, USA
REFERENCE   4  (bases 1 to 5900)
  TITLE     Direct Submission
REFERENCE   5  (bases 1 to 5900)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plasmid"; 
            date: "2009"; volume: "62"; issue: "2"; pages: "88-97"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 3; type: "Journal Article"; journalName: "Submitted 
            (03-MAR-2008) Ophthalmology, University of Pittsburgh, 203 Lothrop 
            St., Pittsburgh, PA 15213, USA"
COMMENT     SGRef: number: 4; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5900
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            complement(399..508)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     terminator      complement(929..956)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(1048..1134)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     primer_bind     1484..1500
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    complement(1504..1560)
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(1570..1586)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(1594..1610)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(1618..1648)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(1663..1684)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      1871..2093
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             2141..3088
                     /codon_start=1
                     /label=Rep101(Ts)
                     /note="temperature-sensitive version of the RepA protein
                     needed for replication with the pSC101 origin (Armstrong et
                     al., 1984)"
                     /translation="MSELVVFKANELATSRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYVQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     CDS             complement(3379..3909)
                     /codon_start=1
                     /label=GmR
                     /note="gentamycin acetyltransferase"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(4098..4126)
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        complement(4144..4248)
                     /label=AmpR promoter
     CDS             complement(4348..5148)
                     /codon_start=1
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(5149..5369)
                     /label=URA3 promoter
     misc_feature    5397..5900
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
ORIGIN
        1 tctaagaaac cattattatc atgacattaa cctataaaaa aggcctctcg agctagagtc
       61 gatcttcgcc agcagggcga ggatcgtggc atcaccgaac cgcgccgtgc gcgggtcgtc
      121 ggtgagccag agtttcagca ggccgcccag gcggcccagg tcgccattga tgcgggccag
      181 ctcgcggacg tgctcatagt ccacgacgcc cgtgattttg tagccctggc cgacggccag
      241 caggtaggcc gacaggctca tgccggccgc cgccgccttt tcctcaatcg ctcttcgttc
      301 gtctggaagg cagtacacct tgataggtgg gctgcccttc ctggttggct tggtttcatc
      361 agccatccgc ttgccctcat ctgttacgcc ggcggtagcc ggccagcctc gcagagcagg
      421 attcccgttg agcaccgcca ggtgcgaata agggacagtg aagaaggaac acccgctcgc
      481 gggtgggcct acttcaccta tcctgcccgg ctgacgccgt tggatacacc aaggaaagtc
      541 tacacgaacc ctttggcaaa atcctgtata tcgtgcgaaa aaggatggat ataccgaaaa
      601 aatcgctata atgaccccga agcagggtta tgcagcggaa aagcgctgct tccctgctgt
      661 tttgtggaat atctaccgac tggaaacagg caaatgcagg aaattactga actgagggga
      721 caggcgagag acgatgccaa agagctacac cgacgagctg gccgagtggg ttgaatcccg
      781 cgcggccaag aagcgccggc gtgatgaggc tgcggttgcg ttcctggcgg tgagggcgga
      841 tgtcgatatg cgtaaggaga aaataccgca tcaggcgcat atttgaatgt atttagaaaa
      901 ataaacaaaa agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt
      961 aatttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg gccgttgctt
     1021 cgcaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt tcaccgacaa
     1081 acaacagata aaacgaaagg cccagtcttt cgactgagcc tttcgtttta tttgatgcct
     1141 ggcagttccc tactctcgca tggggagacc ccacactacc atcggcgcta cggcgtttca
     1201 cttctgagtt cggcatgggg tcaggtggga ccaccgcgct actgccgcca ggcaaattct
     1261 gttttatcag accgcttctg cgttctgatt taatctgtat caggctgaaa atcttctctc
     1321 atccgccaaa acagccaagc tcgccattcg ccattcaggc tgcgcaactg ttgggaaggg
     1381 cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg
     1441 cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt
     1501 gccaagcttg catgcctgca ggtcgactct agaggatccc cgggtaccga gctcgaattc
     1561 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     1621 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     1681 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     1741 ttaatgcagc gatttgcccg agcttgcgag ggtgctactt aagcctttag ggttttaagg
     1801 tctgttttgt agaggagcaa acagcgtttg cgacatcctt ttgtaatact gcggaactga
     1861 ctaaagtagt gagttataca cagggctggg atctattctt tttatctttt tttattcttt
     1921 ctttattcta taaattataa ccacttgaat ataaacaaaa aaaacacaca aaggtctagc
     1981 ggaatttaca gagggtctag cagaatttac aagttttcca gcaaaggtct agcagaattt
     2041 acagataccc acaactcaaa ggaaaaggac tagtaattat cattgactag cccatctcaa
     2101 ttggtatagt gattaaaatc acctagacca attgagatgt atgtctgaat tagttgtttt
     2161 caaagcaaat gaactagcga ctagtcgcta tgacttaacg gagcatgaaa ccaagctaat
     2221 tttatgctgt gtggcactac tcaaccccac gattgaaaac cctacaagga aagaacggac
     2281 ggtatcgttc acttataacc aatacgttca gatgatgaac atcagtaggg aaaatgctta
     2341 tggtgtatta gctaaagcaa ccagagagct gatgacgaga actgtggaaa tcaggaatcc
     2401 tttggttaaa ggctttgaga ttttccagtg gacaaactat gccaagttct caagcgaaaa
     2461 attagaatta gtttttagtg aagagatatt gccttatctt ttccagttaa aaaaattcat
     2521 aaaatataat ctggaacatg ttaagtcttt tgaaaacaaa tactctatga ggatttatga
     2581 gtggttatta aaagaactaa cacaaaagaa aactcacaag gcaaatatag agattagcct
     2641 tgatgaattt aagttcatgt taatgcttga aaataactac catgagttta aaaggcttaa
     2701 ccaatgggtt ttgaaaccaa taagtaaaga tttaaacact tacagcaata tgaaattggt
     2761 ggttgataag cgaggccgcc cgactgatac gttgattttc caagttgaac tagatagaca
     2821 aatggatctc gtaaccgaac ttgagaacaa ccagataaaa atgaatggtg acaaaatacc
     2881 aacaaccatt acatcagatt cctacctaca taacggacta agaaaaacac tacacgatgc
     2941 tttaactgca aaaattcagc tcaccagttt tgaggcaaaa tttttgagtg acatgcaaag
     3001 taagtatgat ctcaatggtt cgttctcatg gctcacgcaa aaacaacgaa ccacactaga
     3061 gaacatactg gctaaatacg gaaggatctg aggttcttat ggctcttgta tctatcagtg
     3121 aagcatcaag actaacaaac aaaagtagaa caactgttca ccgttacata tcaaagggaa
     3181 aactgtccat atgcacagat gaaaacggtg taaaaaagat agatacatca gagcttttac
     3241 gagtttttgg tgcaatcaat ctaaagtata tatgagtaaa cttggtctga caatcgatgc
     3301 gaattggccg cggcgttgtg acaatttacc gaacaactcc gcggccggga agccgatctc
     3361 ggcttgaacg aattgttagg tggcggtact tgggtcgata tcaaagtgca tcacttcttc
     3421 ccgtatgccc aactttgtat agagagccac tgcgggatcg tcaccgtaat ctgcttgcac
     3481 gtagatcaca taagcaccaa gcgcgttggc ctcatgcttg aggagattga tgagcgcggt
     3541 ggcaatgccc tgcctccggt gctcgccgga gactgcgaga tcatagatat agatctcact
     3601 acgcggctgc tcaaacctgg gcagaacgta agccgcgaga gcgccaacaa ccgcttcttg
     3661 gtcgaaggca gcaagcgcga tgaatgtctt actacggagc aagttcccga ggtaatcgga
     3721 gtccggctga tgttgggagt aggtggctac gtctccgaac tcacgaccga aaagatcaag
     3781 agcagcccgc atggatttga cttggtcagg gccgagccta catgtgcgaa tgatgcccat
     3841 acttgagcca cctaactttg ttttagggcg actgccctgc tgcgtaacat cgttgctgct
     3901 gcgtaacatc gttgctgctc cataacatca aacatcgacc cacggcgtaa cgcgcttgct
     3961 gcttggatgc ccgaggcata gactgtacaa aaaaacagtc ataacaagcc atgaaaaccg
     4021 ccactgcgcc gttaccaccg ctgcgttcgg tcaaggttct ggaccagttg cgtgagcgca
     4081 tacgctactt gcattacagt ttacgaaccg aacaggctta tgtcaattcg gttgaatact
     4141 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     4201 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     4261 aaaagtgggt aataactgat ataattaaat tgaagctcta atttgtgagt ttagtataca
     4321 tgcatttact tataatacag ttttttagtt ttgctggccg catcttctca aatatgcttc
     4381 ccagcctgct tttctgtaac gttcaccctc taccttagca tcccttccct ttgcaaatag
     4441 tcctcttcca acaataataa tgtcagatcc tgtagagacc acatcatcca cggttctata
     4501 ctgttgaccc aatgcgtctc ccttgtcatc taaacccaca ccgggtgtca taatcaacca
     4561 atcgtaacct tcatctcttc cacccatgtc tctttgagca ataaagccga taacaaaatc
     4621 tttgtcgctc ttcgcaatgt caacagtacc cttagtatat tctccagtag atagggagcc
     4681 cttgcatgac aattctgcta acatcaaaag gcctctaggt tcctttgtta cttcttctgc
     4741 cgcctgcttc aaaccgctaa caatacctgg gcccaccaca ccgtgtgcat tcgtaatgtc
     4801 tgcccattct gctattctgt atacacccgc agagtactgc aatttgactg tattaccaat
     4861 gtcagcaaat tttctgtctt cgaagagtaa aaaattgtac ttggcggata atgcctttag
     4921 cggcttaact gtgccctcca tggaaaaatc agtcaagata tccacatgtg tttttagtaa
     4981 acaaattttg ggacctaatg cttcaactaa ctccagtaat tccttggtgg tacgaacatc
     5041 caatgaagca cacaagtttg tttgcttttc gtgcatgata ttaaatagct tggcagcaac
     5101 aggactagga tgagtagcag cacgttcctt atatgtagct ttcgacatga tttatcttcg
     5161 tttcctgcag gtttttgttc tgtgcagttg ggttaagaat actgggcaat ttcatgtttc
     5221 ttcaacacta catatgcgta tatataccaa tctaagtctg tgctccttcc ttcgttcttc
     5281 cttctgttcg gagattaccg aatcaaaaaa atttcaaaga aaccgaaatc aaaaaaaaga
     5341 ataaaaaaaa aatgatgaat tgaattgaaa agctagctta tcgatgggtc cttttcatca
     5401 cgtgctataa aaataattat aatttaaatt ttttaatata aatatataaa ttaaaaatag
     5461 aaagtaaaaa aagaaattaa agaaaaaata gtttttgttt tccgaagatg taaaagactc
     5521 tagggggatc gccaacaaat actacctttt atcttgctct tcctgctctc aggtattaat
     5581 gccgaattgt ttcatcttgt ctgtgtagaa gaccacacac gaaaatcctg tgattttaca
     5641 ttttacttat cgttaatcga atgtatatct atttaatctg cttttcttgt ctaataaata
     5701 tatatgtaaa gtacgctttt tgttgaaatt ttttaaacct ttgtttattt ttttttcttc
     5761 attccgtaac tcttctacct tctttattta ctttctaaaa tccaaataca aaacataaaa
     5821 ataaataaac acagagtaaa ttcccaaatt attccatcat taaaagatac gaggcgcgtg
     5881 taagttacag gcaagcgatc
//
