LOCUS       40924_32035        6281 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Suicide vector pMQ200, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6281)
  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 6281)
  AUTHORS   Shanks RMQ., Kadouri DE, O'Toole GA.
  TITLE     Recombineering vectors for a wide range of Gram-negative and 
            -positive bacteria
  JOURNAL   Unpublished
REFERENCE   3  (bases 1 to 6281)
  AUTHORS   Shanks RMQ., Kadouri DE, O'Toole GA.
  TITLE     Direct Submission
  JOURNAL   Submitted (13-OCT-2008) Ophthalmololgy, University of Pittsburgh, 
            203 Lothrop St, Pittsburgh, PA 15213, USA
REFERENCE   4  (bases 1 to 6281)
  TITLE     Direct Submission
REFERENCE   5  (bases 1 to 6281)
  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            "Unpublished"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 
            (13-OCT-2008) Ophthalmololgy, University of Pittsburgh, 203 Lothrop 
            St, Pittsburgh, PA 15213, USA"
COMMENT     SGRef: number: 4; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6281
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..801
                     /codon_start=1
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     oriT            complement(893..1001)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     promoter        complement(1192..1210)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      1418..1806
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     terminator      complement(1889..1975)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     primer_bind     2311..2327
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2337..2355
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    complement(2364..2471)
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     promoter        complement(2484..2502)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(2545..2829)
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     CDS             2856..3731
                     /codon_start=1
                     /label=araC
                     /note="L-arabinose regulatory protein"
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     CDS             4198..4989
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        complement(5148..5256)
                     /label=AmpR promoter
     misc_feature    5293..5796
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
     promoter        6061..6281
                     /label=URA3 promoter
ORIGIN
        1 atgtcgaaag ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag
       61 ctatttaata tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc
      121 accaaggaat tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca
      181 catgtggata tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta
      241 tccgccaagt acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca
      301 gtcaaattgc agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat
      361 gcacacggtg tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta
      421 acaaaggaac ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct
      481 actggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc
      541 ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg
      601 acacccggtg tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg
      661 gatgatgtgg tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag
      721 ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg
      781 agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac
      841 tcacaaatta gagcttcaat ttaattatat cagttattca cgtggagctt atcggccagc
      901 ctcgcagagc aggattcccg ttgagcaccg ccaggtgcga ataagggaca gtgaagaagg
      961 aacacccgct cgcgggtggg cctacttcac ctatcctgcc cggctgacgc cgttggatac
     1021 accaaggaaa gtctacacga accctttggc aaaatcctgt atatcgtgcg aaaaaggatg
     1081 gatataccga aaaaatcgct ataatgaccc cgaagcaggg ttatgcagcg gaaagtatac
     1141 cttaaccgcc ggctgacccg gcggggacga ggcaagctaa acagatcgac ccctatagtg
     1201 agtcgtatta caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     1261 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     1321 cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg
     1381 cgcggaacca gatccccacg atgcgtccgg cgtagaggat ctgaagatca gcagttcaac
     1441 ctgttgatag tacgtactaa gctctcatgt ttcacgtact aagctctcat gtttaacgta
     1501 ctaagctctc atgtttaacg aactaaaccc tcatggctaa cgtactaagc tctcatggct
     1561 aacgtactaa gctctcatgt ttcacgtact aagctctcat gtttgaacaa taaaattaat
     1621 ataaatcagc aacttaaata gcctctaagg ttttaagttt tataagaaaa aaaagaatat
     1681 ataaggcttt taaagctttt aaggtttaac ggttgtggac aacaagccag ggatgtaacg
     1741 cactgagaag cccttagagc ctctcaaagc aattttcagt gacacaggaa cacttaacgg
     1801 ctgacagatg cctggcagtt tatggcgggc gtcctgcccg ccaccctccg ggccgttgct
     1861 tcgcaacgtt caaatccgct cccggcggat ttgtcctact caggagagcg ttcaccgaca
     1921 aacaacagat aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt atttgatgcc
     1981 tggcagttcc ctactctcgc atggggagac cccacactac catcggcgct acggcgtttc
     2041 acttctgagt tcggcatggg gtcaggtggg accaccgcgc tactgccgcc aggcaaattc
     2101 tgttttatca gaccgcttct gcgttctgat ttaatctgta tcaggctgaa aatcttctct
     2161 catccgccaa agtttcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     2221 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     2281 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat
     2341 acgactcact atagggcgaa ttgggtaccg ggccccccct cgaggtcgac ggtatcgata
     2401 agcttgatat cgaattcctg cagcccgggg gatccactag ttctagagcg gccgccaccg
     2461 cggtggagct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     2521 tggtcatttt ttcctcctac gggtatggag aaacagtaga gagttgcgat aaaaagcgtc
     2581 aggtaggatc cgctaatctt atggataaaa atgctatggc atagcaaagt gtgacgccgt
     2641 gcaaataatc aatgtggact tttctgccgt gattatagac acttttgtta cgcgtttttg
     2701 tcatggcttt ggtcccgctt tgttacagaa tgcttttaat aagcggggtt accggtttgg
     2761 ttagcgagaa gagccagtaa aagacgcagt gacggcaatg tctgatgcaa tatggacaat
     2821 tggtttcttc tctgaatggc gggagtatga aaagtatggc tgaagcgcaa aatgatcccc
     2881 tgctgccggg atactcgttt aatgcccatc tggtggcggg tttaacgccg attgaggcca
     2941 acggttatct cgattttttt atcgaccgac cgctgggaat gaaaggttat attctcaatc
     3001 tcaccattcg cggtcagggg gtggtgaaaa atcagggacg agaatttgtt tgccgaccgg
     3061 gtgatatttt gctgttcccg ccaggagaga ttcatcacta cggtcgtcat ccggaggctc
     3121 gcgaatggta tcaccagtgg gtttactttc gtccgcgcgc ctactggcat gaatggctta
     3181 actggccgtc aatatttgcc aatacggggt tctttcgccc ggatgaagcg caccagccgc
     3241 atttcagcga cctgtttggg caaatcatta acgccgggca aggggaaggg cgctattcgg
     3301 agctgctggc gataaatctg cttgagcaat tgttactgcg gcgcatggaa gcgattaacg
     3361 agtcgctcca tccaccgatg gataatcggg tacgcgaggc ttgtcagtac atcagcgatc
     3421 acctggcaga cagcaatttt gatatcgcca gcgtcgcaca gcatgtttgc ttgtcgccgt
     3481 cgcgtctgtc acatcttttc cgccagcagt tagggattag cgtcttaagc tggcgcgagg
     3541 accaacgtat cagccaggcg aagctgcttt tgagcaccac ccggatgcct atcgccaccg
     3601 tcggtcgcaa tgttggtttt gacgatcaac tctatttctc gcgggtattt aaaaaatgca
     3661 ccggggccag cccgagcgag ttccgtgccg gttgtgaaga aaaagtgaat gatgtagccg
     3721 tcaagttgtc ataattggta acgaatcaga caattgacgg cttgacggag tagcataggg
     3781 tttgcagaat ccctgcttcg tccatttgac aggcacatta tgcatcgatt aagctgtcaa
     3841 acatgagcac gctgccgcaa gcactcaggg cgcaagggct gctaaaggaa gcggaacacg
     3901 tagaaagcca gtccgcagaa acggtgctga ccccggatga atgtcagcta ctgggctatc
     3961 tggacaaggg aaaacgcaag cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg
     4021 cgatagctag actgggcggt tttatggaca gcaagcgaac cggaattgcc agctggggcg
     4081 ccctctggta aggttgggaa gccctgcaaa gtaaactgga tggctttctt gccgccaagg
     4141 atctgatggc gcaggggatc aagatctgat caagagacag gatgaggatc gtttcgcatg
     4201 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     4261 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4321 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactccaa
     4381 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4441 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4501 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4561 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4621 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4681 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcggat gcccgacggc
     4741 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4801 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4861 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4921 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4981 gagttcttct gagcgggact ctggggtacg cgtctaggcg gccgcgaagt tcctattccg
     5041 aagttcctct agaagcttct gcagacgggt ttgactggtc aaatttcgaa cgacatcatt
     5101 ctcattgact atactgattt cgtcagactt acgggttgaa tactcatact cttccttttt
     5161 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     5221 atttagaaaa aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     5281 tgggtccttt tcatcacgtg ctataaaaat aattataatt taaatttttt aatataaata
     5341 tataaattaa aaatagaaag taaaaaaaga aattaaagaa aaaatagttt ttgttttccg
     5401 aagatgtaaa agactctagg gggatcgcca acaaatacta ccttttatct tgctcttcct
     5461 gctctcaggt attaatgccg aattgtttca tcttgtctgt gtagaagacc acacacgaaa
     5521 atcctgtgat tttacatttt acttatcgtt aatcgaatgt atatctattt aatctgcttt
     5581 tcttgtctaa taaatatata tgtaaagtac gctttttgtt gaaatttttt aaacctttgt
     5641 ttattttttt ttcttcattc cgtaactctt ctaccttctt tatttacttt ctaaaatcca
     5701 aatacaaaac ataaaaataa ataaacacag agtaaattcc caaattattc catcattaaa
     5761 agatacgagg cgcgtgtaag ttacaggcaa gcgatccgtc ctaagaaacc attattatca
     5821 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg
     5881 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     5941 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     6001 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat accacagctt
     6061 ttcaattcaa ttcatcattt tttttttatt cttttttttg atttcggttt ctttgaaatt
     6121 tttttgattc ggtaatctcc gaacagaagg aagaacgaag gaaggagcac agacttagat
     6181 tggtatatat acgcatatgt agtgttgaag aaacatgaaa ttgcccagta ttcttaaccc
     6241 aactgcacag aacaaaaacc tgcaggaaac gaagataaat c
//
