LOCUS       40924_11366        8081 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pCM433, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8081)
  AUTHORS   Marx CJ.
  TITLE     Development of a broad-host-range sacB-based vector for unmarked 
            allelic exchange
  JOURNAL   BMC Res Notes 1, 1 (2008)
  PUBMED    18710539
REFERENCE   2  (bases 1 to 8081)
  AUTHORS   Marx CJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (23-AUG-2007) Organismic and Evolutionary Biology, Harvard
            University, 3083 Biological Laboratories, 16 Divinity Avenue, 
            Cambridge, MA 02138, USA
REFERENCE   3  (bases 1 to 8081)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8081)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC Res 
            Notes 1, 1 (2008)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (23-AUG-2007) Organismic and Evolutionary Biology, Harvard 
            University, 3083 Biological Laboratories, 16 Divinity Avenue, 
            Cambridge, MA 02138, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8081
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(4..861)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(862..966)
                     /label=AmpR promoter
     CDS             complement(1696..2064)
                     /codon_start=1
                     /label=traJ
                     /note="oriT-recognizing protein"
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            complement(2097..2206)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     promoter        2843..2871
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein
                     gene"
     CDS             2919..4106
                     /codon_start=1
                     /label=TcR
                     /note="tetracycline efflux protein"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSITGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     CDS             complement(4290..4946)
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             complement(5047..6465)
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        complement(6466..6911)
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     rep_origin      complement(7323..7911)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
       61 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
      121 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
      181 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
      241 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      301 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      361 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      421 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
      481 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
      541 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
      601 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
      661 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
      721 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
      781 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
      841 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
      901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      961 tccgcgcaca tttccccgaa aagtgccacc tgacgtctag atctgaattc agctgtacaa
     1021 ttggtaccat ggatgcatat gctgcagctc gagcggccgc gggccctacg tacgcgtgtt
     1081 aaccggtgag ctcactagag gatccagccg accaggcttt ccacgcccgc gtgccgctcc
     1141 atgtcgttcg cgcggttctc ggaaacgcgc tgccgcgttt cgtgattgtc acgctcaagc
     1201 ccgtagtccc gttcgagcgt cgcgcagagg tcagcgaggg cgcggtaggc ccgatacggc
     1261 tcatggatgg tgtttcgggt cgggtgaatc ttgttgatgg cgatatggat gtgcaggttg
     1321 tcggtgtcgt gatgcacggc actgacgcgc tgatgctcgg cgaagccaag cccagcgcag
     1381 atgcggtcct caatcgcgcg caacgtctcc gcgtcgggct tctctcccgc gcggaagcta
     1441 accagcaggt gataggtctt gtcggcctcg gaacgggtgt tgccgtgctg ggtcgccatc
     1501 acctcggcca tgacagcggg cagggtgttt gcctcgcagt tcgtgacgcg cacgtgaccc
     1561 aggcgctcgg tcttgccttg ctcgtcggtg atgtacttca ccagctccgc gaagtcgctc
     1621 ttcttgatgg agcgcatggg gacgtgcttg gcaatcacgc gcaccccccg gccgttttag
     1681 cggctaaaaa agtcatggct ctgccctcgg gcggaccacg cccatcatga ccttgccaag
     1741 ctcgtcctgc ttctcttcga tcttcgccag cagggcgagg atcgtggcat caccgaaccg
     1801 cgccgtgcgc gggtcgtcgg tgagccagag tttcagcagg ccgcccaggc ggcccaggtc
     1861 gccattgatg cgggccagct cgcggacgtg ctcatagtcc acgacgcccg tgattttgta
     1921 gccctggccg acggccagca ggtaggccga caggctcatg ccggccgccg ccgccttttc
     1981 ctcaatcgct cttcgttcgt ctggaaggca gtacaccttg ataggtgggc tgcccttcct
     2041 ggttggcttg gtttcatcag ccatccgctt gccctcatct gttacgccgg cggtagccgg
     2101 ccagcctcgc agagcaggat tcccgttgag caccgccagg tgcgaataag ggacagtgaa
     2161 gaaggaacac ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct gacgccgttg
     2221 gatacaccaa ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc gtgcgaaaaa
     2281 ggatggatat accgaaaaaa tcgctataat gaccccgaag cagggttatg cagcggaaaa
     2341 gcgctgcttc cctgctgttt tgtggaatat ctaccgactg gaaacaggca aatgcaggaa
     2401 attactgaac tgaggggaca ggcgagagac gatgccaaag agctacaccg acgagctggc
     2461 cgagtgggtt gaatcccgcg cggccaagaa gcgccggcgt gatgaggctg cggttgcgtt
     2521 cctggcggtg agggcggatg tcgaggcggc gttagcgtcc ggctatgcgc tcgtcaccat
     2581 ttgggagcac atgcgggaaa cggggaaggt caagttctcc tacgagacgt tccgctcgca
     2641 cgccaggcgg cacatcaagg ccaagcccgc cgatgtgccc gcaccgcagg ccaaggctgc
     2701 ggaacccgcg ccggcaccca agacgccgga gccacggcgg ccgaagcagg ggggcaaggc
     2761 tgaaaagccg gcccccgctg cggccccgac cggcttcacc ttcaacccaa caccggacaa
     2821 aaaggatccc caattctcat gtttgacagc ttatcatcga taagctttaa tgcggtagtt
     2881 tatcacagtt aaattgctaa cgcagtcagg caccgtgtat gaaatctaac aatgcgctca
     2941 tcgtcatcct cggcaccgtc accctggatg ctgtaggcat aggcttggtt atgccggtac
     3001 tgccgggcct cttgcgggat atcgtccatt ccgacagcat cgccagtcac tatggcgtgc
     3061 tgctagcgct atatgcgttg atgcaatttc tatgcgcacc cgttctcgga gcactgtccg
     3121 accgctttgg ccgccgccca gtcctgctcg cttcgctact tggagccact atcgactacg
     3181 cgatcatggc gaccacaccc gtcctgtgga tcctctacgc cggacgcatc gtggccggca
     3241 tcaccggcgc cacaggtgcg gttgctggcg cctatatcgc cgacatcacc gatggggaag
     3301 atcgggctcg ccacttcggg ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc
     3361 ccgtggccgg gggactgttg ggcgccatct ccttgcatgc accattcctt gcggcggcgg
     3421 tgctcaacgg cctcaaccta ctactgggct gcttcctaat gcaggagtcg cataagggag
     3481 agcgtcgacc gatgcccttg agagccttca acccagtcag ctccttccgg tgggcgcggg
     3541 gcatgactat cgtcgccgca cttatgactg tcttctttat catgcaactc gtaggacagg
     3601 tgccggcagc gctctgggtc attttcggcg aggaccgctt tcgctggagc gcgacgatga
     3661 tcggcctgtc gcttgcggta ttcggaatct tgcacgccct cgctcaagcc ttcgtcactg
     3721 gtcccgccac caaacgtttc ggcgagaagc aggccattat cgccggcatg gcggccgacg
     3781 cgctgggcta cgtcttgctg gcgttcgcga cgcgaggctg gatggccttc cccattatga
     3841 ttcttctcgc ttccggcggc atcgggatgc ccgcgttgca ggccatgctg tccaggcagg
     3901 tagatgacga ccatcaggga cagcttcaag gatcgctcgc ggctcttacc agcctaactt
     3961 cgatcactgg accgctgatc gtcacggcga tttatgccgc ctcggcgagc acatggaacg
     4021 ggttggcatg gattgtaggc gccgccctat accttgtctg cctccccgcg ttgcgtcgcg
     4081 gtgcatggag ccgggccacc tcgacctgaa tggaagccgg cggcacctcg ctaacggatt
     4141 caccactcca agaattggag ccaatcaatt cttgcggaga actgtgaatg cgcaaaccaa
     4201 cccttggcag aacatatcca tcgcgtccgc catctccagc agccgcacgc ggcgcatctc
     4261 gggcagcgtt gggtcctggc cgggaattac gccccgccct gccactcatc gcagtactgt
     4321 tgtaattcat taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga
     4381 atcgccagcg gcatcagcac cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg
     4441 ggggcgaaga agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag
     4501 ggattggctg agacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt
     4561 tcaccgtaac acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg
     4621 tattcactcc agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg
     4681 tgaacactat cccatatcac cagctcaccg tctttcattg ccatacggaa ttccggatga
     4741 gcattcatca ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc
     4801 tttacggtct ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattga
     4861 gcaactgact gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg
     4921 gtatatccag tgattttttt ctccatttta gcttccttag ctcctgccct atgggattca
     4981 cctttatgtt gataagaaat aaaagaaaat gccaatagga tatcggcatt ttcttttgcg
     5041 tttttatttg ttaactgtta attgtccttg ttcaaggatg ctgtctttga caacagatgt
     5101 tttcttgcct ttgatgttca gcaggaagct tggcgcaaac gttgattgtt tgtctgcgta
     5161 gaatcctctg tttgtcatat agcttgtaat cacgacattg tttcctttcg cttgaggtac
     5221 agcgaagtgt gagtaagtaa aggttacatc gttaggatca agatccattt ttaacacaag
     5281 gccagttttg ttcagcggct tgtatgggcc agttaaagaa ttagaaacat aaccaagcat
     5341 gtaaatatcg ttagacgtaa tgccgtcaat cgtcattttt gatccgcggg agtcagtgaa
     5401 caggtaccat ttgccgttca ttttaaagac gttcgcgcgt tcaatttcat ctgttactgt
     5461 gttagatgca atcagcggtt tcatcacttt tttcagtgtg taatcatcgt ttagctcaat
     5521 cataccgaga gcgccgtttg ctaactcagc cgtgcgtttt ttatcgcttt gcagaagttt
     5581 ttgactttct tgacggaaga atgatgtgct tttgccatag tatgctttgt taaataaaga
     5641 ttcttcgcct tggtagccat cttcagttcc agtgtttgct tcaaatacta agtatttgtg
     5701 gcctttatct tctacgtagt gaggatctct cagcgtatgg ttgtcgcctg agctgtagtt
     5761 gccttcatcg atgaactgct gtacattttg atacgttttt ccgtcaccgt caaagattga
     5821 tttataatcc tctacaccgt tgatgttcaa agagctgtct gatgctgata cgttaacttg
     5881 tgcagttgtc agtgtttgtt tgccgtaatg tttaccggag aaatcagtgt agaataaacg
     5941 gatttttccg tcagatgtaa atgtggctga acctgaccat tcttgtgttt ggtcttttag
     6001 gatagaatca tttgcatcga atttgtcgct gtctttaaag acgcggccag cgtttttcca
     6061 gctgtcaata gaagtttcgc cgactttttg atagaacatg taaatcgatg tgtcatccgc
     6121 atttttagga tctccggcta atgcaaagac gatgtggtag ccgtgatagt ttgcgacagt
     6181 gccgtcagcg ttttgtaatg gccagctgtc ccaaacgtcc aggccttttg cagaagagat
     6241 atttttaatt gtggacgaat cgaattcagg aacttgatat ttttcatttt tttgctgttc
     6301 agggatttgc agcatatcat ggcgtgtaat atgggaaatg ccgtatgttt ccttatatgg
     6361 cttttggttc gtttctttcg caaacgcttg agttgcgcct cctgccagca gtgcggtagt
     6421 aaaggttaat actgttgctt gttttgcaaa ctttttgatg ttcatcgttc atgtctcctt
     6481 ttttatgtac tgtgttagcg gtctgcttct tccagccctc ctgtttgaag atggcaagtt
     6541 agttacgcac aataaaaaaa gacctaaaat atgtaagggg tgacgccaaa gtatacactt
     6601 tgccctttac acattttagg tcttgcctgc tttatcagta acaaacccgc gcgatttact
     6661 tttcgacctc attctattag actctcgttt ggattgcaac tggtctattt tcctcttttg
     6721 tttgatagaa aatcataaaa ggatttgcag actacgggcc taaagaacta aaaaatctat
     6781 ctgtttcttt tcattctctg tattttttat agtttctgtt gcatgggcat aaagttgcct
     6841 ttttaatcac aattcagaaa atatcataat atctcatttc actaaataat agtgaacggc
     6901 aggtatatgt gatgggttaa aaaggatcga tcctctagcc acgggtgcgc atgatcgtgc
     6961 tcctgtcgtt gaggacccgg ctaggctggc ggggttgcct tactggttag cagaatgaat
     7021 caccgatacg cgagcgaacg tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa
     7081 caacatgaat ggtcttcggt ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc
     7141 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     7201 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     7261 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     7321 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     7381 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     7441 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     7501 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     7561 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     7621 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     7681 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     7741 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
     7801 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     7861 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     7921 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     7981 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     8041 aaatcaatct aaagtatata tgagtaaact tggtctgaca g
//
