LOCUS       40924_20251        6611 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Site-specific excision vector pFLPe2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6611)
  AUTHORS   Choi KH, Mima T, Casart Y, Rholl D, Kumar A, Beacham IR, Schweizer 
            HP.
  TITLE     Genetic tools for select-agent-compliant manipulation of 
            Burkholderia pseudomallei
  JOURNAL   Appl. Environ. Microbiol. 74 (4), 1064-1075 (2008)
  PUBMED    18156318
REFERENCE   2  (bases 1 to 6611)
  AUTHORS   Choi K-H., Schweizer HP.
  TITLE     Genetic tools for Pseudomonas and its related bacteria
  JOURNAL   Unpublished
REFERENCE   3  (bases 1 to 6611)
  AUTHORS   Choi K-H., Schweizer HP.
  TITLE     Direct Submission
  JOURNAL   Submitted (21-SEP-2007) Microbiology, Immunology and Pathology, 
            Colorado State University, 1682 Campus Delivery, Ft. Collins, CO 
            80523-1682, USA
REFERENCE   4  (bases 1 to 6611)
  TITLE     Direct Submission
REFERENCE   5  (bases 1 to 6611)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Appl. 
            Environ. Microbiol."; date: "2008"; volume: "74"; issue: "4"; pages:
            "1064-1075"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 3; type: "Journal Article"; journalName: "Submitted 
            (21-SEP-2007) Microbiology, Immunology and Pathology, Colorado State
            University, 1682 Campus Delivery, Ft. Collins, CO 80523-1682, USA"
COMMENT     SGRef: number: 4; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(86..457)
                     /codon_start=1
                     /label=BleoR
                     /note="antibiotic-binding protein"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(476..523)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     rep_origin      complement(643..994)
                     /direction=LEFT
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
     CDS             1008..1838
                     /codon_start=1
                     /label=pRO1600 Rep
                     /note="replication protein for the broad-host-range plasmid
                     pRO1600 from Pseudomonas aeruginosa"
                     /translation="VASPPMVYKSNALVEAAYCLSVQEQRIVLASISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMGSTHAIRLYELLTQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKARKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     CDS             complement(1949..3229)
                     /codon_start=1
                     /label=FLPo
                     /note="nuclear-targeted site-specific recombinase"
                     /translation="KEIYMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAAELTYL
                     CWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKKLIPAW
                     EFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESIWEITE
                     KILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGVIIQCL
                     VTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEYQLLKD
                     NLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWSDKRAS
                     AVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQLKGSAE
                     GSIRYPAWNGIISQEVLDYLSSYINRRI"
     promoter        complement(3242..3360)
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             3506..4339
                     /codon_start=1
                     /label=rhaS
                     /note="positive regulator of the rhaB promoter"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
     CDS             4416..5261
                     /codon_start=1
                     /label=rhaR
                     /note="transcriptional activator of rhaR and rhaS"
                     /translation="VAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
                     RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
                     PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
                     YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
                     QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPSQWRHLNSQK
                     D"
     primer_bind     complement(5298..5314)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5322..5338)
                     /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(5346..5376)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5391..5412)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     oriT            5796..5904
                     /label=oriT
                     /note="incP origin of transfer"
     rep_origin      complement(5976..6564)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cggtcgagcg gccgccagtg tgatggataa ttcggcttat ctctgatgtt acattgcaca
       61 agataaaata atatcatcat gatcagtcct gctcctcggc cacgaagtgc acgcagttgc
      121 cggccgggtc gcgcagggcg aactcccgcc cccacggctg ctcgccgatc tcggtcatgg
      181 ccggcccgga ggcgtcccgg aagttcgtgg acacgacctc cgaccactcg gcgtacagct
      241 cgtccaggcc gcgcacccac acccaggcca gggtgttgtc cggcaccacc tggtcctgga
      301 ccgcgctgat gaacagggtc acgtcgtccc ggaccacacc ggcgaagtcg tcctccacga
      361 agtcccggga gaacccgagc cggtcggtcc agaactcgac cgctccggcg acgtcgcgcg
      421 cggtgagcac cggaacggca ctggtcaact tggccatggt ttagttcctc accttgtcgt
      481 attatactat gccgatatac tatgccgatg attaattgtc aacacgtgct gatcatgacc
      541 aaaatccctt aacgtgagtt acgcgtcgaa gccgaattat ctgcagaatt cggcttttcg
      601 tgaattcgcc gctggtgccg ctggttggac gccaagggtg aatccgcctc gataccctga
      661 ttactcgctt cctgcgccct ctcaggcggc gataggggac tggtaaaacg gggattgccc
      721 agacgcctcc cccgcccctt caggggcaca aatgcggccc caacggggcc acgtagtggt
      781 gcgttttttg cgtttccacc cttttcttcc ttttcccttt taaacctttt aggacgtcta
      841 caggccacgt aatccgtggc ctgtagagtt taaaaaggga cggatttgtt gccattaagg
      901 gacggatttg ttgttaagaa gggacggatt tgttgttgta aagggacgga tttgttgtat
      961 tgtgggacgc tgatacagtg tccccttata cacaaggaat gtcgaacgtg gcctcacccc
     1021 caatggttta caaaagcaat gccctggtcg aggccgcgta ttgcctcagt gttcaggaac
     1081 agcggatcgt tctggccagt attagccagg tgaagaggag cgagcctgtc accgatgaag
     1141 tgatgtattc agtgacggcg gaggacatag cgacgatggc gggtgtccct atcgaatctt
     1201 cctacaacca gctcaaagaa gcggccctgc gcctgaaacg gcgggaagtc cggttaaccc
     1261 aagagcccaa tggcaagggg aaaagaccga gtgtgatgat taccggctgg gtgcaaacaa
     1321 tcatctaccg ggagggtgag ggccgtgtag aactcaggtt caccaaagac atgctgccgt
     1381 atctgacgga actcaccaaa cagttcacca aatacgcctt ggctgacgtg gccaagatgg
     1441 gcagcaccca cgcgatcagg ctttacgagc tgctcacgca atgggacagc atcggccagc
     1501 gtgaaataga aattgaccag ctgcgaaagt ggtttcaact ggaaggccgg tatccctcga
     1561 tcaaggactt caagttgcga gtgcttgatc cagccgtgac gcagatcaac gagcacagcc
     1621 cgctacaggt ggagtgggcg cagcgaaaga ccgggcgcaa ggtcacacat ctgttgttca
     1681 gttttggacc gaagaagccc gccaaggcgg tgggtaaggc ccctgcgaag cgcaaggcca
     1741 ggaagatttc agatgctgag atcgcgaaac aggctcgccc tggtgagaca tgggaagcgg
     1801 cccgcgctcg actaacccag atgccgctgg atctggccta gaggccaagc cgaattccag
     1861 cacactggcg gccgttacta gtggatccga gctcggatcc actagtaacg gccgccagtg
     1921 tgctggaatt cggcttgatg atggcctata tgcgtctatt tatgtaggat gaaaggtagt
     1981 ctagtacctc ctgtgatatt atcccattcc atgcggggta tcgtatgctt ccttcagcac
     2041 taccctttag ctgttctata tgctgccact cctcaattgg attagtctca tccttcaatg
     2101 ctatcatttc ctttgatatt ggatcatatg catagtaccg agaaactagt gcgaagtagt
     2161 gatcaggtat tgctgttatc tgatgagtat acgttgtcct ggccacggca gaagcacgct
     2221 tatcgctcca atttcccaca acattagtca actccgttag gcccttcatt gaaagaaatg
     2281 aggtcatcaa atgtcttcca atgtgagatt ttgggccatt ttttatagca aagatcggat
     2341 aaggcgcatt tttcttcaaa gctttattgt acgatctgac taagttatct tttaataatt
     2401 ggtattcctg tttattgctt gaagaattgc cggtcctatt tactcgtttt aggactggtt
     2461 cagaattcct caaaaattca tccaaatata caagtggatc gatcctaccc cttgcgctaa
     2521 agaagtatat gtgcctacta acgcttgtct ttgtctctgt cactaaacac tggattatta
     2581 ctcccagata cttattttgg actaatttaa atgatttcgg atcaacgttc ttaatatcgc
     2641 tgaatcttcc acaattgatg aaagtagcta ggaagaggaa ttggtataaa gtttttgttt
     2701 ttgtaaatct cgaagtatac tcaaacgaat ttagtatttt ctcagtgatc tcccagatgc
     2761 tttcaccctc acttagaagt gctttaagca tttttttact gtggctattt cccttatctg
     2821 cttcttccga tgattcgaac tgtaattgca aactacttac aatatcagtg atatcagatt
     2881 gatgtttttg tccgttgtaa ggaataattg taaattccca agcaggaatc aatttcttta
     2941 atgaggcttc cagaattgtt gctttttgcg tcttgtattt aaactggagt gatttattga
     3001 caatatcgaa actcagcgaa ttgcttatga tagtattata gctcatgaat gtggctctct
     3061 tgattgctgt tccgttatgt gtaatcatcc aacataaata ggttagttca gcagcacagg
     3121 atgctatttt ctcacctgaa ggtctttcaa acctttccac aaactgacga acaagcacct
     3181 taggcggggt tttacacagg atatcgaact ggctcatgta tatctccttg ctgaatttca
     3241 ttacgaccag tctaaaaagc gcctgaattc gcgaccttct cgttactgac aggaaaatgg
     3301 gccattggca accagggaaa gatgaacgtg atgatgttca caatttgctg aattgtggtg
     3361 atgtgatgct caccgcattt cctgaaaatt cacgctgtat cttgaaaaat cgacgttttt
     3421 tacgtggttt tccgtcgaaa atttaaggta agaacctgac ctcgtgatta ctatttcgcc
     3481 gtgttgacga catcaggagg ccagtatgac cgtattacat agtgtggatt tttttccgtc
     3541 tggtaacgcg tccgtggcga tagaaccccg gctcccgcag gcggattttc ctgaacatca
     3601 tcatgatttt catgaaattg tgattgtcga acatggcacg ggtattcatg tgtttaatgg
     3661 gcagccctat accatcaccg gtggcacggt ctgtttcgta cgcgatcatg atcggcatct
     3721 gtatgaacat accgataatc tgtgtctgac caatgtgctg tatcgctcgc cggatcgatt
     3781 tcagtttctc gccgggctga atcagttgct gccacaagag ctggatgggc agtatccgtc
     3841 tcactggcgc gttaaccaca gcgtattgca gcaggtgcga cagctggttg cacagatgga
     3901 acagcaggaa ggggaaaatg atttaccctc gaccgccagt cgcgagatct tgtttatgca
     3961 attactgctc ttgctgcgta aaagcagttt gcaggagaac ctggaaaaca gcgcatcacg
     4021 tctcaacttg cttctggcct ggctggagga ccattttgcc gatgaggtga attgggatgc
     4081 cgtggcggat caattttctc tttcactgcg tacgctacat cggcagctta agcagcaaac
     4141 gggactgacg cctcagcgat acctgaaccg cctgcgactg atgaaagccc gacatctgct
     4201 acgccacagc gaggccagcg ttactgacat cgcctatcgc tgtggattca gcgacagtaa
     4261 ccacttttcg acgctttttc gccgagagtt taactggtca ccgcgtgata ttcgccaggg
     4321 acgggatggc tttctgcaat aacgcgaatc ttctcaacgt atttgtacgc catattgcga
     4381 ataatcaact tcgttctctg gccgaggtag ccacggtggc gcatcagtta aaacttctca
     4441 aagatgattt ttttgccagc gaccagcagg cagtcgctgt ggctgaccgt tatccgcaag
     4501 atgtctttgc tgaacataca catgattttt gtgagctggt gattgtctgg cgcggtaatg
     4561 gcctgcatgt actcaacgat cgcccttatc gcattacccg tggcgatctc ttttacattc
     4621 atgctgacga taaacactcc tacgcttccg ttaacgatct ggttttgcag aatattattt
     4681 attgcccgga gcgtctgaag ctgaatcttg actggcaggg ggcgattccg ggatttaacg
     4741 ccagcgcagg gcaaccacac tggcgcttag gtagcatggg gatggcgcag gcgcggcagg
     4801 ttatcggtca gcttgagcat gaaagtagtc agcatgtgcc gtttgctaac gaaatggctg
     4861 agttgctgtt cgggcagttg gtgatgttgc tgaatcgcca tcgttacacc agtgattcgt
     4921 tgccgccaac atccagcgaa acgttgctgg ataagctgat tacccggctg gcggctagcc
     4981 tgaaaagtcc ctttgcgctg gataaatttt gtgatgaggc atcgtgcagt gagcgcgttt
     5041 tgcgtcagca atttcgccag cagactggaa tgaccatcaa tcaatatctg cgacaggtca
     5101 gagtgtgtca tgcgcaatat cttctccagc atagccgcct gttaatcagt gatatttcga
     5161 ccgaatgtgg ctttgaagat agtaactatt tttcggtggt gtttacccgg gaaaccggga
     5221 tgacgcccag ccagtggcgt catctcaatt cgcagaaaga ttaaatgcat atgatcgagc
     5281 tcgaattcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5341 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5401 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5461 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5521 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5581 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5641 atggggattc cttaaggtat actttccgct gcataaccct gcttcggggt cattatagcg
     5701 attttttcgg tatatccatc ctttttcgca cgatatacag gattttgcca aagggttcgt
     5761 gtagactttc cttggtgtat ccaacggcgt cagccgggca ggataggtga agtaggccca
     5821 cccgcgagcg ggtgttcctt cttcactgtc ccttattcgc acctggcggt gctcaacggg
     5881 aatcctgctc tgcgaggctg gccgataagc tagcttatgt gagcaaaagg ccagcaaaag
     5941 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6001 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     6061 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     6121 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6181 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6241 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6301 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6361 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     6421 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6481 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6541 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6601 cagtggaacg a
//
