LOCUS       Exported                9297 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Site-specific excision vector pFLP2, complete sequence.
ACCESSION   AF048702
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9297)
  AUTHORS   Hoang TT, Karkhoff-Schweizer RR, Kutchma AJ, Schweizer HP.
  TITLE     A broad-host-range Flp-FRT recombination system for site-specific 
            excision of chromosomally-located DNA sequences: application for 
            isolation of unmarked Pseudomonas aeruginosa mutants
  JOURNAL   Gene 212 (1), 77-86 (1998)
  PUBMED    9661666
REFERENCE   2  (bases 1 to 9297)
  AUTHORS   Hoang TT, Karkhoff-Schweizer RR, Kutchma AJ, Schweizer HP.
  TITLE     Direct Submission
  JOURNAL   Submitted (17-FEB-1998) Microbiology, Colorado State University, 
            Fort Collins, CO 80523, USA
REFERENCE   3  (bases 1 to 9297)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..9297
                     /organism="Site-specific excision vector pFLP2"
                     /plasmid="pFLP2"
                     /mol_type="genomic DNA"
                     /db_xref="taxon:73560"
     rep_origin      194..596
                     /note="region containing broad-host-range origin of 
                     replication from pRO1600"
     rep_origin      232..583
                     /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             597..1430
                     /codon_start=1
                     /gene="rep"
                     /product="replication protein"
                     /label=rep
                     /protein_id="AAC27030.1"
                     /translation="MASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     gene            597..1430
                     /gene="rep"
                     /label=rep
     primer_bind     1591..1607
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2338..3759)
                     /codon_start=1
                     /gene="sacB"
                     /product="levansucrase"
                     /label=sacB
                     /protein_id="AAC27031.1"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     gene            complement(2338..3759)
                     /gene="sacB"
                     /label=sacB
     promoter        complement(3760..4205)
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             complement(4426..5697)
                     /codon_start=1
                     /gene="flp"
                     /product="Flp recombinase"
                     /label=flp
                     /protein_id="AAC27032.1"
                     /translation="MPQFGILCKTPPKVLVRQFVERFERPSGEKIALCAAELTYLCWMI
                     THNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKKLIPAWEFTI
                     IPYYGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESIWEITEKILN
                     SFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGVIIQCLVTET
                     KTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEYQLLKDNLVR
                     SYNKALKKNAPYSIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWSDKRASAVAR
                     TTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQLKGSAEGSIR
                     YPAWNGIISQEVLDYLSSYINRRI"
     gene            complement(4426..5697)
                     /gene="flp"
                     /label=flp
     RBS             5701..5709
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3 end of the 16S rRNA (Chen et al., 1994)"
     CDS             5798..6511
                     /codon_start=1
                     /gene="cI"
                     /allele="cI-857"
                     /product="lambda repressor"
                     /label=cI
                     /protein_id="AAC27033.1"
                     /translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ
                     SGVGALFNGINALNAYNAALLTKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY
                     PVFSHVQAGMFSPKLRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD
                     GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV
                     VGKVIASQWPEETFG"
     gene            5798..6511
                     /gene="cI"
                     /allele="cI-857"
                     /label=cI
     primer_bind     complement(6800..6816)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6824..6840
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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(6848..6878)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6893..6914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     oriT            7152..7408
                     /label=region containing oriT
                     /note="region containing oriT"
     oriT            7298..7406
                     /note="incP origin of transfer"
     rep_origin      complement(7478..8066)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(8237..9097)
                     /codon_start=1
                     /gene="bla"
                     /product="beta lactamase"
                     /label=bla
                     /protein_id="AAC27034.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(8237..9097)
                     /gene="bla"
                     /label=bla
     promoter        complement(9098..9202)
                     /gene="bla"
                     /label=AmpR promoter