LOCUS       Exported                3792 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pLSP2, complete sequence.
ACCESSION   KU936036
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3792)
  AUTHORS   Lehman SS, Mladinich KM, Boonyakanog A, Mima T, Karkhoff-Schweizer 
            RR, Schweizer HP.
  TITLE     Versatile nourseothricin and streptomycin/spectinomycin resistance 
            gene cassettes and their use in chromosome integration vectors
  JOURNAL   J. Microbiol. Methods 129, 8-13 (2016)
  PUBMED    27457407
REFERENCE   2  (bases 1 to 3792)
  AUTHORS   Lehman SS, Mladinich K, Boonyakanog A, Mima T, Karkhoff-Schweizer 
            RR, Schweizer HP.
  TITLE     Direct Submission
  JOURNAL   Submitted (17-MAR-2016) Molecular Genetics and Microbiology, 
            University of Florida, 2055 Mowry Road, Gainesville, FL 32610, USA
REFERENCE   3  (bases 1 to 3792)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..3792
                     /organism="Cloning vector pLSP2"
                     /mol_type="other DNA"
                     /db_xref="taxon:1884651"
     rep_origin      complement(184..772)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(943..1803)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /protein_id="ANY60774.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(943..1803)
                     /gene="bla"
                     /label=bla
     promoter        complement(1804..1908)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2380..2396
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    2400..2444
                     /note="MCS; multiple cloning site"
     misc_feature    2450..2482
                     /note="loxP; Cre recombinase site"
     regulatory      2550..2603
                     /regulatory_class="promoter"
                     /note="chloramphenicol transacetylase gene promoter"
     regulatory      2567..2572
                     /regulatory_class="minus_35_signal"
     regulatory      2590..2595
                     /regulatory_class="minus_10_signal"
     regulatory      2637..2641
                     /regulatory_class="ribosome_binding_site"
     CDS             2649..3401
                     /codon_start=1
                     /gene="aad9"
                     /product="spectinomycin adenyltransferase"
                     /label=aad9
                     /protein_id="ANY60775.1"
                     /translation="MNTYEQINKVKKILRKHLKNNLIGTYMFGSGVESGLKPNSDLDFL
                     VVVSEPLTDQSKEILIQKIRPISKKIGDKSNLRYIELTIIIQQEMVPWNHPPKQEFIYG
                     EWLQELYEQGYIPQKELNSDLTIMLYQAKRKNKRIYGNYDLEELLPDIPFSDVRRAIMD
                     SSEELIDNYQDDETNSILTLCRMILTMDTGKIIPKDIAGNAVAESSPLEHRERILLAVR
                     SYLGENIEWTNENVNLTINYLNNRLKKL"
     gene            2649..3401
                     /gene="aad9"
                     /label=aad9
                     /note="codon optimized; derived from Enterococcus faecalis"
     regulatory      3402..3443
                     /regulatory_class="terminator"
     misc_feature    3479..3512
                     /note="loxP; Cre recombinase site"
     protein_bind    3479..3512
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    3517..3561
                     /note="MCS; multiple cloning site"
     primer_bind     complement(3574..3590)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3598..3614
                     /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(3622..3652)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3667..3688
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."