LOCUS       Exported                5627 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pRLYC2, complete sequence.
ACCESSION   JQ768507
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5627)
  AUTHORS   Redder P, Linder P.
  TITLE     New Range of Vectors with a Stringent 5-Fluoroorotic Acid-Based 
            Counterselection System for Generating Mutants by Allelic 
            Replacement in Staphylococcus aureus
  JOURNAL   Appl. Environ. Microbiol. 78 (11), 3846-3854 (2012)
  PUBMED    22447609
REFERENCE   2  (bases 1 to 5627)
  AUTHORS   Redder P, Linder P.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-MAR-2012) Department of Microbiology and Molecular 
            Medicine, University of Geneva, Faculty of Medicine, Rue 
            Michel-Servet 1, Geneva 1203, Switzerland
REFERENCE   3  (bases 1 to 5627)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: BioEdit, CAP v. 7.0.9.0
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5627
                     /organism="Cloning vector pRLYC2"
                     /mol_type="other DNA"
                     /db_xref="taxon:1193170"
     CDS             complement(146..469)
                     /codon_start=1
                     /product="LacZalpha A53D"
                     /label=LacZalpha A53D
                     /note="alpha subunit of beta-galactosidase with an A53D 
                     mutation; contains an XcmI site that generates a 3 
                     thymidine overhang and an A53D mutation has been 
                     introduced; this does not block alpha-complementation 
                     activity; allows for blue/white screening"
                     /protein_id="AFK32935.1"
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFDSWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    396..452
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(465..481)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    489..505
                     /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(513..543)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    558..579
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    complement(626..2343)
                     /note="contains pyrF and pyrE genes from Bacillus subtilis 
                     with PspacC promoter; HindIII and BglII sites have been 
                     removed; pyrFE cassette; complements pyrFE deletions in 
                     Staphylococcus aureus"
     misc_feature    complement(2343..3346)
                     /note="CAT194 cassette; chloramphenicol resistance 
                     cassette"
     rep_origin      3457..3667
                     /note="derived from pT181; allows replication in 
                     Staphylococcus aureus if the RepC protein is present; 
                     ORI181"
     rep_origin      3776..4390
                     /note="origin for replication in E. coli; ColE1"
     rep_origin      complement(3808..4396)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4567..5427)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5428..5532)
                     /gene="bla"
                     /label=AmpR promoter