LOCUS       Exported                6537 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Plasmid vector pRP1028, complete sequence.
ACCESSION   KT823412
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6537)
  AUTHORS   Plaut RD, Stibitz S.
  TITLE     Improvements to a Markerless Allelic Exchange System for Bacillus 
            anthracis
  JOURNAL   PLoS ONE 10 (12), E0142758 (2015)
  PUBMED    26624016
REFERENCE   2  (bases 1 to 6537)
  AUTHORS   Plaut RD, Stibitz S.
  TITLE     Direct Submission
  JOURNAL   Submitted (24-SEP-2015) Center for Biologics Evaluation and 
            Research, OVRR, DBPAP, Food and Drug Administration, 10903 New 
            Hampshire Ave, Bldg 52/72, Rm 3310, Silver Spring, MD 20993, USA
REFERENCE   3  (bases 1 to 6537)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6537
                     /organism="Plasmid vector pRP1028"
                     /mol_type="other DNA"
                     /db_xref="taxon:1764028"
     misc_feature    57..74
                     /label=I-SceI site
                     /note="I-SceI site"
     oriT            90..333
                     /label=from D2BpUC9Cam
                     /note="from D2BpUC9Cam"
     oriT            225..333
                     /note="incP origin of transfer"
     promoter        complement(348..366)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(376..392)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      954..2276
                     /label=ts rep from pJRS233, pWV01
                     /note="ts rep from pJRS233, pWV01"
     CDS             complement(954..1103)
                     /codon_start=1
                     /gene="orfD"
                     /product="OrfD"
                     /label=orfD
                     /protein_id="ALQ43911.1"
                     /translation="MTNKEKELFAENEELKKEIKDLKERIERYREMEVELSTTIDLLRG
                     GIIE"
     gene            complement(954..1103)
                     /gene="orfD"
                     /label=orfD
     gene            complement(1100..1798)
                     /gene="repA"
                     /label=repA
     misc_feature    1623..1640
                     /note="mutations in RepA leading to temperature 
                     sensitivity"
     CDS             complement(1865..2026)
                     /codon_start=1
                     /gene="orfC"
                     /product="orfC"
                     /label=orfC
                     /protein_id="ALQ43915.1"
                     /translation="MVISESKKRVMISLTKEQDKKLTDMAKQKGFSKSAVAALAIEEYA
                     RKESEQKK"
     gene            complement(1865..2026)
                     /gene="orfC"
                     /label=orfC
     CDS             complement(2067..2276)
                     /codon_start=1
                     /gene="orfB"
                     /product="orfB"
                     /label=orfB
                     /protein_id="ALQ43916.1"
                     /translation="MGGKEANFASVLRPPIKCRVPIFVPKTLYPNWLKGLRGFSIANES
                     PTFSPTFFINLYLSSFIVVFMITK"
     gene            complement(2067..2276)
                     /gene="orfB"
                     /label=orfB
     CDS             2410..2421
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        2596..2625
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     regulatory      2596..2601
                     /regulatory_class="minus_35_signal"
                     /note="Ptrc"
     regulatory      2619..2623
                     /regulatory_class="minus_10_signal"
                     /note="Ptrc"
     regulatory      2639..2648
                     /regulatory_class="ribosome_binding_site"
     CDS             2656..3438
                     /codon_start=1
                     /gene="spc"
                     /gene_synonym="aad9"
                     /product="spc"
                     /label=spc
                     /note="from Staphylococcus aureus; aad9"
                     /protein_id="ALQ43912.1"
                     /translation="MSNLINGKIPNQAIQTLKIVKDLFGSSIVGVYLFGSAVNGGLRIN
                     SDVDVLVVVNHSLPQLTRKKLTERLMTISGKIGNTDSVRPLEVTVINRSEVVPWQYPPK
                     REFIYGEWLRGEFENGQIQEPSYDPDLAIVLAQARKNSISLFGPDSSSILVSVPLTDIR
                     RAIKDSLPELIEGIKGDERNVILTLARMWQTVTTGEITSKDVAAEWAIPLLPKEHVTLL
                     DIARKGYRGECDDKWEGLYSKVKALVKYMKNSIETSLN"
     gene            2656..3438
                     /gene="spc"
                     /gene_synonym="aad9"
                     /label=spc
                     /note="from Staphylococcus aureus"
     regulatory      3464..3469
                     /regulatory_class="minus_35_signal"
                     /note="PrrnB"
     regulatory      3487..3492
                     /regulatory_class="minus_10_signal"
                     /note="PrrnB"
     regulatory      3512..3517
                     /regulatory_class="minus_35_signal"
                     /note="PFP2"
     regulatory      3535..3540
                     /regulatory_class="minus_10_signal"
                     /note="PFP2"
     regulatory      3553..3562
                     /regulatory_class="ribosome_binding_site"
     CDS             3571..4284
                     /codon_start=1
                     /gene="rfp"
                     /gene_synonym="TurboRFP"
                     /product="rfp"
                     /label=rfp
                     /note="codon-optimized; TurboRFP"
                     /protein_id="ALQ43913.1"
                     /translation="MRGSGSMSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQ
                     TMKIKVVEGGPLPFAFDILATSFMYGSKAFINHTQGIPDFFKQSFPEGFTWERITTYED
                     GGVLTATQDTSFQNGCIIYNVKINGVNFPSNGPVMQKKTRGWEANTEMLYPADGGLRGH
                     SQMALKLVGGGYLHCSFKTTYRSKKPAKNLKMPGFHFVDHRLERIKEADKETYVEQHEM
                     AVAKYCDLPSKLGHR"
     gene            3571..4284
                     /gene="rfp"
                     /gene_synonym="TurboRFP"
                     /label=rfp
                     /note="codon-optimized"
     CDS             complement(4809..5759)
                     /codon_start=1
                     /gene="rep"
                     /product="rep"
                     /label=rep
                     /note="from pSC101"
                     /protein_id="ALQ43914.1"
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     gene            complement(4809..5759)
                     /gene="rep"
                     /label=rep
                     /note="from pSC101"
     rep_origin      complement(5760..6122)
                     /direction=LEFT
                     /label=ori
                     /note="ori"
     regulatory      complement(5790..5795)
                     /regulatory_class="minus_10_signal"
     rep_origin      5807..6029
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     regulatory      complement(5813..5817)
                     /regulatory_class="minus_35_signal"
     misc_feature    complement(6140..6508)
                     /label=par
                     /note="par"