pFR56 vector (V016058)

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LOCUS       62056_10955        7224 bp DNA     circular SYN 02-NOV-2020
DEFINITION  Cloning vector pFR56, complete sequence.
ACCESSION   MT412099
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7224)
  AUTHORS   Rousset F, Fernandez-Rodriguez J, Rocha E, Cabezas-Caballero J, 
            Piastra-Facon F, Clermont O, Denamur E, Bikard D.
  TITLE     Parallel CRISPRi screening of the E. coli core genome reveals a 
            modulation of gene essentiality by horizontal gene transfer
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7224)
  AUTHORS   Rousset F, Fernandez-Rodriguez J, Piastra-Facon F, Bikard D.
  TITLE     Direct Submission
  JOURNAL   Submitted (29-APR-2020) Synthetic Biology Group, Institut Pasteur, 
            28 rue du Dr Roux, Paris 75015, Paris
REFERENCE   3  (bases 1 to 7224)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (29-APR-2020) Synthetic Biology Group, Institut Pasteur, 28 rue du 
            Dr Roux, Paris 75015, Paris"
FEATURES             Location/Qualifiers
     source          1..7224
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      38..73
                     /label=J23110
                     /note="J23110"
                     /regulatory_class="promoter"
     regulatory      78..93
                     /label=B0030 RBS
                     /note="B0030 RBS"
                     /regulatory_class="ribosome_binding_site"
     CDS             99..701
                     /codon_start=1
                     /transl_table=11
                     /product="PhlF repressor"
                     /label=PhlF repressor
                     /protein_id="QOT13791.1"
                     /translation="MARTPSRSSIGSLRSPHTHKAILTSTIEILKECGYSGLSIESVAR
                     RAGASKPTIYRWWTNKAALIAEVYENESEQVRKFPDLGSFKADLDFLLRNLWKVWRETI
                     CGEAFRCVIAEAQLDPATLTQLKDQFMERRREMPKKLVENAISNGELPKDTNRELLLDM
                     IFGFCWYRLLTEQLTVEQDIEEFTFLLINGVCPGTQR"
     regulatory      712..769
                     /label=modified L3S2P11 terminator
                     /note="modified L3S2P11 terminator"
                     /regulatory_class="terminator"
     misc_feature    781..832
                     /label=PhlF promoter
                     /note="PhlF promoter"
     regulatory      797..802
                     /regulatory_class="minus_35_signal"
     misc_feature    802..832
                     /label=phO
                     /note="phO"
     regulatory      820..826
                     /regulatory_class="minus_10_signal"
     misc_feature    832..867
                     /label=RBS
                     /note="RBS"
     CDS             867..4970
                     /label=dCas9
                     /note="catalytically dead mutant of the Cas9 endonuclease
                     from the Streptococcus pyogenes Type II CRISPR/Cas system"
     regulatory      4984..5038
                     /label=terminator ECK120010818
                     /note="terminator ECK120010818"
                     /regulatory_class="terminator"
     misc_feature    5053..5277
                     /label=lambda cos
                     /note="lambda cos"
     misc_feature    complement(5178..5210)
                     /label=cosN site
                     /note="cosN site"
     misc_RNA        complement(5305..5380)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    complement(5381..5401)
                     /label=control spacer with 2 BsaI
                     /note="control spacer with 2 BsaI"
     promoter        complement(5401..5435)
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological
                     Parts BBa_J23119) modified to end with an SpeI site"
     regulatory      complement(5470..5527)
                     /label=terminator ECK120033737
                     /note="terminator ECK120033737"
                     /regulatory_class="terminator"
     CDS             complement(5538..6194)
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     regulatory      complement(6195..6220)
                     /label=synthetic RBS
                     /note="synthetic RBS"
                     /regulatory_class="ribosome_binding_site"
     oriT            complement(6334..6443)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     regulatory      complement(6597..6610)
                     /label=T7 promoter
                     /note="T7 promoter"
                     /regulatory_class="promoter"
     rep_origin      complement(6643..7188)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin."