LOCUS       Exported                9274 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pBACi DNA, complete sequence.
ACCESSION   LC127310
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9274)
  AUTHORS   Satoo Y, Hisatsune J, Sakuma T, Yamamoto T, Sugai M.
  TITLE     Novel CRISPR interference technology, an elementary method for 
            inspects of Staphylococcus aureus clinical isolates
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9274)
  AUTHORS   Satoo Y, Hisatsune J, Sugai M.
  TITLE     Direct Submission
  JOURNAL   Submitted (22-FEB-2016) Contact:Motoyuki Sugai Hiroshima University,
            Bacteriology; Minamiku, Kasumi1-2-3, Hiroshima, Hiroshima 734-8551, 
            Japan URL :http://home.hiroshima-u.ac.jp/saikin/index.html
REFERENCE   3  (bases 1 to 9274)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: CLC Genomics Workbench v. 7
            Sequencing Technology :: Illumina MiSeq
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..9274
                     /organism="Cloning vector pBACi"
                     /mol_type="other DNA"
                     /db_xref="taxon:1807767"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        37..67
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    75..91
                     /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)."
     primer_bind     99..115
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   414..449
                     /rpt_unit_seq="gttttagagctatgctgttttgaatggtcccaaaac"
     misc_feature    complement(450..455)
                     /label=BsaI recognition site
                     /note="BsaI recognition site"
     misc_feature    473..478
                     /label=BsaI recognition site
                     /note="BsaI recognition site"
     repeat_region   480..515
                     /rpt_unit_seq="gttttagagctatgctgttttgaatggtcccaaaac"
     misc_feature    complement(516..647)
                     /label=crRNA leader
                     /note="crRNA leader sequence for the Streptococcus pyogenes
                     CRISPR/Cas system"
     CDS             complement(668..4774)
                     /codon_start=1
                     /gene="dcas9"
                     /product="dCas9"
                     /label=dcas9
                     /protein_id="BBC20719.1"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELKNGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     gene            complement(668..4774)
                     /gene="dcas9"
                     /label=dcas9
     ncRNA           4985..5155
                     /product="tracrRNA"
                     /label=other
                     /ncRNA_class="other"
     misc_RNA        5070..5148
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     primer_bind     complement(5456..5472)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      5812..6426
                     /label=Replication origin for E. coli
                     /note="Replication origin for E. coli"
     rep_origin      5823..6411
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6985..7635)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=cat
                     /protein_id="BBC20720.1"
                     /translation="MNFNKIDLDNWKRKEIFNHYLNQQTTFSITTEIDISVLYRNIKQE
                     GYKFYPAFIFLVTRVINSNTAFRTGYNSDGELGYWDKLEPLYTIFDGVSKTFSGIWTPV
                     KNDFKEFYDLYLSDVEKYNGSGKLFPKTPIPENAFSLSIIPWTSFTGFNLNINNNSNYL
                     LPIITAGKFINKGNSIYLPLSLQVHHSVCDGYHAGLFMNSIQELSDRPNDWLL"
     gene            complement(6985..7635)
                     /gene="cat"
                     /label=cat
     CDS             complement(7914..8918)
                     /codon_start=1
                     /gene="rep"
                     /product="replication protein"
                     /label=rep
                     /note="Rep for S. aureus"
                     /protein_id="BBC20721.1"
                     /translation="MGVSFNIMCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDY
                     FELLHILEFKKAERVKDCAEILEYKQNRETGERKLYRVWFCKSRLCPMCNWRRAMKHGI
                     QSQKVVAEVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLV
                     GFMRATEVTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPN
                     VKVQMIRPKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLIS
                     YGGLLKEIHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
     gene            complement(7914..8918)
                     /gene="rep"
                     /label=rep