LOCUS       Exported                6384 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector RHP-AmpS, complete sequence.
ACCESSION   FJ545754
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6384)
  AUTHORS   Haag AF, Ostermeier C.
  TITLE     Positive Selection Vector for Direct Protein Expression
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6384)
  AUTHORS   Haag AF, Ostermeier C.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-DEC-2008) Novartis Institutes for BioMedical Research,
            Novartis Pharma AG, Forum 1, Novartis Campus, Basel CH-4056, 
            Switzerland
REFERENCE   3  (bases 1 to 6384)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6384
                     /organism="Expression vector RHP-AmpS"
                     /mol_type="other DNA"
                     /db_xref="taxon:660654"
     rep_origin      1..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             complement(549..1364)
                     /codon_start=1
                     /product="kanamycin resistance protein"
                     /label=kanamycin resistance protein
                     /note="Kan(R)"
                     /protein_id="ACT98418.1"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      1486..2074
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    2260..2402
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(2504..2695)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3504..4586)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     CDS             complement(3504..4463)
                     /codon_start=1
                     /product="LacI"
                     /label=LacI
                     /protein_id="ACT98419.1"
                     /translation="MAELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSR
                     ADQLGASVVVSMVERSGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPA
                     LFLDVSDQTPINSIIFSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKY
                     LTRNQIQPIAEREGDWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLR
                     VGADISVVGYDDTEDSSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVS
                     LVKRKTTLAPNTQTASPRALADSLMQLARQVSRLESGQ"
     promoter        complement(4587..4664)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        4973..4991
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      4973..4989
                     /regulatory_class="promoter"
                     /note="T7"
     protein_bind    4992..5016
                     /label=lac repressor protein binding site
                     /bound_moiety="lac repressor protein"
                     /note="lac operator"
     RBS             5031..5053
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     regulatory      5048..5053
                     /regulatory_class="ribosome_binding_site"
     misc_feature    5228..5245
                     /label=His6-tag
                     /note="His6-tag"
     CDS             5228..5245
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    5249..5272
                     /label=PreScission protease site
                     /note="PreScission protease site"
     CDS             5249..5272
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(5298..6155)
                     /codon_start=1
                     /product="BlaD290"
                     /label=BlaD290
                     /protein_id="ACT98420.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKH"
     promoter        complement(6156..6227)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      complement(6156..6190)
                     /regulatory_class="promoter"
                     /note="bla"
     regulatory      6290..6308
                     /regulatory_class="terminator"
                     /note="T7"
     terminator      6301..6348
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"