pEXPR71 vector (V006403)

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LOCUS       40924_18926        8047 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Mammalian expression vector pEXPR71, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8047)
  AUTHORS   Siciliano V.
  TITLE     Engineering modular intracellular protein sensor-actuator devices
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8047)
  AUTHORS   Siciliano V.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-APR-2018) Biological Engineering, Massachusetts 
            Institute of Technology, 500 Tech Square, Cambridge, MA 02139, USA
REFERENCE   3  (bases 1 to 8047)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8047)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (02-APR-2018) Biological Engineering, Massachusetts Institute of 
            Technology, 500 Tech Square, Cambridge, MA 02139, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8047
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(222..810)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(984..1841)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(1842..1946)
                     /label=AmpR promoter
     rep_origin      complement(1972..2427)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     2569..2585
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2595..2613
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    2628..3431
                     /label=HA-L
                     /note="left homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     misc_feature    3510..3535
                     /label=SA
                     /note="splice acceptor site"
     protein_bind    4207..4227
                     /label=attB4
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     protein_bind    4272..4290
                     /label=tet operator
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4307..4325
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4343..4361
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4379..4397
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4414..4432
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4450..4468
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        4480..4518
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     protein_bind    4558..4582
                     /label=attB1
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4598..4921
                     /codon_start=1
                     /label=DD
                     /note="destabilization domain that can be stabilized by
                     Shield1 in the ProteoTuner(TM) system"
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKPE"
     CDS             4934..5635
                     /codon_start=1
                     /label=TEV protease
                     /note="tobacco etch virus protease (Kapust et al., 2001)"
                     /translation="SLFKGPRDYNPISSTICHLTNESDGHTTSLYGIGFGPFIITNKHL
                     FRRNNGTLLVQSLHGVFKVKNTTTLQQHLIDGRDMIIIRMPKDFPPFPQKLKFREPQRE
                     ERICLVTTNFQTKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFIVG
                     IHSASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPEEP
                     FQPVKEATQLMN"
     CDS             6050..6067
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     protein_bind    complement(6080..6104)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    6259..6314
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(6675..6691)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6699..6715
                     /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(6723..6753)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6768..6789
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    6962..7798
                     /label=HA-R
                     /note="right homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     promoter        complement(7828..7846)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7867..7883)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(7891..7907)
                     /label=lac operator
                     /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(7915..7945)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(7960..7981)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."