phr5-AcIE1-iFlp vector (V015913)

LOCUS       62056_12460        5100 bp DNA     circular SYN 24-MAY-2021
DEFINITION  Cloning vector phr5-AcIE1-iFlp, complete sequence.
ACCESSION   MW246025
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5100)
  AUTHORS   Dias MM, Vidigal J, Sequeira DP, Alves PM, Teixeira AP, Roldao A.
  TITLE     Insect High FiveTM cell line development using site-specific flipase
            recombination technology
  JOURNAL   G3 (Bethesda) (2021) In press
  PUBMED    33982066
REFERENCE   2  (bases 1 to 5100)
  AUTHORS   Dias MM, Vidigal J, Sequeira DP, Alves PM, Teixeira AP, Roldao A.
  TITLE     Direct Submission
  JOURNAL   Submitted (11-NOV-2020) Animal Cell Technology Unit, iBET, Instituto
            de Biologia Experimental e Tecnologica, Av. da Republica, Qta. do 
            Marques, ITQB/iBET building, Oeiras 2780-157, Portugal
REFERENCE   3  (bases 1 to 5100)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "G3 
            (Bethesda) (2021) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (11-NOV-2020) Animal Cell Technology Unit, iBET, Instituto de 
            Biologia Experimental e Tecnologica, Av. da Republica, Qta. do 
            Marques, ITQB/iBET building, Oeiras 2780-157, Portugal"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5100
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..483
                     /label=hr5 enhancer
                     /note="baculovirus early transcription enhancer"
     promoter        487..1078
                     /label=IE1 promoter
                     /note="promoter of the ie1 gene from the baculovirus
                     Autographa californica"
     sig_peptide     1084..1143
                     /label=IgM signal sequence
                     /note="signal sequence from mouse IgM heavy chain"
     CDS             1156..1179
                     /codon_start=1
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
                     /translation="WSHPQFEK"
     CDS             1186..1200
                     /codon_start=1
                     /label=enterokinase site
                     /note="enterokinase recognition and cleavage site"
                     /translation="DDDDK"
     CDS             1218..2513
                     /codon_start=1
                     /label=FLPo
                     /note="nuclear-targeted site-specific recombinase"
                     /translation="MAPKKKRKVMPQFGILCKTPPKVLVRQFVERFERPSGEKIASCAA
                     ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK
                     LIPAWEFTIIPYYGQKHQSDITDIVSRLQLQFESSEEADKGNSRSKKMLKALLSEGESI
                     WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV
                     IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY
                     QLLKDNLVRSYNKALKKNAPYSIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS
                     DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL
                     KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI"
     CDS             2569..2595
                     /codon_start=1
                     /label=9xHis
                     /note="9xHis affinity tag"
                     /translation="HHHHHHHHH"
     terminator      2617..2924
                     /label=IE1 terminator
                     /note="terminator of the ie1 gene from the baculovirus 
                     Autographa californica"
     promoter        2954..3058
                     /label=AmpR promoter
     CDS             3059..3916
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4090..4678
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    4966..4987
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        5002..5032
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5040..5056
                     /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)."
     primer_bind     5064..5080
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"