LOCUS       Exported                6485 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  mammalian expression of CTLA4-mEos2.
ACCESSION   .
VERSION     .
KEYWORDS    pirespuro2-CTLA4-mEos2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6485)
  AUTHORS   Fricke F, Beaudouin J, Eils R, Heilemann M
  TITLE     One, two or three? Probing the stoichiometry of membrane proteins by
            single-molecule localization microscopy.
  JOURNAL   Sci Rep. 2015 Sep 11;5:14072. doi: 10.1038/srep14072.
  PUBMED    26358640
REFERENCE   2  (bases 1 to 6485)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep."; 
            date: "2015-09-11"; volume: "5:14072. doi"; pages: " 
            10.1038/srep14072"
FEATURES             Location/Qualifiers
     source          1..6485
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        11..390
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        391..594
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     545..565
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     639..658
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        639..657
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      709..718
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     intron          2128..2357
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2340..2359
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    2518..2980
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2574..2591)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2801..2820
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3013..3612
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3013..3032)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3509..3529
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(3748..3765)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3754..3978
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     4025..4044
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4104..4120)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4104..4120)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4117..4139)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4128..4144
                     /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(4152..4182)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4197..4218
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4335..4352)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4506..5094)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4586..4605)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5265..6125)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5888..5907
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6126..6230)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6305..6324)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"