LOCUS       Exported                6988 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Encodes (N to C): GNSTM glycosylation motif, 3 residue spacer, RVG 
            peptide, 10 residue spacer, Lamp2b (exosomal transmembrane protein),
            3 residue spacer, HA tag..
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA GNSTM-3-RVG-10-Lamp2b-HA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6988)
  AUTHORS   Hung ME, Leonard JN
  TITLE     Stabilization of exosome-targeting peptides via engineered 
            glycosylation.
  JOURNAL   J Biol Chem. 2015 Mar 27;290(13):8166-72. doi: 
            10.1074/jbc.M114.621383. Epub 2015 Feb 5.
  PUBMED    25657008
REFERENCE   2  (bases 1 to 6988)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1074/jbc.M114.621383"; journalName: "J Biol Chem"; date: 
            "2015-03-27- 27"; volume: "290"; issue: "13"; pages: "8166-72"
FEATURES             Location/Qualifiers
     source          1..6988
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        110..489
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        490..693
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     644..664
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     738..757
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        738..756
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      779..788
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2165..2191
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2288..2305)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2294..2518
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2564..2992
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2651..2670)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2861..2882
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3001..3021)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3006..3335
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3186..3321
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3248..3267
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3384..4409
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4539..4660
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4576..4595)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4630..4649
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4709..4725)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4709..4725)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4722..4744)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4733..4749
                     /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(4757..4787)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4802..4823
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4940..4957)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5111..5696)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5191..5210)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5867..6727)
                     /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     6490..6509
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6728..6832)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6907..6926)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"