GAPTrapT2AIresMygro vector (V009961)

LOCUS       40924_1034       12226 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector GAPTrapT2AIresMygro, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12226)
  AUTHORS   Kao T, Labonne T, Niclis JC, Chaurasia R, Lokmic Z, Qian E, Bruveris
            FF, Howden SE, Motazedian A, Schiesser JV, Costa M, Sourris K, Ng E,
            Anderson D, Giudice A, Farlie P, Cheung M, Lamande SR, Penington AJ,
            Parish CL, Thomson LH, Rafii A, Elliott DA, Elefanty AG, Stanley EG.
  TITLE     GAPTrap: A Simple Expression System for Pluripotent Stem Cells and 
            Their Derivatives
  JOURNAL   Stem Cell Reports (2016) In press
  PUBMED    27594589
REFERENCE   2  (bases 1 to 12226)
  AUTHORS   Stanley E.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-AUG-2016) Cell Biology, Murdoch Childrens Research 
            Institute, Royal Children's Hospital, Flemington Road, Melbourne, 
            Victoria 3052, Australia
REFERENCE   3  (bases 1 to 12226)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 12226)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Stem Cell 
            Reports (2016) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (25-AUG-2016) Cell Biology, Murdoch Childrens Research Institute, 
            Royal Children's Hospital, Flemington Road, Melbourne, Victoria 
            3052, Australia"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..12226
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        24..128
                     /label=AmpR promoter
     CDS             129..986
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1160..1748
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    2036..2057
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        2072..2102
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     misc_feature    2110..2126
                     /label=lac operator
                     /note="lac operator"
     protein_bind    2110..2126
                     /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)."
     primer_bind     2134..2150
                     /label=M13 rev
                     /note="M13 rev"
     promoter        2171..2189
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     gene            2216..5559
                     /gene="GAPDH"
                     /label=GAPDH
                     /note="glyceraldehyde-3-phosphate dehydrogenase"
     misc_feature    2219..5556
                     /gene="GAPDH"
                     /label=left arm
                     /note="left arm"
     CDS             5584..5637
                     /codon_start=1
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
                     /translation="EGRGSLLTCGDVEENPGP"
     misc_feature    5660..6246
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             6256..7278
                     /codon_start=1
                     /label=HygR
                     /note="aminoglycoside phosphotransferase from E. coli"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     misc_feature    7310..11545
                     /label=right arm
                     /note="right arm"
     promoter        complement(11583..11601)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(11611..11627)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(11768..12223)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"