LOCUS       Exported                8069 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Retroviral golden gate cloning vector for expression of functional 
            Human T-Cell Receptor genes with NGFR.
ACCESSION   .
VERSION     .
KEYWORDS    MSGV Hu Acceptor PGK-NGFR
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8069)
  AUTHORS   Coren LV, Jain S, Trivett MT, Ohlen C, Ott DE
  TITLE     Production of retroviral constructs for effective transfer and 
            expression of T-cell receptor genes using Golden Gate cloning.
  JOURNAL   Biotechniques. 2015 Mar 1;58(3):135-9. doi: 10.2144/000114265. 
            eCollection 2015.
  PUBMED    25757546
REFERENCE   2  (bases 1 to 8069)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.2144/000114265";
            journalName: "Biotechniques"; date: "2015-03-1- 1"; volume: "58"; 
            issue: "3"; pages: "135-9"
FEATURES             Location/Qualifiers
     source          1..8069
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     LTR             1..518
                     /label=5 LTR
                     /note="5 long terminal repeat from murine embryonic stem 
                     cell virus"
     primer_bind     1358..1380
                     /label=pLXSN 5
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1396..1412
                     /label=pBABE 5
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     1750..1766
                     /label=pBMN 5
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     promoter        2911..3409
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2946..2969)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3342..3361
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     misc_feature    4290..4342
                     /label=MCS
                     /note="multiple cloning site"
     LTR             4386..4900
                     /label=3 LTR
                     /note="3 long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5072..5088
                     /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(5096..5126)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5141..5162
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5279..5296)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5450..6038)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5530..5549)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6209..7069)
                     /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     6832..6851
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7070..7174)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7242..7260
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7298..7320)
                     /label=pGEX 3
                     /note="pGEX vectors, reverse primer"
     primer_bind     7420..7439
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7633..7655
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7647..7664
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7648..7664
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7863..7882
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"