LOCUS       Exported                7151 bp ds-DNA     circular SYN 17-AUG-2021
DEFINITION  Beta-catenin reporter - 3rd generation self-inactivating lentiviral 
            plasmid LEF-1/TCF-responsive promoter driven d2-eGFP (destabilized, 
            half-life of 2h)..
ACCESSION   .
VERSION     .
KEYWORDS    Lentiviral-TOP-dGFP-reporter
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7151)
  AUTHORS   Reya T, Duncan AW, Ailles L, Domen J, Scherer DC, Willert K, Hintz 
            L, Nusse R, Weissman IL
  TITLE     A role for Wnt signalling in self-renewal of haematopoietic stem 
            cells.
  JOURNAL   Nature. 2003 May 22. 423(6938):409-14.
  PUBMED    12717450
REFERENCE   2  (bases 1 to 7151)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2003 May 22. 423(6938):409-14."
FEATURES             Location/Qualifiers
     source          1..7151
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      15..603
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     504..523
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     757..774
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    891..912
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        927..957
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    965..981
                     /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     970..992
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     989..1005
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     989..1005
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1024..1044
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1026..1044
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        1072..1298
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             1299..1479
                     /label=5 LTR (truncated)
                     /note="truncated 5 long terminal repeat (LTR) from HIV-1"
     misc_feature    1526..1651
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2144..2377
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2562..2606
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2873..2990
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     regulatory      3169..3178
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     regulatory      3194..3203
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3200..4045
                     /codon_start=1
                     /product="EGFP destabilized by fusion to residues 422-461 
                     of mouse ornithine decarboxylase, giving an in vivo 
                     half-life of ~2 hours"
                     /label=d2EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKKLSHGFPPEVEEQDDGTLPMSCAQESGMDRHPAACASARINV
                     "
     primer_bind     complement(3245..3266)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3506..3525)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3853..3874
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    4066..4654
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4119..4139)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4537..4548)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4741..4974
                     /label=3 LTR (Delta-U3)
                     /note="self-inactivating 3 long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5046..5167
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5083..5102)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5137..5156
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5207..5342
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5269..5288
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(5362..5381)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5363..5381)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5391..5408)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5391..5407)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5400..5422)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5549..6004
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5636..5655)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5846..5867
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6030..6134
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6135..6995
                     /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     complement(6353..6372)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"