LOCUS       Exported                7151 bp ds-DNA     circular SYN 12-MAY-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
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
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"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     regulatory      3194..3203
                     /regulatory_class="other"
                     /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"
ORIGIN
        1 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
       61 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
      121 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
      181 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
      241 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
      301 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
      361 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
      421 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
      481 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
      541 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
      601 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
      661 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
      721 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
      781 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
      841 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt
      901 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
      961 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag
     1021 cgcgcaatta accctcacta aagggaacaa aagctggagc tgcaagctta atgtagtctt
     1081 atgcaatact cttgtagtct tgcaacatgg taacgatgag ttagcaacat gccttacaag
     1141 gagagaaaaa gcaccgtgca tgccgattgg tggaagtaag gtggtacgat cgtgccttat
     1201 taggaaggca acagacgggt ctgacatgga ttggacgaac cactgaattg ccgcattgca
     1261 gagatattgt atttaagtgc ctagctcgat acataaacgg gtctctctgg ttagaccaga
     1321 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
     1381 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
     1441 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt
     1501 gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg
     1561 cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg
     1621 ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg
     1681 atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt
     1741 atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga
     1801 aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat cagaagaact
     1861 tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa
     1921 agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc
     1981 acagcaagcg gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa
     2041 gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg
     2101 caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg
     2161 ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg
     2221 ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg
     2281 cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc
     2341 tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa
     2401 aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac
     2461 agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct
     2521 taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat
     2581 tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt
     2641 atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg
     2701 tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc
     2761 tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt ggagagagag
     2821 acagagacag atccattcga ttagtgaacg gatctcgacg gtatcggtta acttttaaaa
     2881 gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag
     2941 acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt atcgatcacg
     3001 agactagcct cgatagcccg ggctcgagca gctgaagctt gcatgcctgc aggatcaaag
     3061 ggggtaagat caaagggggt aagatcaaag gtctagaggg tatataatgg atccggtact
     3121 cgagatctgc gatctaagta agcttggcat tccggtactg ttggtaaagc caccatggga
     3181 tgatccaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     3241 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     3301 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
     3361 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
     3421 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     3481 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     3541 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     3601 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     3661 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     3721 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     3781 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
     3841 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     3901 ggacgagctg tacaagaagc ttagccatgg cttcccgccg gaggtggagg agcaggatga
     3961 tggcacgctg cccatgtctt gtgcccagga gagcgggatg gaccgtcacc ctgcagcctg
     4021 tgcttctgct aggatcaatg tgtaggcggc cgcctgcagg tcgacaatca acctctggat
     4081 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     4141 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     4201 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     4261 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     4321 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     4381 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     4441 tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc
     4501 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     4561 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     4621 acgagtcgga tctccctttg ggccgcctcc ccgcctggaa ttcgagctcg gtacctttaa
     4681 gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac
     4741 tggaagggct aattcactcc caacgaagac aagatctgct ttttgcttgt actgggtctc
     4801 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
     4861 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
     4921 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtagta
     4981 gttcatgtca tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg
     5041 agaggaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     5101 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     5161 tatcttatca tgtctggctc tagctatccc gcccctaact ccgcccatcc cgcccctaac
     5221 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     5281 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     5341 cctagggacg tacccaattc gccctatagt gagtcgtatt acgcgcgctc actggccgtc
     5401 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     5461 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     5521 cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg
     5581 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     5641 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     5701 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     5761 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     5821 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     5881 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     5941 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca
     6001 atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     6061 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     6121 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     6181 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     6241 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     6301 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     6361 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     6421 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     6481 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     6541 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     6601 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     6661 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6721 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6781 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6841 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6901 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6961 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     7021 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     7081 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     7141 ccgtagaaaa g
//
