LOCUS Exported 7151 bp ds-DNA circular SYN 16-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 JOURNAL Exported Aug 16, 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 //