LOCUS Exported 7116 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS TOP-GFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7116) AUTHORS Horst D, Chen J, Morikawa T, Ogino S, Kirchner T, Shivdasani RA TITLE Differential WNT activity in colorectal cancer confers limited tumorigenic potential and is regulated by MAPK signaling. JOURNAL Cancer Res. 2012 Feb 8. PUBMED 22318865 REFERENCE 2 (bases 1 to 7116) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7116 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 8..234 /label=RSV promoter /note="Rous sarcoma virus enhancer/promoter" LTR 235..415 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 462..587 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1080..1313 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 1498..1542 /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 1809..1926 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2132..2163 /label=minP /note="minimal TATA-box promoter with low basal activity" regulatory 2226..2235 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2232..2951 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 2232..2948 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2277..2298) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2538..2557) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2885..2906 /label=EGFP-C /note="EGFP, forward primer" misc_feature 2967..3555 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3020..3040) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3438..3449) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 3642..3875 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 3947..4068 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3984..4003) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4038..4057 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin 4108..4243 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4170..4189 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(4263..4282) /label=T7 /note="T7 promoter, forward primer" promoter complement(4264..4282) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(4292..4309) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(4292..4308) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4301..4323) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4450..4905 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4537..4556) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4747..4768 /label=F1ori-F /note="F1 origin, forward primer" promoter 4931..5035 /gene="bla" /label=AmpR promoter CDS 5036..5896 /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(5254..5273) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6067..6655 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6556..6575 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6809..6826 /label=L4440 /note="L4440 vector, forward primer" protein_bind 6943..6964 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6979..7009 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 7017..7033 /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 7022..7044 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 7041..7057 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 7041..7057 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 7076..7096 /label=T3 /note="T3 promoter, forward primer" promoter 7078..7096 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" ORIGIN 1 agcttaatgt agtcttatgc aatactcttg tagtcttgca acatggtaac gatgagttag 61 caacatgcct tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg 121 tacgatcgtg ccttattagg aaggcaacag acgggtctga catggattgg acgaaccact 181 gaattgccgc attgcagaga tattgtattt aagtgcctag ctcgatacat aaacgggtct 241 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt 301 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac 361 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc 421 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc 481 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa 541 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg 601 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata 661 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc 721 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga 781 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc 841 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca 901 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg 961 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga 1021 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata 1081 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg 1141 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg 1201 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag 1261 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt 1321 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt 1381 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt 1441 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag 1501 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata 1561 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta 1621 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta 1681 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa 1741 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat 1801 cggttaactt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta 1861 gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac aaaaattcaa 1921 aattttatcg atcacgagac tagcctcgag aagcttgata tcacgacgcg tggtaccgag 1981 ctcttacgcg agatcaaagg gggtaagatc aaagggggta agatcaaagg ggcgcgagat 2041 caaagggggt aagatcaaag ggggtaagat caaagggggt aagatcaaag gggcgcgccc 2101 gcgtgctagc ccgggctcga gatctagact ctagagggta tataatggaa gctcgaattc 2161 cagcttggca ttccggtact gttggtaaaa agcttggcat tccggtactg ttggatccac 2221 cggtcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg 2281 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg 2341 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc 2401 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg 2461 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc 2521 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg 2581 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca 2641 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca 2701 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg 2761 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc 2821 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg 2881 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc 2941 tgtacaagta aagcggccgc gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat 3001 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc 3061 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct 3121 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca 3181 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt 3241 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg 3301 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga 3361 agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt 3421 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc 3481 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt 3541 gggccgcctc cccgcctgga attcgagctc ggtaccttta agaccaatga cttacaaggc 3601 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc 3661 ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag accagatctg 3721 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc 3781 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct 3841 cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc atcttattat 3901 tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact tgtttattgc 3961 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt 4021 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggct 4081 ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 4141 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc 4201 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac gtacccaatt 4261 cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa cgtcgtgact 4321 gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct 4381 ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg 4441 gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca 4501 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct 4561 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt 4621 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac 4681 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct 4741 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt 4801 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac 4861 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg 4921 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc 4981 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag 5041 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt 5101 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt 5161 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga 5221 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat 5281 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga 5341 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag 5401 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg 5461 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg 5521 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt 5581 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg 5641 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc 5701 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg 5761 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac 5821 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact 5881 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa 5941 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa 6001 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg 6061 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc 6121 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac 6181 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca 6241 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt 6301 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc 6361 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg 6421 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc 6481 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac 6541 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct 6601 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc 6661 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt 6721 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac 6781 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg 6841 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga 6901 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac 6961 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt 7021 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agcgcgcaat 7081 taaccctcac taaagggaac aaaagctgga gctgca //