LOCUS Exported 7516 bp ds-DNA circular SYN 18-JAN-2022 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS LV-Cre pLKO.1 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7516) AUTHORS Beronja S, Livshits G, Williams S, Fuchs E TITLE Rapid functional dissection of genetic networks via tissue-specific transduction and RNAi in mouse embryos. JOURNAL Nat Med. 2010 Jul . 16(7):821-7. PUBMED 20526348 REFERENCE 2 (bases 1 to 7516) AUTHORS . TITLE Direct Submission JOURNAL Exported Jan 18, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7516 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 41..344 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 345..548 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 499..519 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 545..569 /label=LNCX /note="Human CMV promoter, forward primer" CDS 643..663 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 664..1692 /codon_start=1 /product="site-specific recombinase" /label=Cre /note="Cre is a site-specific recombinase from bacteriophage P1. Recombination occurs at loxP sequences." /translation="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLQ SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED GD" LTR 1771..2004 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 2076..2197 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2113..2132) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2167..2186 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin 2237..2372 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2299..2318 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(2392..2411) /label=T7 /note="T7 promoter, forward primer" promoter complement(2393..2411) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2421..2438) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2421..2437) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2430..2452) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 2579..3034 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2666..2685) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2876..2897 /label=F1ori-F /note="F1 origin, forward primer" promoter 3060..3164 /gene="bla" /label=AmpR promoter CDS 3165..4025 /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(3383..3402) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4196..4784 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4685..4704 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 4938..4955 /label=L4440 /note="L4440 vector, forward primer" protein_bind 5072..5093 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5108..5138 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5146..5162 /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 5151..5173 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 5170..5186 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5170..5186 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 5205..5225 /label=T3 /note="T3 promoter, forward primer" promoter 5207..5225 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" promoter 5253..5479 /label=RSV promoter /note="Rous sarcoma virus enhancer/promoter" LTR 5480..5660 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 5707..5832 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 6325..6558 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 6743..6787 /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" promoter 7085..7325 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 7085..7105 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 7256..7275 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_feature 7396..7513 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" ORIGIN 1 gtttattaca gggacagcag gctagcgctt gggagttccg cgttacataa cttacggtaa 61 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg 121 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt 181 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg 241 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc 301 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc 361 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca 421 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta 481 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa 541 gcagagctcg tttagtgaac cgtcagatcg cctggagacg ccatccacgc tgttttgacc 601 tccatagaag aacgcgtcac cgactctaga ggatctacca tgcccaagaa gaagaggaag 661 gtgtccaatt tactgaccgt acaccaaaat ttgcctgcat taccggtcga tgcaacgagt 721 gatgaggttc gcaagaacct gatggacatg ttcagggatc gccaggcgtt ttctgagcat 781 acctggaaaa tgcttcagtc cgtttgccgg tcgtgggcgg catggtgcaa gttgaataac 841 cggaaatggt ttcccgcaga acctgaagat gttcgcgatt atcttctata tcttcaggcg 901 cgcggtctgg cagtaaaaac tatccagcaa catttgggcc agctaaacat gcttcatcgt 961 cggtccgggc tgccacgacc aagtgacagc aatgctgttt cactggttat gcggcggatc 1021 cgaaaagaaa acgttgatgc cggtgaacgt gcaaaacagg ctctagcgtt cgaacgcact 1081 gatttcgacc aggttcgttc actcatggaa aatagcgatc gctgccagga tatacgtaat 1141 ctggcatttc tggggattgc ttataacacc ctgttacgta tagccgaaat tgccaggatc 1201 agggttaaag atatctcacg tactgacggt gggagaatgt taatccatat tggcagaacg 1261 aaaacgctgg ttagcaccgc aggtgtagag aaggcactta gcctgggggt aactaaactg 1321 gtcgagcgat ggatttccgt ctctggtgta gctgatgatc cgaataacta cctgttttgc 1381 cgggtcagaa aaaatggtgt tgccgcgcca tctgccacca gccagctatc aactcgcgcc 1441 ctggaaggga tttttgaagc aactcatcga ttgatttacg gcgctaagga tgactctggt 1501 cagagatacc tggcctggtc tggacacagt gcccgtgtcg gagccgcgcg agatatggcc 1561 cgcgctggag tttcaatacc ggagatcatg caagctggtg gctggaccaa tgtaaatatt 1621 gtcatgaact atatccgtaa cctggatagt gaaacagggg caatggtgcg cctgctggaa 1681 gatggcgatt agatgcatcg gtacctttaa gaccaatgac ttacaaggca gctgtagatc 1741 ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc caacgaagac 1801 aagatctgct ttttgcttgt actgggtctc tctggttaga ccagatctga gcctgggagc 1861 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc 1921 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt 1981 agtcagtgtg gaaaatctct agcagtagta gttcatgtca tcttattatt cagtatttat 2041 aacttgcaaa gaaatgaata tcagagagtg agaggaactt gtttattgca gcttataatg 2101 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 2161 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggctc tagctatccc 2221 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca 2281 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt 2341 ccagaagtag tgaggaggct tttttggagg cctagggacg tacccaattc gccctatagt 2401 gagtcgtatt acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct 2461 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc 2521 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac 2581 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct 2641 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg 2701 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt 2761 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca 2821 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga 2881 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa 2941 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac 3001 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc 3061 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac 3121 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt 3181 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag 3241 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg 3301 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa 3361 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc 3421 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag 3481 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa 3541 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc 3601 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg 3661 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa 3721 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa 3781 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg 3841 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag 3901 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg 3961 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt 4021 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt 4081 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac 4141 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag 4201 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg 4261 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca 4321 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga 4381 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca 4441 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc 4501 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca 4561 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa 4621 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc 4681 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc 4741 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg 4801 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat 4861 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca 4921 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca 4981 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg 5041 actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac 5101 cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac 5161 aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt aaccctcact 5221 aaagggaaca aaagctggag ctgcaagctt aatgtagtct tatgcaatac tcttgtagtc 5281 ttgcaacatg gtaacgatga gttagcaaca tgccttacaa ggagagaaaa agcaccgtgc 5341 atgccgattg gtggaagtaa ggtggtacga tcgtgcctta ttaggaaggc aacagacggg 5401 tctgacatgg attggacgaa ccactgaatt gccgcattgc agagatattg tatttaagtg 5461 cctagctcga tacataaacg ggtctctctg gttagaccag atctgagcct gggagctctc 5521 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt 5581 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc 5641 agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa agggaaacca 5701 gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag aggcgagggg 5761 cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga gagagatggg 5821 tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa aattcggtta 5881 aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag cagggagcta 5941 gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag acaaatactg 6001 ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca 6061 gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa ggaagcttta 6121 gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc ggccgctgat 6181 cttcagacct ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa 6241 agtagtaaaa attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca 6301 gagagaaaaa agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg 6361 aagcactatg ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg 6421 tatagtgcag cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca 6481 actcacagtc tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct 6541 aaaggatcaa cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc 6601 tgtgccttgg aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac 6661 ctggatggag tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga 6721 agaatcgcaa aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc 6781 aagtttgtgg aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat 6841 gatagtagga ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag 6901 agttaggcag ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc 6961 cgacaggccc gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg 7021 attagtgaac ggatctcgac ggtatcgatc acgagactag cctcgagcgg ccgccccctt 7081 caccgagggc ctatttccca tgattccttc atatttgcat atacgataca aggctgttag 7141 agagataatt ggaattaatt tgactgtaaa cacaaagata ttagtacaaa atacgtgacg 7201 tagaaagtaa taatttcttg ggtagtttgc agttttaaaa ttatgtttta aaatggacta 7261 tcatatgctt accgtaactt gaaagtattt cgatttcttg gctttatata tcttgtggaa 7321 aggacgaaac accggtccgc aggtatgcac gcgtgaattc tcgacctcga gacaaatggc 7381 agtattcatc cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag 7441 aatagtagac ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa 7501 aattcaaaat tttcgg //