LOCUS Exported 5903 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mAmber-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS . TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /product="mVenus YFP with monomerizing A206K mutation (Nagai et al., 2002)" /note="mAmber" /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGCGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 1081 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagctc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctcggctgc ggcctgcagt 1201 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg 1441 tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca 1501 acatcgagga cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccaag ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1681 ctctcggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc //