LOCUS V027813 6965 bp DNA circular SYN 01-JAN-1980 DEFINITION synthetic circular DNA. ACCESSION V027813 VERSION V027813 KEYWORDS . SOURCE . ORGANISM . . FEATURES Location/Qualifiers enhancer 235..614 /note="human cytomegalovirus immediate early enhancer" /label="CMV enhancer" promoter 615..818 /note="human cytomegalovirus (CMV) immediate early promoter" /label="CMV promoter" promoter 863..881 /note="promoter for bacteriophage T7 RNA polymerase" /label="T7 promoter" CDS 905..1663 /label="IL12A(NM_000882)" /note="IL12A(NM_000882)" /gene="IL12A" CDS 1670..1735 /codon_start=1 /product="three tandem FLAG® epitope tags, followed by an enterokinase cleavage site" /transl_table=1 /translation="DYKDHDGDYKDHDIDYKDDDDK" /label="3xFLAG" CDS 1745..1801 /codon_start=1 /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /product="2A peptide from porcine teschovirus-1 polyprotein" /transl_table=1 /translation="ATNFSLLKQAGDVEENPGP" /label="P2A" CDS 1814..2533 /codon_start=1 /note="mammalian codon-optimized" /product="the original enhanced GFP (Yang et al., 1996)" /transl_table=1 /translation="M,V,SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKL TLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFK DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNG IKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMV LLEFVTAAGITLGMDELYK*" /label="EGFP" polyA_signal 2565..2789 /note="bovine growth hormone polyadenylation signal" /label="bGH poly(A) signal" rep_origin 2835..3263 /direction=RIGHT /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" /label="f1 ori" promoter 3277..3606 /note="SV40 enhancer and early promoter" /label="SV40 promoter" CDS 3673..4467 /codon_start=1 /gene="aph(3')-II (or nptII)" /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin®)" /product="aminoglycoside phosphotransferase from Tn5" /transl_table=1 /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*" /label="NeoR/KanR" polyA_signal 4641..4762 /note="SV40 polyadenylation signal" /label="SV40 poly(A) signal" primer_bind complement(4811..4827) /note="common sequencing primer, one of multiple similar variants" /label="M13 rev" protein_bind 4835..4851 /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-β-D-thiogalactopyranoside (IPTG)." /label="lac operator" promoter complement(4859..4889) /note="promoter for the E. coli lac operon" /label="lac promoter" protein_bind 4904..4925 /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." /label="CAP binding site" rep_origin complement(5213..5798) /direction=LEFT /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" /label="ori" CDS complement(5969..6829) /codon_start=1 /gene="bla" /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /product="β-lactamase" /transl_table=1 /translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA SLIKHW*" /label="AmpR" promoter complement(6830..6934) /gene="bla" /label="AmpR 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 ctggctaact agagaaccca 841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc 901 caccatgtgg ccccctgggt cagcctccca gccaccgccc tcacctgccg cggccacagg 961 tctgcatcca gcggctcgcc ctgtgtccct gcagtgccgg ctcagcatgt gtccagcgcg 1021 cagcctcctc cttgtggcta ccctggtcct cctggaccac ctcagtttgg ccagaaacct 1081 ccccgtggcc actccagacc caggaatgtt cccatgcctt caccactccc aaaacctgct 1141 gagggccgtc agcaacatgc tccagaaggc cagacaaact ctagaatttt acccttgcac 1201 ttctgaagag attgatcatg aagatatcac aaaagataaa accagcacag tggaggcctg 1261 tttaccattg gaattaacca agaatgagag ttgcctaaat tccagagaga cctctttcat 1321 aactaatggg agttgcctgg cctccagaaa gacctctttt atgatggccc tgtgccttag 1381 tagtatttat gaagacttga agatgtacca ggtggagttc aagaccatga atgcaaagct 1441 tctgatggat cctaagaggc agatctttct agatcaaaac atgctggcag ttattgatga 1501 gctgatgcag gccctgaatt tcaacagtga gactgtgcca caaaaatcct cccttgaaga 1561 accggatttt tataaaacta aaatcaagct ctgcatactt cttcatgctt tcagaattcg 1621 ggcagtgact attgatagag tgatgagcta tctgaatgct tccctcgagg actacaaaga 1681 ccatgacggt gattataaag atcatgacat cgactacaag gatgacgatg acaagggaag 1741 cggagccact aacttctccc tgttgaaaca agcaggggat gtcgaagaga atcccgggcc 1801 accggtcgcc accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct 1861 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg 1921 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt 1981 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc 2041 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga 2101 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga 2161 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa 2221 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga 2281 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag 2341 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct 2401 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg 2461 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga 2521 gctgtacaag taagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg 2581 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc 2641 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc 2701 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag 2761 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc 2821 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac 2881 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc 2941 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt 3001 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg 3061 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac 3121 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta 3181 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat 3241 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag 3301 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc 3361 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat 3421 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt 3481 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc 3541 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt 3601 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga 3661 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg 3721 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg 3781 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc 3841 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct 3901 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa 3961 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg 4021 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa 4081 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat 4141 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg 4201 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc 4261 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac 4321 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg 4381 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc 4441 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag 4501 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg 4561 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc 4621 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca 4681 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt 4741 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg 4801 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca 4861 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca 4921 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc 4981 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt 5041 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact 5101 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag 5161 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata 5221 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc 5281 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg 5341 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc 5401 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg 5461 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc 5521 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga 5581 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg 5641 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa 5701 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ttttttgttt 5761 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta 5821 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat 5881 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa 5941 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct 6001 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta 6061 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct 6121 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg 6181 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa 6241 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt 6301 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta 6361 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca 6421 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta 6481 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct 6541 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg 6601 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac 6661 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact 6721 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa 6781 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt 6841 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat 6901 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg 6961 acgtc //