LOCUS pCBio 6834 bp ds-DNA circular 12-JUN-2008 DEFINITION Mammalian expression vector pCBio, complete sequence. ACCESSION DQ520291 VERSION DQ520291.1 GI:104744492 KEYWORDS . SOURCE Mammalian expression vector pCBio ORGANISM Mammalian expression vector pCBio other sequences; artificial sequences; vectors. REFERENCE 1 (bases 1 to 6834) AUTHORS Kulman,J.D., Satake,M. and Harris,J.E. TITLE A versatile system for site-specific enzymatic biotinylation and regulated expression of proteins in cultured mammalian cells JOURNAL Protein Expr. Purif. 52 (2), 320-328 (2007) PUBMED 17084093 REFERENCE 2 (bases 1 to 6834) AUTHORS Kulman,J.D., Satake,M. and Harris,J.E. TITLE Direct Submission JOURNAL Submitted (28-APR-2006) Biochemistry, University of Washington, N.E. Pacific St., Seattle, WA 98195, USA COMMENT COMMENT ApEinfo:methylated:1 FEATURES Location/Qualifiers promoter 7..439 /note="tetracycline-responsive promoter" /label=tetracycline-responsive promoter /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 protein_bind 15..33 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink protein_bind 57..75 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat(1) /ApEinfo_label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink protein_bind 99..117 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat(2) /ApEinfo_label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink misc_feature 5733..5782 /label=SV40 Promoter /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 misc_feature 5783..5836 /label=SV40 Promoter(1) /ApEinfo_label=SV40 Promoter /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 protein_bind 141..159 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat(3) /ApEinfo_label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink protein_bind 183..201 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat(4) /ApEinfo_label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink protein_bind 225..243 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat(5) /ApEinfo_label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink protein_bind 267..285 /note="tetO repeat" /bound_moiety="TetR" /label=tetO repeat(6) /ApEinfo_label=tetO repeat /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink misc_feature 320..439 /note="minimal CMV promoter" /label=minimal CMV promoter /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 TATA_signal 342..349 /label=TATA_signal /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink misc_feature 471..490 /note="multiple cloning site; MCS" /label=multiple cloning site; MCS /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 misc_feature 491..529 /note="biotin acceptor peptide (BioTag)-encoding" /label=biotin acceptor peptide (BioTag)-encoding /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 misc_feature 591..1177 /note="encephalomyocarditis virus internal ribosome entry site; IRES" /label=EMCV-R IRES /ApEinfo_fwdcolor=#00fff8 /ApEinfo_revcolor=#00fff8 gene 1209..2222 /gene="birA" /label=birA /ApEinfo_fwdcolor=#00ff6e /ApEinfo_revcolor=#00ff6e RBS 1209..1217 /gene="birA" /note="Kozak site" /label=birA(1) /ApEinfo_label=birA /ApEinfo_fwdcolor=pink /ApEinfo_revcolor=pink CDS 1218..2222 /gene="birA" /note="holocarboxylase synthetase" /codon_start=1 /product="biotin ligase" /protein_id="ABF74577.1" /db_xref="GI:104744493" /translation="MDYKDDDDKSPRSMKDNTVPLKLIALLANGEFHSGEQLGETLGM SRAAINKHIQTLRDWGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDS TNQYLLDRIGELKSGDACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAI GLSLVIGIVMAEVLRKLGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGA GINMAMRRVEESVVNQGWITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLS RWEKLDNFINRPVKLIIGDKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK " /label=biotin ligase /ApEinfo_fwdcolor=#ffff9e /ApEinfo_revcolor=#ffff9e misc_feature 1221..1244 /gene="birA" /note="FLAG epitope tag-encoding" /label=birA(2) /ApEinfo_label=birA /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 misc_feature 1257..2222 /gene="birA" /note="Escherichia coli BirA (biotin ligase)" /label=birA(3) /ApEinfo_label=birA /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 gene complement(4432..5292) /gene="amp" /label=amp /ApEinfo_fwdcolor=#00ff6e /ApEinfo_revcolor=#00ff6e CDS complement(4432..5292) /gene="amp" /note="confers ampicillin resistance" /codon_start=1 /product="beta-lactamase" /protein_id="ABF74578.1" /db_xref="GI:104744494" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW" /label=beta-lactamase /ApEinfo_fwdcolor=#ffff9e /ApEinfo_revcolor=#ffff9e gene 5924..6523 /gene="pur" /label=pur /ApEinfo_fwdcolor=#00ff6e /ApEinfo_revcolor=#00ff6e CDS 5924..6523 /gene="pur" /note="confers puromycin resistance" /codon_start=1 /product="puromycin N-acetyltransferase" /protein_id="ABF74579.1" /db_xref="GI:104744495" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSR LAAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFL ETSAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" /label=puromycin N-acetyltransferase /ApEinfo_fwdcolor=#ffff9e /ApEinfo_revcolor=#ffff9e polyA_signal 2272..3439 /note="beta-globin poly-A signal" /label=beta-globin poly-A signal /ApEinfo_fwdcolor=#ff00cb /ApEinfo_revcolor=#ff00cb misc_feature 371..371 /label=putative CMV transcription start site (+1) /ApEinfo_fwdcolor=#ff00ff /ApEinfo_revcolor=#ff00ff misc_feature 3080..3080 /label=poly(A) tail appended after this base /ApEinfo_fwdcolor=cyan /ApEinfo_revcolor=cyan misc_feature 6662..6834 /label=inverted SV40 late poly(A) signal /ApEinfo_fwdcolor=cyan /ApEinfo_revcolor=cyan misc_feature 5502..5732 /label=SV40 enhancer /ApEinfo_fwdcolor=#ff0000 /ApEinfo_revcolor=#ff0000 misc_feature 5733..5782 /label=SV40 transcription start sites region /ApEinfo_fwdcolor=#ffb1be /ApEinfo_revcolor=#ffb1be misc_feature 5733..5733 /label=SV40 transcription start (+1) /ApEinfo_fwdcolor=#ff00ff /ApEinfo_revcolor=#ff00ff misc_feature 5782..5782 /label=SV40 transcription start (+1)(1) /ApEinfo_label=SV40 transcription start (+1) /ApEinfo_fwdcolor=#ff00ff /ApEinfo_revcolor=#ff00ff misc_feature 5777..5777 /label=SV40 transcription start (+1)(2) /ApEinfo_label=SV40 transcription start (+1) /ApEinfo_fwdcolor=#ff00ff /ApEinfo_revcolor=#ff00ff misc_feature 5771..5771 /label=SV40 transcription start (+1)(3) /ApEinfo_label=SV40 transcription start (+1) /ApEinfo_fwdcolor=#fe00ff /ApEinfo_revcolor=#fe00ff ORIGIN 1 ctcgagttta ccactcccta tcagtgatag agaaaagtga aagtcgagtt taccactccc 61 tatcagtgat agagaaaagt gaaagtcgag tttaccactc cctatcagtg atagagaaaa 121 gtgaaagtcg agtttaccac tccctatcag tgatagagaa aagtgaaagt cgagtttacc 181 actccctatc agtgatagag aaaagtgaaa gtcgagttta ccactcccta tcagtgatag 241 agaaaagtga aagtcgagtt taccactccc tatcagtgat agagaaaagt gaaagtcgag 301 ctcggtaccc gggtcgaggt aggcgtgtac ggtgggaggc ctatataagc agagctcgtt 361 tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac 421 accgggaccg atccagcctc cgcggccccg aattcgagct cggtacccgg ggatccgata 481 tcgcggccgc ttgaacgaca tttttgaagc gcaaaaaatt gaatggcatt aaggccgcgc 541 tagcctcgag aattcacgcg tcgagcatgc atctagggcg gccaattccg cccctctccc 601 tccccccccc ctaacgttac tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc 661 tatatgtgat tttccaccat attgccgtct tttggcaatg tgagggcccg gaaacctggc 721 cctgtcttct tgacgagcat tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt 781 ctgttgaatg tcgtgaagga agcagttcct ctggaagctt cttgaagaca aacaacgtct 841 gtagcgaccc tttgcaggca gcggaacccc ccacctggcg acaggtgcct ctgcggccaa 901 aagccacgtg tataagatac acctgcaaag gcggcacaac cccagtgcca cgttgtgagt 961 tggatagttg tggaaagagt caaatggctc tcctcaagcg tattcaacaa ggggctgaag 1021 gatgcccaga aggtacccca ttgtatggga tctgatctgg ggcctcggtg cacatgcttt 1081 acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt 1141 ttcctttgaa aaacacgatg ataagcttgc cacaacccaa acttaagctt ggtaccgagc 1201 tcggatctgc cgccaccatg gattacaagg atgacgacga taagagcccg agatccatga 1261 aggataacac cgtgccactg aaattgattg ccctgttagc gaacggtgaa tttcactctg 1321 gcgagcagtt gggtgaaacg ctgggaatga gccgggcggc tattaataaa cacattcaga 1381 cactgcgtga ctggggcgtt gatgtcttta ccgttccggg taaaggatac agcctgcctg 1441 agcctatcca gttacttaat gctaaacaga tattgggtca gctggatggc ggtagtgtag 1501 ccgtgctgcc agtgattgac tccacgaatc agtaccttct tgatcgtatc ggagagctta 1561 aatcgggcga tgcttgcatt gcagaatacc agcaggctgg ccgtggtcgc cggggtcgga 1621 aatggttttc gccttttggc gcaaacttat atttgtcgat gttctggcgt ctggaacaag 1681 gcccggcggc ggcgattggt ttaagtctgg ttatcggtat cgtgatggcg gaagtattac 1741 gcaagctggg tgcagataaa gttcgtgtta aatggcctaa tgacctctat ctgcaggatc 1801 gcaagctggc aggcattctg gtggagctga ctggcaaaac tggcgatgcg gcgcaaatag 1861 tcattggagc cgggatcaac atggcaatgc gccgtgttga agagagtgtc gttaatcagg 1921 ggtggatcac gctgcaggaa gcggggatca atctcgatcg taatacgttg gcggccatgc 1981 taatacgtga attacgtgct gcgttggaac tcttcgaaca agaaggattg gcaccttatc 2041 tgtcgcgctg ggaaaagctg gataatttta ttaatcgccc agtgaaactt atcattggtg 2101 ataaagaaat atttggcatt tcacgcggaa tagacaaaca gggggcttta ttacttgagc 2161 aggatggaat aataaaaccc tggatgggcg gtgaaatatc cctgcgtagt gcagaaaaat 2221 aagaattcag atctctgcag ctcgagtcta gagggcccgt ttatctctag agctgagaac 2281 ttcagggtga gtttggggac ccttgattgt tctttctttt tcgctattgt aaaattcatg 2341 ttatatggag ggggcaaagt tttcagggtg ttgtttagaa tgggaagatg tcccttgtat 2401 caccatggac cctcatgata attttgtttc tttcactttc tactctgttg acaaccattg 2461 tctcctctta ttttcttttc attttctgta acttttttcg ttaaacttta gcttgcattt 2521 gtaacgaatt tttaaattca ctttcgttta tttgtcagat tgtaagtact ttctctaatc 2581 actttttttt caaggcaatc agggtaatta tattgtactt cagcacagtt ttagagaaca 2641 attgttataa ttaaatgata aggtagaata tttctgcata taaattctgg ctggcgtgga 2701 aatattctta ttggtagaaa caactacatc ctggtaatca tcctgccttt ctctttatgg 2761 ttacaatgat atacactgtt tgagatgagg ataaaatact ctgagtccaa accgggcccc 2821 tctgctaacc atgttcatgc cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt 2881 gttgtgctgt ctcatcattt tggcaaagaa ttcactcctc aggtgcaggc tgcctatcag 2941 aaggtggtgg ctggtgtggc caatgccctg gctcacaaat accactgaga tctttttccc 3001 tctgccaaaa attatgggga catcatgaag ccccttgagc atctgacttc tgggtaataa 3061 aggaaattta ttttcattgc aatagtgtgt gggaattttt tgtgtctctc actcggaagg 3121 acatatggga gggcaaatca tttaaaacat cagaatgagt atttggttta gagtttggca 3181 acatatgcca tatgctggct gccatgaaca aaggtggcta taaagaggtc atcagtatat 3241 gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat 3301 tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca 3361 tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct 3421 tctcttatga actcgactgc attaatgaat cggccaacgc gcggggagag gcggtttgcg 3481 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg 3541 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa 3601 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc 3661 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc 3721 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag 3781 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct 3841 cccttcggga agcgtggcgc tttctcaatg ctcacgctgt aggtatctca gttcggtgta 3901 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc 3961 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc 4021 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt 4081 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct 4141 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc 4201 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca 4261 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta 4321 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa 4381 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg 4441 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg 4501 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc 4561 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc 4621 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa 4681 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc 4741 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg 4801 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc 4861 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat 4921 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg 4981 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc 5041 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg 5101 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat 5161 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg 5221 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg 5281 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct 5341 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac 5401 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta 5461 taaaaatagg cgtatcacga ggccctttcg tcttcactcg agtgtgtcag ttagggtgtg 5521 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag 5581 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc 5641 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc 5701 ccagttccgc ccattctccg ccccatggct gaCTAATTTT TTTTATTTAT GCAGAGGCCG 5761 AGGCCGCCTC GGCCTCTGAG CTATTCCAGA AGTAGTGAGG AGGCTTTTTT GGAGGCCTAG 5821 GCTTTTGCAA AAAGCTtgca tgcctgcagg tcggccgcca cgaccggtgc cgccaccatc 5881 ccctgaccca cgcccctgac ccctcacaag gagacgacct tccatgaccg agtacaagcc 5941 cacggtgcgc ctcgccaccc gcgacgacgt cccccgggcc gtacgcaccc tcgccgccgc 6001 gttcgccgac taccccgcca cgcgccacac cgtcgacccg gaccgccaca tcgagcgggt 6061 caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt 6121 cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc 6181 ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca 6241 gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc 6301 caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc 6361 cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg 6421 caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga 6481 aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgacgcccgc cccacgaccc 6541 gcagcgcccg accgaaagga gcgcacgacc ccatggctcc gaccgaagcc gacccgggcg 6601 gccccgccga ccccgcaccc gcccccgagg cccaccgact ctagaggatc ataatcagcc 6661 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc 6721 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt 6781 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgc //