LOCUS Exported 5895 bp ds-DNA circular SYN 01-MAR-2022 DEFINITION Tobacco Etch Viral Protease. ACCESSION . VERSION . KEYWORDS CMVTO-TEVP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) TITLE Protease-controlled secretion and display of intercellular signals JOURNAL Unpublished REFERENCE 2 (bases 1 to 5895) AUTHORS . TITLE Direct Submission JOURNAL Exported Mar 1, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(189..208) /label=F1ori-R /note="F1 origin, reverse primer" protein_bind 340..387 /label=FRT /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="FLP-mediated recombination occurs in the 8-bp core sequence TCTAGAAA (Turan and Bode, 2011)." CDS 395..1417 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /label=HygR /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 1547..1668 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1584..1603) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1638..1657 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(1717..1733) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1717..1733) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1730..1752) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1741..1757 /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)." promoter complement(1765..1795) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1810..1831 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(1948..1965) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2119..2707) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2199..2218) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2878..3738) /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 3501..3520 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3739..3843) /gene="bla" /label=AmpR promoter primer_bind complement(3918..3937) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 4104..4483 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4484..4687 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4638..4658 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" protein_bind 4689..4707 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 4710..4728 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" primer_bind 4738..4762 /label=LNCX /note="Human CMV promoter, forward primer" regulatory 4854..4863 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 4869..5576 /codon_start=1 /product="tobacco etch virus protease (Kapust et al., 2001)" /label=TEV protease /translation="GESLFKGPRDYNPISSTICHLTNESDGHTTSLYGIGFGPFIITNK HLFRRNNGTLLVQSLHGVFKVKNTTTLQQHLIDGRDMIIIRMPKDFPPFPQKLKFREPQ REERICLVTTNFQTKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFI VGIHSASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPE EPFQPVKEATQLMN" CDS 5597..5626 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" primer_bind complement(5682..5699) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 5688..17 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" ORIGIN 1 tgcggtgggc tctatggaga tctatcgatg ttctggggat gcggtgggct ctatggcttc 61 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc 121 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct 181 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg 241 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga 301 ccccaaaaaa cttgattagg gtgatggttc acgtacctag aagttcctat tccgaagttc 361 ctattctcta gaaagtatag gaacttcctt ggccaaaaag cctgaactca ccgcgacgtc 421 tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc agctctcgga 481 gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt 541 aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact ttgcatcggc 601 cgcgctcccg attccggaag tgcttgacat tggggaattc agcgagagcc tgacctattg 661 catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc 721 tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc ttagccagac 781 gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat ggcgtgattt 841 catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg acgacaccgt 901 cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg actgccccga 961 agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg acaatggccg 1021 cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat acgaggtcgc 1081 caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc gctacttcga 1141 gcggaggcat ccggagcttg caggatcgcc gcggctccgg gcgtatatgc tccgcattgg 1201 tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag cttgggcgca 1261 gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta cacaaatcgc 1321 ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa 1381 ccgacgcccc agcactcgtc cgagggcaaa ggaatagcac gtactacgag atttcgattc 1441 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat 1501 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc 1561 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt 1621 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat 1681 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa 1741 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg 1801 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca 1861 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg 1921 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg 1981 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg 2041 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa 2101 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg 2161 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc 2221 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc 2281 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc 2341 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg 2401 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc 2461 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga 2521 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc 2581 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac 2641 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg 2701 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc 2761 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa 2821 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta 2881 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt 2941 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag 3001 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca 3061 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc 3121 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt 3181 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag 3241 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt 3301 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat 3361 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt 3421 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc 3481 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat 3541 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag 3601 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt 3661 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg 3721 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta 3781 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc 3841 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc 3901 ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc 3961 cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg 4021 caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt 4081 cgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat 4141 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa 4201 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt 4261 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta 4321 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt 4381 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc 4441 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca 4501 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat 4561 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa 4621 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag 4681 cagagctctc cctatcagtg atagagatct ccctatcagt gatagagatc gtcgacgagc 4741 tcgtttagtg aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag 4801 aagacaccgg gaccgatcca gcctccggac tctagcgttt aaacttaagc ttcgccacca 4861 tgggatccgg ggagagtttg tttaaggggc caagggacta taacccaata tcatccacta 4921 tttgccacct cactaacgag agcgatggac atacaacctc tctctacggg ataggcttcg 4981 gtcctttcat cattaccaat aagcatctgt ttcgccgaaa caacggtact cttctggttc 5041 aatctcttca tggtgtcttc aaggtgaaaa acaccactac gcttcagcaa cacctgattg 5101 atggtaggga tatgataatt atcagaatgc cgaaagattt cccacctttt ccacagaagc 5161 tgaaattcag ggaaccgcag agagaggaga ggatttgttt ggtaacgacc aacttccaga 5221 cgaagagtat gagttctatg gtgtccgaca ctagctgcac gttcccctca agtgatggga 5281 tattctggaa acactggata cagactaaag acggacagtg tggaagccca ttggtttcca 5341 cccgagatgg ttttatcgtg ggtatccata gcgcctctaa tttcacaaac acgaacaact 5401 atttcacttc agtgcccaaa aactttatgg agctgctcac aaaccaagaa gcgcagcagt 5461 gggtaagcgg ttggagactt aacgctgact cagttctctg gggggggcac aaagtattca 5521 tggtaaagcc agaggagcca ttccaaccag tcaaagaagc cacacaactt atgaacagct 5581 agctcgagga gggcccgaac aaaaactcat ctcagaagag gatctgaata gcgccgtcga 5641 ccatcatcat aatcatcatt gagtttaaac ccgctgatca gcctcgactg tgccttctag 5701 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac 5761 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca 5821 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag 5881 caggcatgct gggga //