LOCUS Exported 7032 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pCT302 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7032) AUTHORS Midelfort KS, Hernandez HH, Lippow SM, Tidor B, Drennan CL, Wittrup KD TITLE Substantial energetic improvement with minimal structural perturbation in a high affinity mutant antibody. JOURNAL J Mol Biol. 2004 Oct 22;343(3):685-701. PUBMED 15465055 REFERENCE 2 (bases 1 to 7032) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7032 /organism="synthetic DNA construct" /mol_type="other DNA" CDS complement(159..833) /codon_start=1 /gene="S. cerevisiae TRP1" /product="phosphoribosylanthranilate isomerase, required for tryptophan biosynthesis" /label=TRP1 /note="yeast auxotrophic marker" /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA KK" promoter complement(834..1114) /gene="S. cerevisiae TRP1" /label=TRP1 promoter primer_bind complement(1131..1150) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 1250..1272 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(1310..1328) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" misc_feature 1370..1873 /label=CEN/ARS /note="S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence" promoter 1910..2014 /gene="bla" /label=AmpR promoter CDS 2015..2875 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(2233..2252) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 3046..3634 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3788..3805 /label=L4440 /note="L4440 vector, forward primer" promoter 3958..3988 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3996..4012 /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 4001..4023 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 4020..4036 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 4020..4036 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" promoter 4057..4075 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" promoter 4240..4904 /gene="GAL1 and GAL10 genes of S. cerevisiae" /label=GAL1,10 promoter /note="divergent inducible promoter, regulated by Gal4" protein_bind 4456..4573 /label=UAS /bound_moiety="Gal4" /note="upstream activating sequence mediating Gal4-dependent induction" primer_bind 4868..4891 /label=GAL1 /note="S. cerevisiae GAL1 promoter, forward primer" CDS 5223..5234 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" CDS 5235..5261 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 6090..6119 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" primer_bind complement(6452..6471) /label=T7 /note="T7 promoter, forward primer" promoter complement(6453..6471) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(6478..6495) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(6478..6494) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(6487..6509) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 6639..62 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(6726..6745) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 6936..6957 /label=F1ori-F /note="F1 origin, forward primer" ORIGIN 1 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat 61 ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg caggcaagtg 121 cacaaacaat acttaaataa atactactca gtaataacct atttcttagc atttttgacg 181 aaatttgcta ttttgttaga gtcttttaca ccatttgtct ccacacctcc gcttacatca 241 acaccaataa cgccatttaa tctaagcgca tcaccaacat tttctggcgt cagtccacca 301 gctaacataa aatgtaagct ttcggggctc tcttgccttc caacccagtc agaaatcgag 361 ttccaatcca aaagttcacc tgtcccacct gcttctgaat caaacaaggg aataaacgaa 421 tgaggtttct gtgaagctgc actgagtagt atgttgcagt cttttggaaa tacgagtctt 481 ttaataactg gcaaaccgag gaactcttgg tattcttgcc acgactcatc tccatgcagt 541 tggacgatat caatgccgta atcattgacc agagccaaaa catcctcctt aggttgatta 601 cgaaacacgc caaccaagta tttcggagtg cctgaactat ttttatatgc ttttacaaga 661 cttgaaattt tccttgcaat aaccgggtca attgttctct ttctattggg cacacatata 721 atacccagca agtcagcatc ggaatctaga gcacattctg cggcctctgt gctctgcaag 781 ccgcaaactt tcaccaatgg accagaacta cctgtgaaat taataacaga catactccaa 841 gctgcctttg tgtgcttaat cacgtatact cacgtgctca atagtcacca atgccctccc 901 tcttggccct ctccttttct tttttcgacc gaattaattc ttaatcggca aaaaaagaaa 961 agctccggat caagattgta cgtaaggtga caagctattt ttcaataaag aatatcttcc 1021 actactgcca tctggcgtca taactgcaaa gtacacatat attacgatgc tgtctattaa 1081 atgcttccta tattatatat atagtaatgt cgtttatggt gcactctcag tacaatctgc 1141 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga 1201 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc 1261 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata 1321 cgcctatttt tataggttaa tgtcatgata ataatggttt cttaggacgg atcgcttgcc 1381 tgtaacttac acgcgcctcg tatcttttaa tgatggaata atttgggaat ttactctgtg 1441 tttatttatt tttatgtttt gtatttggat tttagaaagt aaataaagaa ggtagaagag 1501 ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa atttcaacaa aaagcgtact 1561 ttacatatat atttattaga caagaaaagc agattaaata gatatacatt cgattaacga 1621 taagtaaaat gtaaaatcac aggattttcg tgtgtggtct tctacacaga caagatgaaa 1681 caattcggca ttaatacctg agagcaggaa gagcaagata aaaggtagta tttgttggcg 1741 atccccctag agtcttttac atcttcggaa aacaaaaact attttttctt taatttcttt 1801 ttttactttc tatttttaat ttatatattt atattaaaaa atttaaatta taattatttt 1861 tatagcacgt gatgaaaagg acccaggtgg cacttttcgg ggaaatgtgc gcggaacccc 1921 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg 1981 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 2041 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 2101 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct 2161 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 2221 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 2281 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 2341 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 2401 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 2461 ttttcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 2521 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 2581 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 2641 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 2701 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 2761 agatggtaag ccctcccgta tcgtagttat ctacacgacg ggcagtcagg caactatgga 2821 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 2881 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 2941 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc 3001 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt 3061 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt 3121 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 3181 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 3241 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 3301 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 3361 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 3421 atacctacag cgtgagcatt gagaaagcgc cacgcttccc gaagggagaa aggcggacag 3481 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc caggggggaa 3541 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 3601 gtgatgctcg tcaggggggc cgagcctatg gaaaaacgcc agcaacgcgg cctttttacg 3661 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 3721 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 3781 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 3841 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 3901 gggcagtgag cgcaacgcaa ttaatgtgag ttacctcact cattaggcac cccaggcttt 3961 acactttatg cttccggctc ctatgttgtg tggaattgtg agcggataac aatttcacac 4021 aggaaacagc tatgaccatg attacgccaa gctcggaatt aaccctcact aaagggaaca 4081 aaagctgggt acccgacagg ttatcagcaa caacacagtc atatccattc tcaattagct 4141 ctaccacagt gtgtgaacca atgtatccag caccacctgt aaccaaaaca attttagaag 4201 tactttcact ttgtaactga gctgtcattt atattgaatt ttcaaaaatt cttacttttt 4261 ttttggatgg acgcaaagaa gtttaataat catattacat ggcattacca ccatatacat 4321 atccatatac atatccatat ctaatcttac ttatatgttg tggaaatgta aagagcccca 4381 ttatcttagc ctaaaaaaac cttctctttg gaactttcag taatacgctt aactgctcat 4441 tgctatattg aagtacggat tagaagccgc cgagcgggtg acagccctcc gaaggaagac 4501 tctcctccgt gcgtcctcgt cttcaccggt cgcgttcctg aaacgcagat gtgcctcgcg 4561 ccgcactgct ccgaacaata aagattctac aatactagct tttatggtta tgaagaggaa 4621 aaattggcag taacctggcc ccacaaacct tcaaatgaac gaatcaaatt aacaaccata 4681 ggatgataat gcgattagtt ttttagcctt atttctgggg taattaatca gcgaagcgat 4741 gatttttgat ctattaacag atatataaat gcaaaaactg cataaccact ttaactaata 4801 ctttcaacat tttcggtttg tattacttct tattcaaatg taataaaagt atcaacaaaa 4861 aattgttaat atacctctat actttaacgt caaggagaaa aaaccccgga tcgaattccc 4921 tacttcatac attttcaatt aagatgcagt tacttcgctg tttttcaata ttttctgtta 4981 ttgcttcagt tttagcacag gaactgacaa ctatatgcga gcaaatcccc tcaccaactt 5041 tagaatcgac gccgtactct ttgtcaacga ctactatttt ggccaacggg aaggcaatgc 5101 aaggagtttt tgaatattac aaatcagtaa cgtttgtcag taattgcggt tctcacccct 5161 caacaactag caaaggcagc cccataaaca cacagtatgt ttttaaggac aatagctcga 5221 cgattgaagg tagataccca tacgacgttc cagactacgc tctgcaggct agtggtggtg 5281 gtggttctgg tggtggtggt tctggtggtg gtggttctgc tagcgacgtc gttatgactc 5341 aaacaccact atcacttcct gttagtctag gtgatcaagc ctccatctct tgcagatcta 5401 gtcagagcct tgtacacagt aatggaaaca cctatttacg ttggtacctg cagaagccag 5461 gccagtctcc aaaggtcctg atctacaaag tttccaaccg attttctggg gtcccagaca 5521 ggttcagtgg cagtggatca gggacagatt tcacactcaa gatcagcaga gtggaggctg 5581 aggatctggg agtttatttc tgctctcaaa gtacacatgt tccgtggacg ttcggtggag 5641 gcaccaagct tgaaattaag tcctctgctg atgatgctaa gaaggatgct gctaagaagg 5701 atgatgctaa gaaagatgat gctaagaaag atggtgacgt caaactggat gagactggag 5761 gaggcttggt gcaacctggg aggcccatga aactctcctg tgttgcctct ggattcactt 5821 ttagtgacta ctggatgaac tgggtccgcc agtctccaga gaaaggactg gagtgggtag 5881 cacaaattag aaacaaacct tataattatg aaacatatta ttcagattct gtgaaaggca 5941 gattcaccat ctcaagagat gattccaaaa gtagtgtcta cctgcaaatg aacaacttaa 6001 gagttgaaga catgggtatc tattactgta cgggttctta ctatggtatg gactactggg 6061 gtcaaggaac ctcagtcacc gtctcctcag aacaaaagct tatttctgaa gaagacttgt 6121 aatagctcga gatctgataa caacagtgta gatgtaacaa aatcgacttt gttcccactg 6181 tacttttagc tcgtacaaaa tacaatatac ttttcatttc tccgtaaaca acatgttttc 6241 ccatgtaata tccttttcta tttttcgttc cgttaccaac tttacacata ctttatatag 6301 ctattcactt ctatacacta aaaaactaag acaattttaa ttttgctgcc tgccatattt 6361 caatttgtta taaattccta taatttatcc tattagtagc taaaaaaaga tgaatgtgaa 6421 tcgaatccta agagaattga gctccaattc gccctatagt gagtcgtatt acaattcact 6481 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct 6541 tgcagcacat ccccccttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc 6601 ttcccaacag ttgcgcagcc tgaatggcga atggcgcgac gcgccctgta gcggcgcatt 6661 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc 6721 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca 6781 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc 6841 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt 6901 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac 6961 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc 7021 ctattggtta aa //