LOCUS Exported 9032 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS Flag-HA-USP13 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 9032) AUTHORS Sowa ME, Bennett EJ, Gygi SP, Harper JW TITLE Defining the human deubiquitinating enzyme interaction landscape. JOURNAL Cell. 2009 Jul 23. 138(2):389-403. PUBMED 19615732 REFERENCE 2 (bases 1 to 9032) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..9032 /organism="synthetic DNA construct" /mol_type="other DNA" LTR 65..579 /label=3' LTR /note="3' long terminal repeat from murine embryonic stem cell virus" protein_bind 741..757 /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(765..795) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 810..831 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(948..965) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(1119..1707) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1199..1218) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1878..2738) /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 2501..2520 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2739..2843) /gene="bla" /label=AmpR promoter primer_bind 2911..2929 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(2967..2989) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 3089..3108 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3302..3324 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 3452..3471 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" LTR 3671..4187 /label=5' LTR /note="5' long terminal repeat from murine embryonic stem cell virus" misc_feature 4251..4592 /label=MESV Psi /note="packaging signal of murine embryonic stem cell virus" CDS 4659..5075 /label=gag (truncated) /note="truncated Moloney murine leukemia virus (MMLV) gag gene lacking the start codon" primer_bind 5005..5027 /label=pLXSN 5' /note="Murine stem cell virus, forward primer. Also called MSCV" primer_bind 5043..5059 /label=pBABE 5' /note="Psi packaging signal, forward primer for pBABE vectors" regulatory 5088..5097 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 5097..5120 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 5133..5159 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" protein_bind 5175..5199 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" protein_bind complement(7804..7828) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" misc_feature 7858..8410 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(8002..8019) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 8229..8248 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 8422..9021 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(8422..8441) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 8918..8938 /label=Puro-F /note="Puromycin resistance gene, forward primer" ORIGIN 1 cgacctgcag ccaagcttat cgataaaata aaagatttta tttagtctcc agaaaaaggg 61 gggaatgaaa gaccccacct gtaggtttgg caagctagct taagtaacgc cattttgcaa 121 ggcatggaaa atacataact gagaatagag aagttcagat caaggttagg aacagagaga 181 cagcagaata tgggccaaac aggatatctg tggtaagcag ttcctgcccc ggctcagggc 241 caagaacaga tggtccccag atgcggtccc gccctcagca gtttctagag aaccatcaga 301 tgtttccagg gtgccccaag gacctgaaat gaccctgtgc cttatttgaa ctaaccaatc 361 agttcgcttc tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa aagagcccac 421 aacccctcac tcggcgcgcc agtcctccga tagactgcgt cgcccgggta cccgtgtatc 481 caataaaccc tcttgcagtt gcatccgact tgtggtctcg ctgttccttg ggagggtctc 541 ctctgagtga ttgactaccc gtcagcgggg gtctttcatg ggtaacagtt tcttgaagtt 601 ggagaacaac attctgaggg taggagtcga atattaagta atcctgactc aattagccac 661 tgttttgaat ccacatactc caatactcct gaaatagttc attatggaca gcgcagaaag 721 agctggggag aattgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga 781 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg 841 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc 901 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac 961 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata 1021 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa 1081 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct 1141 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa 1201 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg 1261 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca 1321 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa 1381 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg 1441 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg 1501 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga 1561 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc 1621 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag 1681 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac 1741 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc 1801 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag 1861 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt 1921 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag 1981 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca 2041 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact 2101 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca 2161 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg 2221 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc 2281 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg 2341 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca 2401 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt 2461 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc 2521 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc 2581 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca 2641 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa 2701 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat 2761 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa 2821 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa 2881 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc 2941 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca 3001 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt 3061 ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac 3121 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgccat 3181 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta 3241 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt 3301 tcccagtcac gacgttgtaa aacgacggcg caaggaatgg tgcatgcaag gagatggcgc 3361 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga 3421 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa 3481 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg cgattagtcc 3541 aatttgttaa agacaggata tcagtggtcc aggctctagt tttgactcaa caatatcacc 3601 agctgaagcc tatagagtac gagccataga taaaataaaa gattttattt agtctccaga 3661 aaaagggggg aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat 3721 tttgcaaggc atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac 3781 agagagacag cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc 3841 tcagggccaa gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac 3901 catcagatgt ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta 3961 accaatcagt tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag 4021 agcccacaac ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc 4081 gtattcccaa taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg 4141 agggtctcct cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc 4201 gagatttgga gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca 4261 gcggtcgttt cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc 4321 gcctgcgtct gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg 4381 acgagttctg aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt 4441 tttgtggccc gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg 4501 ttctggtagg agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg 4561 tttggaaccg aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc 4621 tctgtctgac tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct 4681 taagtttgac cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag 4741 atgtcaagaa gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg 4801 gatggccgcg agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct 4861 tttcacctgg cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct 4921 tggcttttga cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc 4981 ttcctccatc cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct 5041 ccctttatcc agccctcact ccttctctag gcgccggaat tagatctgcc accatggact 5101 acaaggatga cgatgacaag ctcgatggag gatacccata cgatgttcca gattacgctg 5161 ctcgaggtta tcaaacaagt ttgtacaaaa aagttggcgc gcccaccatg cagcgccggg 5221 gcgccctgtt cggcatgccg ggcggcagcg gaggcaggaa gatggctgca ggagacatcg 5281 gcgagctgct agtgccccac atgcccacga tccgcgtgcc caggtccggc gacagggtct 5341 acaagaacga gtgcgccttc tcctacgact ctcccaattc tgaaggtgga ctctatgtat 5401 gcatgaatac atttttggcc tttggaaggg aacatgttga aagacatttt cgaaaaactg 5461 gacagagtgt atacatgcac ctgaaaagac atgtgcgaga gaaggtaaga ggggcgtctg 5521 gtggagcgtt accaaaaagg aggaattcca agattttttt agatctagat actgatgacg 5581 atttaaatag cgacgattat gaatatgaag atgaagccaa acttgttata ttcccagatc 5641 actatgaaat agcactacca aatattgagg agttaccagc cctggtaaca attgcttgtg 5701 atgcagttct cagctcaaaa tctccataca gaaagcagga cccagacacg tgggaaaatg 5761 aattgccagt atctaaatat gccaacaacc tcacccagct ggacaatgga gtcaggattc 5821 ctccaagtgg ttggaagtgt gccagatgcg acctgcgaga aaacctctgg ttgaatctga 5881 ctgacggctc tgtcctgtgt ggaaagtggt tctttgacag ctctgggggc aacgggcatg 5941 cgctggagca ttacagagac atgggctacc cactagccgt gaaactggga accatcactc 6001 ctgacggggc agatgtttat tcttttcaag aagaagaacc tgttttggat cctcatttgg 6061 ccaagcactt agcgcatttt ggaattgata tgcttcatat gcatgggaca gagaatgggc 6121 tccaggacaa tgacatcaag ctgagggtca gtgagtggga agtgatccag gagtcgggca 6181 cgaaactgaa gccaatgtat ggtcctggct acacgggtct gaagaacctg ggcaacagct 6241 gctatctcag ctctgtcatg caggccatct tcagcatccc agaattccag agagcgtatg 6301 taggaaacct tcccagaata tttgactact cgcctttaga tccaacacaa gatttcaaca 6361 cacagatgac taagttagga catggccttc tctcaggcca gtattcaaag cctccggtga 6421 aatctgaact cattgaacag gtgatgaagg aggagcacaa gccacagcag aacgggatct 6481 ctccgcgcat gtttaaggcc tttgtaagca agagccaccc ggaattctcc tctaacaggc 6541 agcaagatgc ccaggaattc ttcttgcacc tggtgaatct agtagagagg aaccgcatcg 6601 gctcagaaaa cccaagcgat gtttttcgtt ttttggtgga agaacgcatt cagtgctgtc 6661 agacccggaa agtccgctac acggagaggg tggattacct gatgcagtta cctgtggcca 6721 tggaggcggc aaccaacaag gatgaactga tcgcttatga actaacgaga agggaagcag 6781 aagcaaacag aagacccctt cctgagttgg tacgtgccaa gataccattt agtgcctgcc 6841 ttcaggcctt ctctgaacca gaaaatgttg atgatttctg gagcagtgcc ctacaagcaa 6901 agtctgcggg tgtgaaaaca tctcgctttg cttcattccc tgaatacttg gtagtgcaga 6961 taaagaagtt cacttttggt cttgactggg ttcccaaaaa atttgatgtt tctattgata 7021 tgccagacct acttgatatc aaccatctcc gagccagggg gttacagcca ggagaggaag 7081 aacttccaga catcagcccc cccatagtca ttcctgatga ctcaaaagat cgcctgatga 7141 accaattgat agacccatca gacatcgatg agtcatcagt gatgcagctg gccgagatgg 7201 gtttcccgct ggaagcatgt cgcaaggctg tgtacttcac tggaaatatg ggcgccgagg 7261 tggccttcaa ctggatcatt gttcacatgg aagagccaga ttttgctgag ccgctgacca 7321 tgcctggtta tggaggggca gcttctgctg gagcctctgt ttttggtgct tctggactgg 7381 ataaccaacc tccagaggaa atcgtagcta tcatcacctc catgggattt cagcgaaatc 7441 aggctattca ggcactacga gcaacgaata ataacctgga aagagcactg gattggatct 7501 ttagccaccc tgagtttgaa gaagacagtg attttgtgat tgagatggag aataatgcca 7561 atgcaaacat tatttctgag gccaagcccg aaggacctag agtcaaggat ggatctggaa 7621 catatgagct atttgcattc atcagtcaca tgggaacatc cacaatgagt ggtcattaca 7681 tttgccatat caaaaaggaa ggaagatggg tgatttacaa tgaccacaaa gtttgtgcct 7741 cagaaaggcc ccctaaagac ctgggctaca tgtactttta ccgcaggata ccaagcttaa 7801 ttaatccaac tttcttgtac aaagtggttc gataacgaat tccgcccccc ccccctaacg 7861 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca 7921 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga 7981 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga 8041 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca 8101 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag 8161 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa 8221 gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc cagaaggtac 8281 cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt gtttagtcga 8341 ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac 8401 gatgataata tggccacaac catgaccgag tacaagccca cggtgcgcct cgccacccgc 8461 gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg 8521 cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc 8581 ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg 8641 gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg 8701 cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg 8761 gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac 8821 caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc 8881 gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg 8941 ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg 9001 acccgcaagc ccggtgcctg atgcatcgat gt //