LOCUS Exported 10547 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses pentamerised full-length protein ectodomain with no N-linked glycans in mammalian cells. Contains a C-terminal rat Cd4 d3+4 tag, COMP pentamerisation domain, flag- and 6xHis tags. . ACCESSION . VERSION . KEYWORDS SIAP-1-COMP-blac-flag-his SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 10547) TITLE Pentamerised full-length protein ectodomains with no N-linked glycans REFERENCE 2 (bases 1 to 10547) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..10547 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 310..329 /label=pMT2-F /note="Synthetic intron, forward primer" regulatory 352..361 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 4081..4866 /codon_start=1 /gene="bla(M)" /product="beta-lactamase lacking the signal sequence" /label=bla(M) /note="allows cytosolic expression of beta-lactamase" /translation="PETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMST FKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMS DNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATT LRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAA LGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW" CDS 4873..4938 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS 4945..4962 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 4998..5053 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(5052..5071) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(7289..7307) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 7375..7479 /gene="bla" /label=AmpR promoter CDS 7480..8340 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" rep_origin 8511..9099 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 9000..9019 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 9253..9270 /label=L4440 /note="L4440 vector, forward primer" protein_bind 9387..9408 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 9423..9453 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 9461..9477 /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 9466..9488 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 9485..9501 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 9485..9501 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" enhancer 9772..10151 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 10152..10355 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 10306..10326 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 10352..10376 /label=LNCX /note="Human CMV promoter, forward primer" ORIGIN 1 ctcgagaaag gcgtctaacc agtcacagtc gcaaggtagg ctgagcaccg tggcgggcgg 61 cagcgggtgg cggtcggggt tgtttctggc ggaggtgctg ctgatgatgt aattaaagta 121 ggcggtcttg agacggcgga tggtcgaggt gaggtgtggc aggcttgaga tccagctgtt 181 ggggtgagta ctccctctca aaagcgggca ttacttctgc gctaagattg tcagtttcca 241 aaaacgagga ggatttgata ttcacctggc ccgatctggc catacacttg agtgacaatg 301 acatccactt tgcctttctc tccacaggtg tccactccca ggtccaagtt tgccaccatg 361 gagtttcaga cccaggtact catgtccctg ctgctctgca tgtctggtgc ggccgcctac 421 aacatcaaga gcggcaacac cgccaacaat atcaaatacg tgaacgtgct ggacaacgac 481 cgggacatca acacccacag cgtgctgccc gaggtggaaa acgtgatcga gcggaaggac 541 atctaccggc agatcaactt catggaaacc ttcgtgtcca gcaacaacat gatgcacgac 601 agagagaagc acaccagcaa cgacgccggc agctacgaga tcaccggcat cgtggacggc 661 atgaagatcg gccaccccat cagcgtggcc ctgggcagcc agtacagcaa ctacttcgac 721 tacctgcaga tcgtgcacct ggactacacc aacagccggt tcagcttcac cgtgggcgag 781 ggcaagtact acctgcggac ctacggcagc acctacatga cccctagcgc catcaagatc 841 aaggtgccct gcgagaagtg caagttcatc aacagcgagt acagcggcat catcaagatt 901 atcccttacg agacaaacaa caacctgttc atctacaact gggtgctgca gaccagcagc 961 cctctggccc tggaaaacat caataccgtg ttcagcgacg aggccgacct gatccacggc 1021 aacagcctga gcgaggaatt caagatcgac agcagcgccg ctgccaccag cctgaacacc 1081 ttctatggca tcgtgctgca cggcatctgg tcctccgagt acgccgagcg gctgctgacc 1141 gtgatctccg agttccccga ctgcgtgaag atgagcgccc acgacaagaa cgccagaagc 1201 aagcagcgga agaaccagaa gtggatcctg gtcaacgagg acctgggctc cttcgacatg 1261 aagatggaag tgtgcgagga agtgaactgc gactactccg ccatcatcca cgtgtccaag 1321 cacgccttcg agtactccaa gaaactggtg cacaaccggg gcagaaacgg ccggtactac 1381 agcagacggg tggaaaagat cctgatccgg gccctgctga gcctggactt cagcctgttt 1441 atcacctact tccagcagaa acacggcgtg accctgctgg accctcagta cgactacgag 1501 ctgatcacca acatggccgg ctacagctcc aacaactacc agagctggaa ccacaacctg 1561 gaagaactgg tggaactggc caccagctgg gacgagtacc ccaagggcct gcagaaggtg 1621 cagggcctga gctacctgct gcggagaaag aacggcgcca agcaccccgt gtaccccaca 1681 gcccctgccg tggcctttcc agccggcagc cagaacaacg ccttcatcga gttcatggaa 1741 tccgccttcg tgaactacgt ggacatcagc cacctggtca tccacgaggt ggcccacttc 1801 atctgggtca acaccgtgtc caaagaactg aaagagaaat ggatccagat cgggcagtgg 1861 tacaaagagc ccctgagccc cagcgaatgg gccacaaagc tggaagtgga attcgtgtcc 1921 gcctacgccc atgataagaa ccccgccgag gacttcgccg agagcatggc cacctacgtg 1981 ctgaacagca agctgctgaa ctccagaagc ttcgacaagt tcaagtggat tcaggacaat 2041 ctgttcggcg gaggcttcta catcaccacc ggcacccaca agttcgacgt gatcaacctg 2101 ggcaatgagg tgtactactt ccccggcaaa gtgaccagag tgcgggccaa ggtgctgggc 2161 tcccctaccg aggacaagct ggtcaagatc tacatcagcc tgctgtccag cgacggcagc 2221 gagggctgtg ccaagcacgg ctacgcccgg atcttcagcg agcagcagac cttccgggac 2281 ctgtacttcc acaccgagga tagaagcccc tgcagccaca agctgtacgg cgagttcacc 2341 atgaacaagc acgagagcag aggccggtgg accgccgagt ccatgatctt caccggcgag 2401 aacaacatcg agagatacgt gggcctgggc tctttccact tctacctgta cgtgaacaac 2461 cagaacgagg acgtggaaaa gcctatccct ctgctggact ccatcagcat ctacacacac 2521 aacgccgccg agactaacga cgctctgctg cggctgcacg tgatggtgct ggaaaatgaa 2581 ctgatcaaag agcacggcgg accttacgcc agcttcgccg cccacgagaa caaggcctac 2641 tcctacgaga gccggaccta caagatgtac cctcccgagt tcaacaccct gatgctgaag 2701 gccgactact tcatcagaga tatcaacacc agaggcttcc gcgaagtgaa catggacagc 2761 tgcaagagct acaccaatat ggacacccgg aacctgaagt gcttccaggt gctgaacccc 2821 gtgaccatcc ccaagtactg tatcggctcc acctactttc tgagacaggt gtccatcgag 2881 gatatcgccg gcaatctgga aaccgtgaat atcgccagcg acaagtacag cgccagactg 2941 caccctatcg gcgtgcggga caagcagaaa cccgtggtgt ccaacgtgcg ggtgtcctcc 3001 aagcccgcca acgagtacca cgacggcgag acaatcgtgt ccctgagctt caacgtgcac 3061 gacaacctgg ccggcgtgta ctacatcttc gtgtacctgc gggatcccca cggcggaaag 3121 cacagaagcg acatcgaccg ggccagcctg cctaccggca cagagaacaa gcagatcaat 3181 cacaagatcc tgctgcccaa gggctccatg ggcggcacct ggatgctgga agagatcaag 3241 gccgtggact cctgcaagaa cgaggcccgg aatatctaca cccactccgt gtacgtgcag 3301 aacgacggcg cgccgtcgac ctccatcacg gcctataaga gtgaggggga gtcagcggag 3361 ttctccttcc cactcaacct tggagaggaa agcctgcagg gagagttgag atggaaggca 3421 gagaaggctc cttcttccca gtcctggatc accttctccc taaagaacca aaaggtgtct 3481 gtgcagaagt ctactagcaa ccccaagttc cagctgtccg aaacgctccc actcaccctt 3541 cagatacccc aggtctccct tcagtttgct ggttctggca acctgaccct gactctggac 3601 agagggatac tgtatcagga agtgaacctg gtggtgatga aagtgactca gcccgacagc 3661 aacactttga cctgtgaggt gatgggaccc acctcaccca agatgagact gatcttgaag 3721 caggagaatc aggaggccag ggtctccagg caggagaaag tgattcaagt gcaggcccct 3781 gaagcagggg tgtggcaatg tctactgagt gaaggtgaag aggtcaagat ggactccaag 3841 atccaggttt tatccaaagg gttgaattcc ggcggcggct ccggaggcgg taccggtggc 3901 ggcgacctag ccccacagat gcttcgagaa ctccaggaga ctaatgcggc gctgcaagac 3961 gtgagagagc tcttgcgaca gcaggtcaag gagatcacct tcctgaagaa tacggtgatg 4021 gaatgtgacg cctgcggtct agacagaaac ctgcctccac tagcacctct gggcccacac 4081 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac 4141 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt 4201 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc 4261 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca 4321 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc 4381 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag 4441 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa 4501 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg 4561 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa 4621 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg 4681 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt 4741 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt 4801 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag 4861 cattggggcc ctgactacaa agaccatgac ggtgattata aagatcatga catcgattac 4921 aaggatgacg atgacaaggg ccctcaccac caccatcacc actaataata aggatccccc 4981 gacctcgacc tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt 5041 tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttggtcg agatccctcg 5101 gagatctcta gctagaggat cgatccccgc cccggacgaa ctaaacctga ctacgacatc 5161 tctgcccctt cttcgcgggg cagtgcatgt aatcccttca gttggttggt acaacttgcc 5221 aactgggccc tgttccacat gtgacacggg gggggaccaa acacaaaggg gttctctgac 5281 tgtagttgac atccttataa atggatgtgc acatttgcca acactgagtg gctttcatcc 5341 tggagcagac tttgcagtct gtggactgca acacaacatt gcctttatgt gtaactcttg 5401 gctgaagctc ttacaccaat gctgggggac atgtacctcc caggggccca ggaagactac 5461 gggaggctac accaacgtca atcagagggg cctgtgtagc taccgataag cggaccctca 5521 agagggcatt agcaatagtg tttataaggc ccccttgtta accctaaacg ggtagcatat 5581 gcttcccggg tagtagtata tactatccag actaacccta attcaatagc atatgttacc 5641 caacgggaag catatgctat cgaattaggg ttagtaaaag ggtcctaagg aacagcgata 5701 tctcccaccc catgagctgt cacggtttta tttacatggg gtcaggattc cacgagggta 5761 gtgaaccatt ttagtcacaa gggcagtggc tgaagatcaa ggagcgggca gtgaactctc 5821 ctgaatcttc gcctgcttct tcattctcct tcgtttagct aatagaataa ctgctgagtt 5881 gtgaacagta aggtgtatgt gaggtgctcg aaaacaaggt ttcaggtgac gcccccagaa 5941 taaaatttgg acggggggtt cagtggtggc attgtgctat gacaccaata taaccctcac 6001 aaaccccttg ggcaataaat actagtgtag gaatgaaaca ttctgaatat ctttaacaat 6061 agaaatccat ggggtgggga caagccgtaa agactggatg tccatctcac acgaatttat 6121 ggctatgggc aacacataat cctagtgcaa tatgatactg gggttattaa gatgtgtccc 6181 aggcagggac caagacaggt gaaccatgtt gttacactct atttgtaaca aggggaaaga 6241 gagtggacgc cgacagcagc ggactccact ggttgtctct aacacccccg aaaattaaac 6301 ggggctccac gccaatgggg cccataaaca aagacaagtg gccactcttt tttttgaaat 6361 tgtggagtgg gggcacgcgt cagcccccac acgccgccct gcggttttgg actgtaaaat 6421 aagggtgtaa taacttggct gattgtaacc ccgctaacca ctgcggtcaa accacttgcc 6481 cacaaaacca ctaatggcac cccggggaat acctgcataa gtaggtgggc gggccaagat 6541 aggggcgcga ttgctgcgat ctggaggaca aattacacac acttgcgcct gagcgccaag 6601 cacagggttg ttggtcctca tattcacgag gtcgctgaga gcacggtggg ctaatgttgc 6661 catgggtagc atatactacc caaatatctg gatagcatat gctatcctaa tctatatctg 6721 ggtagcatag gctatcctaa tctatatctg ggtagcatat gctatcctaa tctatatctg 6781 ggtagtatat gctatcctaa tttatatctg ggtagcatag gctatcctaa tctatatctg 6841 ggtagcatat gctatcctaa tctatatctg ggtagtatat gctatcctaa tctgtatccg 6901 ggtagcatat gctatcctaa tagagattag ggtagtatat gctatcctaa tttatatctg 6961 ggtagcatat actacccaaa tatctggata gcatatgcta tcctaatcta tatctgggta 7021 gcatatgcta tcctaatcta tatctgggta gcataggcta tcctaatcta tatctgggta 7081 gcatatgcta tcctaatcta tatctgggta gtatatgcta tcctaattta tatctgggta 7141 gcataggcta tcctaatcta tatctgggta gcatatgcta tcctaatcta tatctgggta 7201 gtatatgcta tcctaatctg tatccgggta gcatatgcta tcctcatgat aagctgtcaa 7261 acatgagaat tttcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat 7321 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga 7381 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa 7441 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt 7501 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg 7561 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg 7621 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg 7681 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag 7741 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca 7801 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg 7861 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc 7921 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg 7981 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg 8041 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac 8101 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg 8161 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg 8221 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact 8281 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa 8341 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt 8401 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag 8461 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct 8521 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt 8581 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg 8641 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct 8701 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc 8761 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg 8821 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa 8881 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg 8941 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg 9001 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga 9061 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt 9121 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct 9181 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga 9241 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg 9301 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg 9361 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag 9421 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt 9481 cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt cgaccaattc 9541 tcatgtttga cagcttatca tcgcagatcc gggcaacgtt gttgccattg ctgcaggcgc 9601 agaactggta ggtatggaag atctatacat tgaatcaata ttggcaatta gccatattag 9661 tcattggtta tatagcataa atcaatattg gctattggcc attgcatacg ttgtatctat 9721 atcataatat gtacatttat attggctcat gtccaatatg accgccatgt tgacattgat 9781 tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg 9841 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc 9901 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt 9961 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc 10021 atatgccaag tccgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg 10081 cccagtacat gaccttacgg gactttccta cttggcagta catctacgta ttagtcatcg 10141 ctattaccat ggtgatgcgg ttttggcagt acaccaatgg gcgtggatag cggtttgact 10201 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa 10261 atcaacggga ctttccaaaa tgtcgtaata accccgcccc gttgacgcaa atgggcggta 10321 ggcgtgtacg gtgggaggtc tatataagca gagctcgttt agtgaaccgt cagatcctca 10381 ctctcttccg catcgctgtc tgcgagggcc agctgttggg ctcgcggttg aggacaaact 10441 cttcgcggtc tttccagtac tcttggatcg gaaacccgtc ggcctccgaa cggtactccg 10501 ccaccgaggg acctgagcga gtccgcatcg accggatcgg aaaacct //