LOCUS Exported 8143 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 PF3D7_1333300-COMP-blac-flag-his SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8143) TITLE Pentamerised full-length protein ectodomains with no N-linked glycans REFERENCE 2 (bases 1 to 8143) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8143 /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 1675..2460 /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 2467..2532 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS 2539..2556 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 2594..2649 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(2648..2667) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(4885..4903) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4971..5075 /gene="bla" /label=AmpR promoter CDS 5076..5936 /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 6107..6695 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6596..6615 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6849..6866 /label=L4440 /note="L4440 vector, forward primer" protein_bind 6983..7004 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 7019..7049 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 7057..7073 /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 7062..7084 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 7081..7097 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 7081..7097 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" enhancer 7368..7747 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 7748..7951 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 7902..7922 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 7948..7972 /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 aacggcctgg aaatctacat caagatgaag ggcaagaaac tgaagtgcat caaagagcgg 481 atcaacaagg acaccctggt catcagcaag ttcaagacca accacaagaa cgccctggtg 541 tccctgtaca tctacgacgt ggacgtgaac gagaagaact tccacaccca gcccaagctg 601 cccatcttcg agacagtgaa caagcacgac atcaagaccg ccttcaccac cttctacagc 661 agcagctaca gcttctgtgc ctacaacaac gccgacaagc tgatcgaggt gtacttcgag 721 attaagtacg gcaccgaggc ccgggactac gcccaggtgg ccaagaccga gcacctgaac 781 cagggcaccc tgtacctgaa gcagatcctg gaccagatga acatcttcca cgtgaacctg 841 aagaggatcc gcagcgacga ggacaaagag aagaaggccc acgataagct gaacgacgcc 901 ggcgcgccgt cgacctccat cacggcctat aagagtgagg gggagtcagc ggagttctcc 961 ttcccactca accttggaga ggaaagcctg cagggagagt tgagatggaa ggcagagaag 1021 gctccttctt cccagtcctg gatcaccttc tccctaaaga accaaaaggt gtctgtgcag 1081 aagtctacta gcaaccccaa gttccagctg tccgaaacgc tcccactcac ccttcagata 1141 ccccaggtct cccttcagtt tgctggttct ggcaacctga ccctgactct ggacagaggg 1201 atactgtatc aggaagtgaa cctggtggtg atgaaagtga ctcagcccga cagcaacact 1261 ttgacctgtg aggtgatggg acccacctca cccaagatga gactgatctt gaagcaggag 1321 aatcaggagg ccagggtctc caggcaggag aaagtgattc aagtgcaggc ccctgaagca 1381 ggggtgtggc aatgtctact gagtgaaggt gaagaggtca agatggactc caagatccag 1441 gttttatcca aagggttgaa ttccggcggc ggctccggag gcggtaccgg tggcggcgac 1501 ctagccccac agatgcttcg agaactccag gagactaatg cggcgctgca agacgtgaga 1561 gagctcttgc gacagcaggt caaggagatc accttcctga agaatacggt gatggaatgt 1621 gacgcctgcg gtctagacag aaacctgcct ccactagcac ctctgggccc acacccagaa 1681 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa 1741 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg 1801 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa 1861 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc 1921 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc 1981 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta 2041 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag 2101 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca 2161 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata 2221 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc 2281 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca 2341 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca 2401 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg 2461 ggccctgact acaaagacca tgacggtgat tataaagatc atgacatcga ttacaaggat 2521 gacgatgaca agggccctca ccaccaccat caccactaat taattaagga tcccccgacc 2581 tcgacctctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg 2641 tgtctctcac tcggaaggac atatgggagg gcaaatcatt tggtcgagat ccctcggaga 2701 tctctagcta gaggatcgat ccccgccccg gacgaactaa acctgactac gacatctctg 2761 ccccttcttc gcggggcagt gcatgtaatc ccttcagttg gttggtacaa cttgccaact 2821 gggccctgtt ccacatgtga cacggggggg gaccaaacac aaaggggttc tctgactgta 2881 gttgacatcc ttataaatgg atgtgcacat ttgccaacac tgagtggctt tcatcctgga 2941 gcagactttg cagtctgtgg actgcaacac aacattgcct ttatgtgtaa ctcttggctg 3001 aagctcttac accaatgctg ggggacatgt acctcccagg ggcccaggaa gactacggga 3061 ggctacacca acgtcaatca gaggggcctg tgtagctacc gataagcgga ccctcaagag 3121 ggcattagca atagtgttta taaggccccc ttgttaaccc taaacgggta gcatatgctt 3181 cccgggtagt agtatatact atccagacta accctaattc aatagcatat gttacccaac 3241 gggaagcata tgctatcgaa ttagggttag taaaagggtc ctaaggaaca gcgatatctc 3301 ccaccccatg agctgtcacg gttttattta catggggtca ggattccacg agggtagtga 3361 accattttag tcacaagggc agtggctgaa gatcaaggag cgggcagtga actctcctga 3421 atcttcgcct gcttcttcat tctccttcgt ttagctaata gaataactgc tgagttgtga 3481 acagtaaggt gtatgtgagg tgctcgaaaa caaggtttca ggtgacgccc ccagaataaa 3541 atttggacgg ggggttcagt ggtggcattg tgctatgaca ccaatataac cctcacaaac 3601 cccttgggca ataaatacta gtgtaggaat gaaacattct gaatatcttt aacaatagaa 3661 atccatgggg tggggacaag ccgtaaagac tggatgtcca tctcacacga atttatggct 3721 atgggcaaca cataatccta gtgcaatatg atactggggt tattaagatg tgtcccaggc 3781 agggaccaag acaggtgaac catgttgtta cactctattt gtaacaaggg gaaagagagt 3841 ggacgccgac agcagcggac tccactggtt gtctctaaca cccccgaaaa ttaaacgggg 3901 ctccacgcca atggggccca taaacaaaga caagtggcca ctcttttttt tgaaattgtg 3961 gagtgggggc acgcgtcagc ccccacacgc cgccctgcgg ttttggactg taaaataagg 4021 gtgtaataac ttggctgatt gtaaccccgc taaccactgc ggtcaaacca cttgcccaca 4081 aaaccactaa tggcaccccg gggaatacct gcataagtag gtgggcgggc caagataggg 4141 gcgcgattgc tgcgatctgg aggacaaatt acacacactt gcgcctgagc gccaagcaca 4201 gggttgttgg tcctcatatt cacgaggtcg ctgagagcac ggtgggctaa tgttgccatg 4261 ggtagcatat actacccaaa tatctggata gcatatgcta tcctaatcta tatctgggta 4321 gcataggcta tcctaatcta tatctgggta gcatatgcta tcctaatcta tatctgggta 4381 gtatatgcta tcctaattta tatctgggta gcataggcta tcctaatcta tatctgggta 4441 gcatatgcta tcctaatcta tatctgggta gtatatgcta tcctaatctg tatccgggta 4501 gcatatgcta tcctaataga gattagggta gtatatgcta tcctaattta tatctgggta 4561 gcatatacta cccaaatatc tggatagcat atgctatcct aatctatatc tgggtagcat 4621 atgctatcct aatctatatc tgggtagcat aggctatcct aatctatatc tgggtagcat 4681 atgctatcct aatctatatc tgggtagtat atgctatcct aatttatatc tgggtagcat 4741 aggctatcct aatctatatc tgggtagcat atgctatcct aatctatatc tgggtagtat 4801 atgctatcct aatctgtatc cgggtagcat atgctatcct catgataagc tgtcaaacat 4861 gagaattttc ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca 4921 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc 4981 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct 5041 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg 5101 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg 5161 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc 5221 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca 5281 cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac 5341 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa 5401 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg 5461 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt 5521 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg 5581 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc 5641 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga 5701 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta 5761 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc 5821 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg 5881 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt 5941 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa 6001 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt 6061 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 6121 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 6181 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga 6241 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag 6301 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata 6361 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg 6421 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga 6481 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca 6541 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa 6601 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt 6661 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac 6721 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt 6781 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga 6841 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc 6901 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag 6961 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt 7021 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca 7081 caggaaacag ctatgaccat gattacgcca agctctagct agaggtcgac caattctcat 7141 gtttgacagc ttatcatcgc agatccgggc aacgttgttg ccattgctgc aggcgcagaa 7201 ctggtaggta tggaagatct atacattgaa tcaatattgg caattagcca tattagtcat 7261 tggttatata gcataaatca atattggcta ttggccattg catacgttgt atctatatca 7321 taatatgtac atttatattg gctcatgtcc aatatgaccg ccatgttgac attgattatt 7381 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt 7441 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc 7501 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg 7561 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat 7621 gccaagtccg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca 7681 gtacatgacc ttacgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat 7741 taccatggtg atgcggtttt ggcagtacac caatgggcgt ggatagcggt ttgactcacg 7801 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca 7861 acgggacttt ccaaaatgtc gtaataaccc cgccccgttg acgcaaatgg gcggtaggcg 7921 tgtacggtgg gaggtctata taagcagagc tcgtttagtg aaccgtcaga tcctcactct 7981 cttccgcatc gctgtctgcg agggccagct gttgggctcg cggttgagga caaactcttc 8041 gcggtctttc cagtactctt ggatcggaaa cccgtcggcc tccgaacggt actccgccac 8101 cgagggacct gagcgagtcc gcatcgaccg gatcggaaaa cct //