LOCUS Exported 8644 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_0422100-COMP-blac-flag-his SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8644) TITLE Pentamerised full-length protein ectodomains with no N-linked glycans REFERENCE 2 (bases 1 to 8644) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8644 /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 2176..2961 /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 2968..3033 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS 3040..3057 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 3095..3150 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(3149..3168) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(5386..5404) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5472..5576 /gene="bla" /label=AmpR promoter CDS 5577..6437 /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 6608..7196 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7097..7116 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 7350..7367 /label=L4440 /note="L4440 vector, forward primer" protein_bind 7484..7505 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 7520..7550 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 7558..7574 /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 7563..7585 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 7582..7598 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 7582..7598 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" enhancer 7869..8248 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 8249..8452 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 8403..8423 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 8449..8473 /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 ggccgccaac 421 ttcgtgctga gcaacgccaa caccaacaac gtgaacaaga acggcgccaa caagaagaac 481 aacgctaaca acaataatat gaacaaggcc ggcgataaga agaaggccac caagggcaag 541 aagggaaacg ccaacgacaa tgtgaacgag ttcaacttca atttcaacga cctgctcaac 601 gagatcaaca acgccgccac caacgataac aataacaaca acaatgacaa taataacgac 661 aacaacaaca acaataacaa caacaacaac aattacaatg gcaagaacga gatcaatgag 721 aacaacttca acgaagagta caacaagctc gagaagaact tccagatgaa cgccgaggac 781 gatgacaaca accggggcga catcaacatg gaccagttca acgagaagaa aaacaacctg 841 aacgaggata ataacaagat gaagcccaac gccaatgact accagtataa cgagaacgac 901 gccgacaccg acagcgaaga ggacctgacc aacatctgga acaagaatat gcagaacttc 961 gagcccagca ccctgctgac cttcgagatc cccagcaaca gcgaggaata cctgttcgaa 1021 gaggtgcccg aagtgaacac ctacttccgg ggcgtgttct actccaacaa cgaggccgac 1081 gacaacatca tcgagtttct gatcagcgac cccgacggcc aggtcatcta caagaaagag 1141 gccagcgagg gcatcttcta cttctacacc aagaaagtgg gcatctacac cctgaccctc 1201 aagaacaaca agtggatggg caagaaactg accaccgtgg ccctgggcct gggcgagaat 1261 cccgccctga gaagcgacca cgtgaaggac ttcaccaact acatccagcg gatcgtggcc 1321 gagacaaaga agctgaaaaa cgagattaag tacctgagca gcaagcacat gcggcacatc 1381 gagaagatga agaagatcac cggcgcgccg tcgacctcca tcacggccta taagagtgag 1441 ggggagtcag cggagttctc cttcccactc aaccttggag aggaaagcct gcagggagag 1501 ttgagatgga aggcagagaa ggctccttct tcccagtcct ggatcacctt ctccctaaag 1561 aaccaaaagg tgtctgtgca gaagtctact agcaacccca agttccagct gtccgaaacg 1621 ctcccactca cccttcagat accccaggtc tcccttcagt ttgctggttc tggcaacctg 1681 accctgactc tggacagagg gatactgtat caggaagtga acctggtggt gatgaaagtg 1741 actcagcccg acagcaacac tttgacctgt gaggtgatgg gacccacctc acccaagatg 1801 agactgatct tgaagcagga gaatcaggag gccagggtct ccaggcagga gaaagtgatt 1861 caagtgcagg cccctgaagc aggggtgtgg caatgtctac tgagtgaagg tgaagaggtc 1921 aagatggact ccaagatcca ggttttatcc aaagggttga attccggcgg cggctccgga 1981 ggcggtaccg gtggcggcga cctagcccca cagatgcttc gagaactcca ggagactaat 2041 gcggcgctgc aagacgtgag agagctcttg cgacagcagg tcaaggagat caccttcctg 2101 aagaatacgg tgatggaatg tgacgcctgc ggtctagaca gaaacctgcc tccactagca 2161 cctctgggcc cacacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt 2221 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc 2281 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta 2341 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac 2401 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa 2461 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg 2521 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc 2581 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg 2641 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta 2701 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg 2761 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg 2821 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc 2881 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt 2941 gcctcactga ttaagcattg gggccctgac tacaaagacc atgacggtga ttataaagat 3001 catgacatcg attacaagga tgacgatgac aagggccctc accaccacca tcaccactaa 3061 ttaattaagg atcccccgac ctcgacctct ggctaataaa ggaaatttat tttcattgca 3121 atagtgtgtt ggaatttttt gtgtctctca ctcggaagga catatgggag ggcaaatcat 3181 ttggtcgaga tccctcggag atctctagct agaggatcga tccccgcccc ggacgaacta 3241 aacctgacta cgacatctct gccccttctt cgcggggcag tgcatgtaat cccttcagtt 3301 ggttggtaca acttgccaac tgggccctgt tccacatgtg acacgggggg ggaccaaaca 3361 caaaggggtt ctctgactgt agttgacatc cttataaatg gatgtgcaca tttgccaaca 3421 ctgagtggct ttcatcctgg agcagacttt gcagtctgtg gactgcaaca caacattgcc 3481 tttatgtgta actcttggct gaagctctta caccaatgct gggggacatg tacctcccag 3541 gggcccagga agactacggg aggctacacc aacgtcaatc agaggggcct gtgtagctac 3601 cgataagcgg accctcaaga gggcattagc aatagtgttt ataaggcccc cttgttaacc 3661 ctaaacgggt agcatatgct tcccgggtag tagtatatac tatccagact aaccctaatt 3721 caatagcata tgttacccaa cgggaagcat atgctatcga attagggtta gtaaaagggt 3781 cctaaggaac agcgatatct cccaccccat gagctgtcac ggttttattt acatggggtc 3841 aggattccac gagggtagtg aaccatttta gtcacaaggg cagtggctga agatcaagga 3901 gcgggcagtg aactctcctg aatcttcgcc tgcttcttca ttctccttcg tttagctaat 3961 agaataactg ctgagttgtg aacagtaagg tgtatgtgag gtgctcgaaa acaaggtttc 4021 aggtgacgcc cccagaataa aatttggacg gggggttcag tggtggcatt gtgctatgac 4081 accaatataa ccctcacaaa ccccttgggc aataaatact agtgtaggaa tgaaacattc 4141 tgaatatctt taacaataga aatccatggg gtggggacaa gccgtaaaga ctggatgtcc 4201 atctcacacg aatttatggc tatgggcaac acataatcct agtgcaatat gatactgggg 4261 ttattaagat gtgtcccagg cagggaccaa gacaggtgaa ccatgttgtt acactctatt 4321 tgtaacaagg ggaaagagag tggacgccga cagcagcgga ctccactggt tgtctctaac 4381 acccccgaaa attaaacggg gctccacgcc aatggggccc ataaacaaag acaagtggcc 4441 actctttttt ttgaaattgt ggagtggggg cacgcgtcag cccccacacg ccgccctgcg 4501 gttttggact gtaaaataag ggtgtaataa cttggctgat tgtaaccccg ctaaccactg 4561 cggtcaaacc acttgcccac aaaaccacta atggcacccc ggggaatacc tgcataagta 4621 ggtgggcggg ccaagatagg ggcgcgattg ctgcgatctg gaggacaaat tacacacact 4681 tgcgcctgag cgccaagcac agggttgttg gtcctcatat tcacgaggtc gctgagagca 4741 cggtgggcta atgttgccat gggtagcata tactacccaa atatctggat agcatatgct 4801 atcctaatct atatctgggt agcataggct atcctaatct atatctgggt agcatatgct 4861 atcctaatct atatctgggt agtatatgct atcctaattt atatctgggt agcataggct 4921 atcctaatct atatctgggt agcatatgct atcctaatct atatctgggt agtatatgct 4981 atcctaatct gtatccgggt agcatatgct atcctaatag agattagggt agtatatgct 5041 atcctaattt atatctgggt agcatatact acccaaatat ctggatagca tatgctatcc 5101 taatctatat ctgggtagca tatgctatcc taatctatat ctgggtagca taggctatcc 5161 taatctatat ctgggtagca tatgctatcc taatctatat ctgggtagta tatgctatcc 5221 taatttatat ctgggtagca taggctatcc taatctatat ctgggtagca tatgctatcc 5281 taatctatat ctgggtagta tatgctatcc taatctgtat ccgggtagca tatgctatcc 5341 tcatgataag ctgtcaaaca tgagaatttt cttgaagacg aaagggcctc gtgatacgcc 5401 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc 5461 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc 5521 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga 5581 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt 5641 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag 5701 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag 5761 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg 5821 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg 5881 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca 5941 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag 6001 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc 6061 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg 6121 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc 6181 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg 6241 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg 6301 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga 6361 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac 6421 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa 6481 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca 6541 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag 6601 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac 6661 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa 6721 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc 6781 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag 6841 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac 6901 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc 6961 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc 7021 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca 7081 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc 7141 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg 7201 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct 7261 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata 7321 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc 7381 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg 7441 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca 7501 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg 7561 tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc aagctctagc 7621 tagaggtcga ccaattctca tgtttgacag cttatcatcg cagatccggg caacgttgtt 7681 gccattgctg caggcgcaga actggtaggt atggaagatc tatacattga atcaatattg 7741 gcaattagcc atattagtca ttggttatat agcataaatc aatattggct attggccatt 7801 gcatacgttg tatctatatc ataatatgta catttatatt ggctcatgtc caatatgacc 7861 gccatgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt 7921 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg 7981 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc 8041 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc 8101 agtacatcaa gtgtatcata tgccaagtcc gccccctatt gacgtcaatg acggtaaatg 8161 gcccgcctgg cattatgccc agtacatgac cttacgggac tttcctactt ggcagtacat 8221 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca ccaatgggcg 8281 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag 8341 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaataacc ccgccccgtt 8401 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcgtttagt 8461 gaaccgtcag atcctcactc tcttccgcat cgctgtctgc gagggccagc tgttgggctc 8521 gcggttgagg acaaactctt cgcggtcttt ccagtactct tggatcggaa acccgtcggc 8581 ctccgaacgg tactccgcca ccgagggacc tgagcgagtc cgcatcgacc ggatcggaaa 8641 acct //