LOCUS       sequence-223542-        9203 bp    DNA     circular SYN 15-APR-2025
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    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9203)
  AUTHORS   Knockel J, Dundas K, Yang ASP, Galaway F, Metcalf T, Gemert GV, 
            Sauerwein RW, Rayner JC, Billker O, Wright GJ
  TITLE     Systematic Identification of Plasmodium Falciparum Sporozoite 
            Membrane Protein Interactions Reveals an Essential Role for the p24 
            Complex in Host Infection.
  JOURNAL   Mol Cell Proteomics. 2021;20:100038. doi: 10.1074/mcp.RA120.002432. 
            Epub 2021 Jan 27.
  PUBMED    33515807
REFERENCE   2  (bases 1 to 9203)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Cell 
            Proteomics."; date: "2021"; volume: "20:100038. doi"; pages: " 
            10.1074/mcp.RA120.002432. Epub 2021 Jan 27"
COMMENT     Sequence Label: PIESP15-COMP-blac-flag-his
FEATURES             Location/Qualifiers
     source          1..9203
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     310..329
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      352..361
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2737..3522
                     /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             3529..3594
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3601..3618
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    3654..3709
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3708..3727)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5945..5963)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6031..6135
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6136..6204,6205..6996)
                     /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      7167..7755
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7656..7675
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7909..7926
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8043..8064
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8079..8084,8085..8102,8103..8109)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8117..8133
                     /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     8122..8144
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8141..8157
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8141..8157
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        8428..8807
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8808..9011
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8962..8982
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     9008..9032
                     /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 ggccgccaag
      421 aacagcgaca ccaagaacaa cgcctacaac gactactaca acacctacct gagcaagatc
      481 cacagcttca aggccccttt caacgtcgag aacctggaaa acggctggaa cgtggactac
      541 agcagcgtgt ccaccaacaa gcacgtggtg ctgatcccca gcatctacaa cagacggggc
      601 gtgctgtaca acaagaaccc catcaactcc gacaacttca tcatcgagtt caatttcttc
      661 atggaacgga agtactacaa agagaagatc gacgagaagt actataagga cgagaagaac
      721 gacgtgcacg agatcgtgta caagaccaac agcgacgaca acctggccac cagcaacaag
      781 gccaacaaca aagagcagaa gcagatcaag cggaacggct tcgccctgtg gatcctgaag
      841 gaccccttcg tgatcaaaga ggccggcagc aacgtcaacc tggtgctgca ggaagaagag
      901 tacaacatct tcggctacaa gaacaggttc aacggcatcg gcatcctgtt ccagctgaag
      961 aagtctacca acaaagctaa caataaggct aataacaaca acaattacta ctactataac
     1021 aactacaacg atatcctgaa cgtggccatc atcctgaaca acggcgacaa gagcatcaat
     1081 agcgacgact tcaccatctt caagaacatc cccatcgaca tgattatgat caacaacctg
     1141 atccacacca agatcatctg caacaagaaa gagatcatca ttgccttcta ctaccccaag
     1201 agcaacaact atatcaacac cctcatcatg aagaacaata tcgccaccga gaattatatc
     1261 gccttcacct ccttcaatta caaagaggac gacaacaaaa acatcgacta caatgacgtg
     1321 aacaacaagt attacgagaa ccagaacggc tacctgccca ccttcgtggg catcaaacag
     1381 gtgctgacct tcaacctgaa caacaacaat aacaacaaca acaatatcaa catcaatgag
     1441 gccaacatta acaataataa ttataacaac aacaacaaca atgataacaa caagatcaac
     1501 tacgaccaga actacaacaa ggacatcagc gccaacgaca tcaacgccgc cttcaacgat
     1561 gtgctcaaca acatcaaccc cgagaagaat accatcaacc aggccatcga cgtgctgaac
     1621 agcatcacca agctgctgca gaaattcatg aaccaccaga tcaacagcga gaagaagctc
     1681 ctccagaaca tcaacatcct caacgagaag ctgcagttcg tgcagaccga gctgaaagtg
     1741 atcaagaaga atctggccaa tcaggccgag gaacccaaga acctgcagaa gatgttctcc
     1801 acagagctga ccggcctgaa gaacctgttt cacacccaca cccagcacca caagaagaac
     1861 atcgaggaca tcacaaacaa gctgaccaag aagattgagc ggaacaacga gctgaagatc
     1921 ctgaccgaga aggcccagaa actggaacag atcatcaaca agggcgcgcc gtcgacctcc
     1981 atcacggcct ataagagtga gggggagtca gcggagttct ccttcccact caaccttgga
     2041 gaggaaagcc tgcagggaga gttgagatgg aaggcagaga aggctccttc ttcccagtcc
     2101 tggatcacct tctccctaaa gaaccaaaag gtgtctgtgc agaagtctac tagcaacccc
     2161 aagttccagc tgtccgaaac gctcccactc acccttcaga taccccaggt ctcccttcag
     2221 tttgctggtt ctggcaacct gaccctgact ctggacagag ggatactgta tcaggaagtg
     2281 aacctggtgg tgatgaaagt gactcagccc gacagcaaca ctttgacctg tgaggtgatg
     2341 ggacccacct cacccaagat gagactgatc ttgaagcagg agaatcagga ggccagggtc
     2401 tccaggcagg agaaagtgat tcaagtgcag gcccctgaag caggggtgtg gcaatgtcta
     2461 ctgagtgaag gtgaagaggt caagatggac tccaagatcc aggttttatc caaagggttg
     2521 aattccggcg gcggctccgg aggcggtacc ggtggcggcg acctagcccc acagatgctt
     2581 cgagaactcc aggagactaa tgcggcgctg caagacgtga gagagctctt gcgacagcag
     2641 gtcaaggaga tcaccttcct gaagaatacg gtgatggaat gtgacgcctg cggtctagac
     2701 agaaacctgc ctccactagc acctctgggc ccacacccag aaacgctggt gaaagtaaaa
     2761 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     2821 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     2881 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     2941 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     3001 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     3061 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     3121 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     3181 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     3241 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     3301 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     3361 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     3421 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     3481 agacagatcg ctgagatagg tgcctcactg attaagcatt ggggccctga ctacaaagac
     3541 catgacggtg attataaaga tcatgacatc gattacaagg atgacgatga caagggccct
     3601 caccaccacc atcaccacta ataataagga tcccccgacc tcgacctctg gctaataaag
     3661 gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac
     3721 atatgggagg gcaaatcatt tggtcgagat ccctcggaga tctctagcta gaggatcgat
     3781 ccccgccccg gacgaactaa acctgactac gacatctctg ccccttcttc gcggggcagt
     3841 gcatgtaatc ccttcagttg gttggtacaa cttgccaact gggccctgtt ccacatgtga
     3901 cacggggggg gaccaaacac aaaggggttc tctgactgta gttgacatcc ttataaatgg
     3961 atgtgcacat ttgccaacac tgagtggctt tcatcctgga gcagactttg cagtctgtgg
     4021 actgcaacac aacattgcct ttatgtgtaa ctcttggctg aagctcttac accaatgctg
     4081 ggggacatgt acctcccagg ggcccaggaa gactacggga ggctacacca acgtcaatca
     4141 gaggggcctg tgtagctacc gataagcgga ccctcaagag ggcattagca atagtgttta
     4201 taaggccccc ttgttaaccc taaacgggta gcatatgctt cccgggtagt agtatatact
     4261 atccagacta accctaattc aatagcatat gttacccaac gggaagcata tgctatcgaa
     4321 ttagggttag taaaagggtc ctaaggaaca gcgatatctc ccaccccatg agctgtcacg
     4381 gttttattta catggggtca ggattccacg agggtagtga accattttag tcacaagggc
     4441 agtggctgaa gatcaaggag cgggcagtga actctcctga atcttcgcct gcttcttcat
     4501 tctccttcgt ttagctaata gaataactgc tgagttgtga acagtaaggt gtatgtgagg
     4561 tgctcgaaaa caaggtttca ggtgacgccc ccagaataaa atttggacgg ggggttcagt
     4621 ggtggcattg tgctatgaca ccaatataac cctcacaaac cccttgggca ataaatacta
     4681 gtgtaggaat gaaacattct gaatatcttt aacaatagaa atccatgggg tggggacaag
     4741 ccgtaaagac tggatgtcca tctcacacga atttatggct atgggcaaca cataatccta
     4801 gtgcaatatg atactggggt tattaagatg tgtcccaggc agggaccaag acaggtgaac
     4861 catgttgtta cactctattt gtaacaaggg gaaagagagt ggacgccgac agcagcggac
     4921 tccactggtt gtctctaaca cccccgaaaa ttaaacgggg ctccacgcca atggggccca
     4981 taaacaaaga caagtggcca ctcttttttt tgaaattgtg gagtgggggc acgcgtcagc
     5041 ccccacacgc cgccctgcgg ttttggactg taaaataagg gtgtaataac ttggctgatt
     5101 gtaaccccgc taaccactgc ggtcaaacca cttgcccaca aaaccactaa tggcaccccg
     5161 gggaatacct gcataagtag gtgggcgggc caagataggg gcgcgattgc tgcgatctgg
     5221 aggacaaatt acacacactt gcgcctgagc gccaagcaca gggttgttgg tcctcatatt
     5281 cacgaggtcg ctgagagcac ggtgggctaa tgttgccatg ggtagcatat actacccaaa
     5341 tatctggata gcatatgcta tcctaatcta tatctgggta gcataggcta tcctaatcta
     5401 tatctgggta gcatatgcta tcctaatcta tatctgggta gtatatgcta tcctaattta
     5461 tatctgggta gcataggcta tcctaatcta tatctgggta gcatatgcta tcctaatcta
     5521 tatctgggta gtatatgcta tcctaatctg tatccgggta gcatatgcta tcctaataga
     5581 gattagggta gtatatgcta tcctaattta tatctgggta gcatatacta cccaaatatc
     5641 tggatagcat atgctatcct aatctatatc tgggtagcat atgctatcct aatctatatc
     5701 tgggtagcat aggctatcct aatctatatc tgggtagcat atgctatcct aatctatatc
     5761 tgggtagtat atgctatcct aatttatatc tgggtagcat aggctatcct aatctatatc
     5821 tgggtagcat atgctatcct aatctatatc tgggtagtat atgctatcct aatctgtatc
     5881 cgggtagcat atgctatcct catgataagc tgtcaaacat gagaattttc ttgaagacga
     5941 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     6001 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     6061 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     6121 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     6181 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     6241 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     6301 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     6361 ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg catacactat
     6421 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     6481 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     6541 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     6601 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     6661 cgtgacacca cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa
     6721 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     6781 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     6841 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     6901 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     6961 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     7021 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     7081 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     7141 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     7201 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     7261 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     7321 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     7381 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     7441 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     7501 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     7561 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     7621 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     7681 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     7741 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     7801 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     7861 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     7921 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     7981 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     8041 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     8101 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
     8161 gattacgcca agctctagct agaggtcgac caattctcat gtttgacagc ttatcatcgc
     8221 agatccgggc aacgttgttg ccattgctgc aggcgcagaa ctggtaggta tggaagatct
     8281 atacattgaa tcaatattgg caattagcca tattagtcat tggttatata gcataaatca
     8341 atattggcta ttggccattg catacgttgt atctatatca taatatgtac atttatattg
     8401 gctcatgtcc aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat
     8461 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
     8521 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
     8581 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
     8641 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg
     8701 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact
     8761 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     8821 ggcagtacac caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     8881 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     8941 gtaataaccc cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     9001 taagcagagc tcgtttagtg aaccgtcaga tcctcactct cttccgcatc gctgtctgcg
     9061 agggccagct gttgggctcg cggttgagga caaactcttc gcggtctttc cagtactctt
     9121 ggatcggaaa cccgtcggcc tccgaacggt actccgccac cgagggacct gagcgagtcc
     9181 gcatcgaccg gatcggaaaa cct
//
