LOCUS       sequence-221989-        8663 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 8663)
  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 8663)
  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: GAP50-COMP-blac-flag-his
FEATURES             Location/Qualifiers
     source          1..8663
                     /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             2197..2982
                     /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             2989..3054
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3061..3078
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    3114..3169
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3168..3187)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5405..5423)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5491..5595
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5596..5664,5665..6456)
                     /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      6627..7215
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7116..7135
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7369..7386
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7503..7524
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7539..7544,7545..7562,7563..7569)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7577..7593
                     /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     7582..7604
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7601..7617
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7601..7617
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        7888..8267
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8268..8471
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8422..8442
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     8468..8492
                     /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 ggccgcccag
      421 ctgagatttg cctctctggg cgactggggc aaggacacca agggccagat cctgaacgcc
      481 aagtacttca agcagttcat caagaacgag cgcgtgacct tcatcgtgtc tcccggcagc
      541 aacttcatcg acggcgtgaa gggcctgaac gaccccgcct ggaagaacct gtacgaggac
      601 gtgtacagcg aagagaaggg cgatatgtac atgcctttct tcaccgtgct gggcacccgg
      661 gactggaccg gcaactacaa tgcccagctg ctgaagggcc agggcatcta catcgagaag
      721 aacggcgaga catctatcga gaaggacgcc gacgccacca actaccccaa gtggatcatg
      781 cccaactact ggtatcacta cttcacccac tttaccgtgt ccagcggccc cagcatcgtg
      841 aaaaccggcc acaaggacct ggccgctgcc ttcatcttca tcgatacctg ggtgctgagc
      901 agcaatttcc cctacaagaa gatccacgag aaggcctgga acgatctgaa gtcccagctg
      961 tccgtggcca agaagattgc cgacttcatc atcgtcgtgg gcgaccagcc catctacagc
     1021 agcggctaca gccggggcag cagctacctg gcctactacc tgctgcctct gctgaaagac
     1081 gccgaggtgg acctgtacat cagcggccac gacaacaaca tggaagtgat cgaggacaac
     1141 gacatggccc acatcacctg tggcagcggc agcatgagcc agggcaagag cggcatgaag
     1201 aacagcaaga gcctgttctt tagcagcgat atcggcttct gcgtgcacga gctgagcaac
     1261 aacggcatcg tgaccaagtt cgtgtcctcc aagaaaggcg aagtgatcta cacccacaag
     1321 ctgaacatca agaagaagaa aaccctggac aaagtgaacg ccctgcagca cttcgccgcc
     1381 ctgcccaacg tggaactgac cgacgtgcca agctccggcc ccggcgcgcc gtcgacctcc
     1441 atcacggcct ataagagtga gggggagtca gcggagttct ccttcccact caaccttgga
     1501 gaggaaagcc tgcagggaga gttgagatgg aaggcagaga aggctccttc ttcccagtcc
     1561 tggatcacct tctccctaaa gaaccaaaag gtgtctgtgc agaagtctac tagcaacccc
     1621 aagttccagc tgtccgaaac gctcccactc acccttcaga taccccaggt ctcccttcag
     1681 tttgctggtt ctggcaacct gaccctgact ctggacagag ggatactgta tcaggaagtg
     1741 aacctggtgg tgatgaaagt gactcagccc gacagcaaca ctttgacctg tgaggtgatg
     1801 ggacccacct cacccaagat gagactgatc ttgaagcagg agaatcagga ggccagggtc
     1861 tccaggcagg agaaagtgat tcaagtgcag gcccctgaag caggggtgtg gcaatgtcta
     1921 ctgagtgaag gtgaagaggt caagatggac tccaagatcc aggttttatc caaagggttg
     1981 aattccggcg gcggctccgg aggcggtacc ggtggcggcg acctagcccc acagatgctt
     2041 cgagaactcc aggagactaa tgcggcgctg caagacgtga gagagctctt gcgacagcag
     2101 gtcaaggaga tcaccttcct gaagaatacg gtgatggaat gtgacgcctg cggtctagac
     2161 agaaacctgc ctccactagc acctctgggc ccacacccag aaacgctggt gaaagtaaaa
     2221 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     2281 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     2341 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     2401 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     2461 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     2521 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     2581 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     2641 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     2701 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     2761 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     2821 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     2881 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     2941 agacagatcg ctgagatagg tgcctcactg attaagcatt ggggccctga ctacaaagac
     3001 catgacggtg attataaaga tcatgacatc gattacaagg atgacgatga caagggccct
     3061 caccaccacc atcaccacta ataataagga tcccccgacc tcgacctctg gctaataaag
     3121 gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac
     3181 atatgggagg gcaaatcatt tggtcgagat ccctcggaga tctctagcta gaggatcgat
     3241 ccccgccccg gacgaactaa acctgactac gacatctctg ccccttcttc gcggggcagt
     3301 gcatgtaatc ccttcagttg gttggtacaa cttgccaact gggccctgtt ccacatgtga
     3361 cacggggggg gaccaaacac aaaggggttc tctgactgta gttgacatcc ttataaatgg
     3421 atgtgcacat ttgccaacac tgagtggctt tcatcctgga gcagactttg cagtctgtgg
     3481 actgcaacac aacattgcct ttatgtgtaa ctcttggctg aagctcttac accaatgctg
     3541 ggggacatgt acctcccagg ggcccaggaa gactacggga ggctacacca acgtcaatca
     3601 gaggggcctg tgtagctacc gataagcgga ccctcaagag ggcattagca atagtgttta
     3661 taaggccccc ttgttaaccc taaacgggta gcatatgctt cccgggtagt agtatatact
     3721 atccagacta accctaattc aatagcatat gttacccaac gggaagcata tgctatcgaa
     3781 ttagggttag taaaagggtc ctaaggaaca gcgatatctc ccaccccatg agctgtcacg
     3841 gttttattta catggggtca ggattccacg agggtagtga accattttag tcacaagggc
     3901 agtggctgaa gatcaaggag cgggcagtga actctcctga atcttcgcct gcttcttcat
     3961 tctccttcgt ttagctaata gaataactgc tgagttgtga acagtaaggt gtatgtgagg
     4021 tgctcgaaaa caaggtttca ggtgacgccc ccagaataaa atttggacgg ggggttcagt
     4081 ggtggcattg tgctatgaca ccaatataac cctcacaaac cccttgggca ataaatacta
     4141 gtgtaggaat gaaacattct gaatatcttt aacaatagaa atccatgggg tggggacaag
     4201 ccgtaaagac tggatgtcca tctcacacga atttatggct atgggcaaca cataatccta
     4261 gtgcaatatg atactggggt tattaagatg tgtcccaggc agggaccaag acaggtgaac
     4321 catgttgtta cactctattt gtaacaaggg gaaagagagt ggacgccgac agcagcggac
     4381 tccactggtt gtctctaaca cccccgaaaa ttaaacgggg ctccacgcca atggggccca
     4441 taaacaaaga caagtggcca ctcttttttt tgaaattgtg gagtgggggc acgcgtcagc
     4501 ccccacacgc cgccctgcgg ttttggactg taaaataagg gtgtaataac ttggctgatt
     4561 gtaaccccgc taaccactgc ggtcaaacca cttgcccaca aaaccactaa tggcaccccg
     4621 gggaatacct gcataagtag gtgggcgggc caagataggg gcgcgattgc tgcgatctgg
     4681 aggacaaatt acacacactt gcgcctgagc gccaagcaca gggttgttgg tcctcatatt
     4741 cacgaggtcg ctgagagcac ggtgggctaa tgttgccatg ggtagcatat actacccaaa
     4801 tatctggata gcatatgcta tcctaatcta tatctgggta gcataggcta tcctaatcta
     4861 tatctgggta gcatatgcta tcctaatcta tatctgggta gtatatgcta tcctaattta
     4921 tatctgggta gcataggcta tcctaatcta tatctgggta gcatatgcta tcctaatcta
     4981 tatctgggta gtatatgcta tcctaatctg tatccgggta gcatatgcta tcctaataga
     5041 gattagggta gtatatgcta tcctaattta tatctgggta gcatatacta cccaaatatc
     5101 tggatagcat atgctatcct aatctatatc tgggtagcat atgctatcct aatctatatc
     5161 tgggtagcat aggctatcct aatctatatc tgggtagcat atgctatcct aatctatatc
     5221 tgggtagtat atgctatcct aatttatatc tgggtagcat aggctatcct aatctatatc
     5281 tgggtagcat atgctatcct aatctatatc tgggtagtat atgctatcct aatctgtatc
     5341 cgggtagcat atgctatcct catgataagc tgtcaaacat gagaattttc ttgaagacga
     5401 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     5461 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     5521 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     5581 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     5641 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5701 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5761 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5821 ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg catacactat
     5881 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5941 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     6001 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     6061 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     6121 cgtgacacca cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa
     6181 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     6241 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     6301 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     6361 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     6421 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     6481 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     6541 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     6601 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     6661 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     6721 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     6781 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     6841 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     6901 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     6961 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     7021 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     7081 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     7141 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     7201 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     7261 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     7321 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     7381 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     7441 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     7501 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     7561 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
     7621 gattacgcca agctctagct agaggtcgac caattctcat gtttgacagc ttatcatcgc
     7681 agatccgggc aacgttgttg ccattgctgc aggcgcagaa ctggtaggta tggaagatct
     7741 atacattgaa tcaatattgg caattagcca tattagtcat tggttatata gcataaatca
     7801 atattggcta ttggccattg catacgttgt atctatatca taatatgtac atttatattg
     7861 gctcatgtcc aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat
     7921 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
     7981 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
     8041 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
     8101 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg
     8161 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact
     8221 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     8281 ggcagtacac caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     8341 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     8401 gtaataaccc cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     8461 taagcagagc tcgtttagtg aaccgtcaga tcctcactct cttccgcatc gctgtctgcg
     8521 agggccagct gttgggctcg cggttgagga caaactcttc gcggtctttc cagtactctt
     8581 ggatcggaaa cccgtcggcc tccgaacggt actccgccac cgagggacct gagcgagtcc
     8641 gcatcgaccg gatcggaaaa cct
//
