LOCUS       sequence-327668-        8453 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human expression vector encoding full length SARS CoV-2 spike 
            protein, containing biotinylation site and 8His-tag. Modification of
            Addgene plasmid #154754..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8453)
  AUTHORS   Reyes RA, Clarke K, Gonzales SJ, Cantwell AM, Garza R, Catano G, 
            Tragus RE, Patterson TF, Bol S, Bunnik EM
  TITLE     SARS-CoV-2 spike-specific memory B cells express higher levels of 
            T-bet and FcRL5 after non-severe COVID-19 as compared to severe 
            disease.
  JOURNAL   PLoS One. 2021 Dec 22;16(12):e0261656. doi: 
            10.1371/journal.pone.0261656. eCollection 2021.
  PUBMED    34936684
REFERENCE   2  (bases 1 to 8453)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS One.";
            date: "2021-12-22"; volume: "16(12):e0261656. doi"; pages: " 
            10.1371/journal.pone.0261656. eCollection 2021"
COMMENT     Sequence Label: SARS CoV-2 HexaPro spike (bio-His)
FEATURES             Location/Qualifiers
     source          1..8453
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        28..407
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          807..1823
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1747..1769
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     CDS             5706..5729
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(5831..5850)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5896..5951
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5950..5969)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(6312..6328)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6312..6328)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6325..6347)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6336..6352
                     /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)."
     promoter        complement(join(6360..6366,6367..6384,6385..6390))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6405..6426
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6555..6572)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6726..7314)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6806..6825)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7485..8276,8277..8345))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8108..8127
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8346..8450)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atttccccga aaagtgccac ctgggtcgac attgattatt gactagttat taatagtaat
       61 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      121 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      181 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      241 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      301 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
      361 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc gaggtgagcc
      421 ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat
      481 ttatttttta attattttgt gcagcgatgg gggcgggggg gggggggggg sgcgccsgnn
      541 gggggggggg gggggggggg gggggggggg gsggggggng gggggggggg gggkkggkgg
      601 gskkggkrgg kggggggggg gggggggggg gggggggggg gggggggggc gcgcggnggg
      661 gggggggggg ggggggcgag gggcggggcg gggcgaggcg gagaggtgcg gcggcagcca
      721 atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg cggcggccct
      781 ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc
      841 gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg
      901 tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt taatgacggc
      961 ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg
     1021 gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt ggggagcgcc gcgtgcggct
     1081 ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt gcgctccgca
     1141 gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg ggctgcgagg
     1201 ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg agcagggggt gtgggcgcgt
     1261 cggtcgggct gcaacccccc ctgcaccccc ctccccgagt tgctgagcac ggcccggctt
     1321 cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc
     1381 ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggagggc tcgggggagg
     1441 ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc ggcgagccgc agccattgcc
     1501 ttttatggta atcgtgcgag agggcgcagg gacttccttt gtcccaaatc tgtgcggagc
     1561 cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg
     1621 ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc
     1681 tccctctcca gcctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca
     1741 gggcggggtt cggcttctgg cgtgtgaccg gcggctctag cgcctctgct aaccatgttc
     1801 atgccttctt ctttttccta cagctcctgg gcaacgtgct ggttgttgtg ctgtctcatc
     1861 attttggcaa agaattgcgg ccgtctcagg ccgagttcgg taccgccacc atgttcgtgt
     1921 tcctggtgct cctgcctctg gtgagcagcc agtgcgtgaa cctgaccacc cgaacccagc
     1981 tcccaccagc ctacaccaac agctttacac ggggcgtgta ctaccctgac aaggtgttca
     2041 gatctagcgt cctgcacagc actcaggacc tcttcctgcc gttcttcagc aacgtgacat
     2101 ggttccacgc catccacgtg agcggcacaa acggaaccaa gcggtttgat aaccccgtcc
     2161 tgccattcaa tgatggagtt tacttcgcca gtaccgagaa gagtaacatc atccggggct
     2221 ggatcttcgg caccaccctg gatagcaaaa cacagagcct cctgatcgtg aacaatgcca
     2281 cgaacgtcgt gatcaaggtg tgcgagttcc agttttgcaa tgatcctttc ctgggtgtgt
     2341 actaccacaa gaacaacaag agctggatgg aaagcgagtt cagagtctac agcagcgcca
     2401 acaactgcac attcgagtac gtctctcagc cttttctgat ggaccttgag gggaaacaag
     2461 gcaacttcaa gaacctgaga gaattcgtgt tcaagaacat cgacggctac ttcaaaatct
     2521 actccaagca cacacccatc aacctggtcc gggacctccc tcagggcttc agcgccctgg
     2581 aacccctggt cgacctgccc ataggcatca acataacgcg gttccaaacc ctgctggccc
     2641 tgcatagatc ctacctgact cctggcgaca gcagcagcgg atggaccgcc ggagctgcag
     2701 cctactatgt gggctacctg caacctagaa ccttcctgct gaagtacaac gagaacggca
     2761 caatcacaga cgccgtcgac tgcgccctgg accctctctc tgagacaaag tgcaccctga
     2821 agtccttcac cgtggaaaag ggcatctacc agaccagcaa cttccgggtg cagcctacag
     2881 agagcatcgt gcgatttcca aacattacca acctctgccc cttcggcgag gtgtttaacg
     2941 ccacaagatt tgcctccgtt tacgcctgga atagaaagag aatcagcaat tgtgtggccg
     3001 actactccgt gctgtataac agcgcctctt tcagcacctt caagtgctac ggcgtttccc
     3061 caacaaagct gaatgacctg tgcttcacca acgtgtacgc cgactccttc gtaattagag
     3121 gcgatgaggt gcggcagatc gcaccaggcc agaccggtaa gatcgctgac tacaactata
     3181 agctgcctga tgattttaca ggctgcgtga tcgcctggaa ctctaacaac ctggatagca
     3241 aggtgggcgg caactacaac tacctgtacc ggctgtttcg caagtctaac ctgaaacctt
     3301 tcgagagaga catctccaca gagatctacc aggccggttc tacaccttgt aacggggtgg
     3361 aaggcttcaa ctgttacttc cctctgcaaa gctacggctt ccagcctacc aatggagtcg
     3421 gctaccagcc ataccgggtg gtcgtgctgt ccttcgagtt actccacgcc cccgccaccg
     3481 tctgcggtcc taagaagtcc accaatctgg ttaagaacaa atgcgtgaac ttcaacttca
     3541 acggcctgac cgggaccggc gtgctgaccg aaagcaacaa aaagttcctc cccttccagc
     3601 agttcggccg tgatatcgct gacaccacag atgccgtcag agatccacag accctggaaa
     3661 tcctggatat tacaccctgc tccttcggag gagtttctgt gatcaccccc gggaccaata
     3721 ccagcaacca ggtggctgtg ctgtaccaag atgttaactg caccgaggtt cctgtggcca
     3781 tccacgccga tcagctgaca cctacttgga gagtgtactc cactggctcc aatgtgttcc
     3841 agaccagggc cggatgtctg atcggcgccg agcacgtgaa taacagttac gagtgcgaca
     3901 tccctatcgg cgccggcatc tgtgccagct accagaccca gacaaacagc cctgggtctg
     3961 cttcctctgt agctagccag agcatcatcg cctacaccat gagcctgggc gcagagaaca
     4021 gcgtggccta ttccaacaac tctatcgcca ttcccaccaa ctttacaatt agcgtcacaa
     4081 cagagatcct gcccgtgagc atgaccaaga ccagcgtgga ctgtacaatg tacatctgtg
     4141 gcgacagcac tgaatgcagc aacctgctgc tgcaatacgg ctccttttgc acccaactga
     4201 accgggcgct gaccggaatc gccgtggaac aggacaaaaa tacccaggag gtgttcgccc
     4261 aagtgaagca gatctacaag accccaccta tcaaggactt cggcggcttt aactttagcc
     4321 agattctccc tgatccttct aagcctagca agcggagccc tatcgaggat ctgctgttca
     4381 acaaggtcac cctggccgat gccggcttta tcaaacagta tggcgattgc ctgggcgaca
     4441 tagccgccag agatctgatc tgcgcccaga aattcaacgg cctgacagtt ctcccacctc
     4501 tgctgaccga cgagatgatc gctcagtaca cctctgccct gctggctggc accatcacat
     4561 ctgggtggac atttggcgcc ggccccgccc tgcagatccc ctttcccatg cagatggcct
     4621 atagattcaa cggaatcggc gtgacccaga acgtgctgta tgaaaaccag aagctgatcg
     4681 ctaaccagtt caattctgcc atcggcaaga tccaggactc cctctcctct acccccagcg
     4741 ccctgggcaa actgcaggac gtggtgaatc agaacgccca agccctgaac accctggtga
     4801 agcagctcag cagcaatttt ggcgccatca gctctgtgct gaacgatatc ctgtctagac
     4861 tggaccctcc agaagccgaa gtccagatcg atagactgat cacaggcaga ctgcagtccc
     4921 tgcaaaccta cgtgacccaa cagctgatca gggccgctga aataagagcc agcgccaatc
     4981 tcgccgctac caagatgtcc gagtgtgtgc tgggacagtc taaacgcgtt gacttctgcg
     5041 gcaaaggcta tcacctgatg agcttccccc agagcgcgcc gcacggcgtg gtgttcctgc
     5101 atgtgacata cgtgcctgcc caagagaaga atttcacaac cgcccctgcc atctgccacg
     5161 acggcaaggc ccacttccca agagagggcg ttttcgtttc caatggcaca cactggttcg
     5221 tgacacaaag aaacttctac gaaccccaga ttatcaccac cgacaacacc ttcgtgagtg
     5281 gcaattgtga cgtggtcatc ggaatcgtga acaacacagt gtacgaccct ctgcaacctg
     5341 agctggactc ttttaaggaa gagctggaca agtactttaa aaaccacacc agccccgatg
     5401 tggacctggg cgacatcagt ggcattaacg ccagcgtggt gaacatccaa aaggaaatcg
     5461 acagactgaa cgaggtggcc aagaacctga acgagtccct gatcgacctg caggagctcg
     5521 gcaaatacga gcagggatcc ggatacatcc ccgaggcccc cagagatggc caggcctacg
     5581 tgcggaagga cggcgagtgg gtactgctga gcacattcct gggcagatcc ggaggcggtg
     5641 gttccctgca tcatattctg gatgcacaga aaatggtgtg gaatcatcgt ggcggtggag
     5701 gttcccatca ccaccatcac caccatcatt gataatgact cgagcgataa ttcactcctc
     5761 aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc caatgccctg gctcacaaat
     5821 accactgaga tctttttccc tctgccaaaa attatgggga catcatgaag ccccttgagc
     5881 atctgacttc tggctaataa aggaaattta ttttcattgc aatagtgtgt tggaattttt
     5941 tgtgtctctc actcggaagg acatatggga gggcaaatca tttaaaacat cagaatgagt
     6001 atttggttta gagtttggca acatatgccc atatgctggc tgccatgaac aaaggttggc
     6061 tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta ttccatagaa
     6121 aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt ttttctttaa
     6181 catccctaaa attttcctta catgttttac tagccagatt tttcctcctc tcctgactac
     6241 tcccagtcat agctgtccct cttctcttat ggagatccct cgacctgcag cccaagcttg
     6301 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     6361 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     6421 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagcgg
     6481 atcgatccgc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     6541 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     6601 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     6661 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6721 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6781 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     6841 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     6901 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6961 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     7021 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     7081 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     7141 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     7201 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     7261 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     7321 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     7381 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     7441 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     7501 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     7561 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7621 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7681 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7741 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7801 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     7861 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     7921 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7981 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     8041 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     8101 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     8161 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     8221 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     8281 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     8341 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     8401 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cac
//
