LOCUS       sequence-383904-        6693 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Sars-CoV-2 ORF8_R52I in MAC-C destination vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6693)
  AUTHORS   Varjosalo M, Huuskonen S, Liu X, Redchuk T, Salokas K, Lundberg R, 
            Maljanen S, Belik M, Reinholm A, Kolehmainen P, Tripathi G, Laine P,
            Belanov S, Auvinen P, Vartiainen M, Laine L, Osterlund P, Julkkunen 
            I, Kakkola L
  TITLE     The comprehensive SARS-CoV-2 'hijackome' knowledge base— reveals 
            significant changes in host cell protein expression and activation 
            by multiple SARS-CoV-2 variants
  JOURNAL   10 May 2024, Research Square
REFERENCE   2  (bases 1 to 6693)
  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: "10 May 
            2024, Research Square"
COMMENT     Sequence Label: MAC-C Sars-CoV-2 ORF8_R52I
FEATURES             Location/Qualifiers
     source          1..6693
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(243..262)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    394..441
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             449..1471
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             449..1471
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    1601..1722
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1638..1657)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1692..1711
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1771..1787)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1771..1787)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1784..1806)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1795..1811
                     /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(1819..1825,1826..1843,1844..1849))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1864..1885
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2002..2019)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2173..2761)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2253..2272)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2932..3723,3724..3792))
                     /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     3555..3574
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3793..3897)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3972..3991)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4163..4542
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4543..4746
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4697..4717
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    4748..4766
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4769..4787
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     4797..4821
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    4964..4988
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(5358..5382)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             5429..5452
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             5489..5512
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             5543..5569
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             5543..5569
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(5570..5917,5918..5920,5921..6529)
                     /codon_start=1
                     /product="promiscuous R118G mutant of the E. coli biotin 
                     protein ligase (Cronan, 2005)"
                     /label=BirA(R118G)
                     /note="also known as BirA* or BioID"
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     primer_bind     complement(6573..6590)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    join(6579..6693,1..110)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
       61 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
      121 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
      181 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
      241 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
      301 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
      361 aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa gttcctattc
      421 tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga
      481 gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga
      541 agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag
      601 ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct
      661 cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc
      721 ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct
      781 gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg
      841 gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg
      901 cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc
      961 gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg
     1021 gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac
     1081 agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat
     1141 cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag
     1201 gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga
     1261 ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg
     1321 atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag
     1381 aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg
     1441 ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg attccaccgc
     1501 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     1561 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     1621 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     1681 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     1741 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     1801 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     1861 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     1921 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     1981 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     2041 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     2101 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     2161 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     2221 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     2281 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     2341 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     2401 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     2461 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     2521 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     2581 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     2641 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     2701 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     2761 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     2821 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     2881 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     2941 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     3001 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     3061 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     3121 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     3181 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     3241 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     3301 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     3361 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     3421 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     3481 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     3541 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     3601 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     3661 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     3721 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     3781 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     3841 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     3901 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
     3961 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     4021 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     4081 ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
     4141 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     4201 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     4261 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     4321 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     4381 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     4441 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     4501 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     4561 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     4621 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     4681 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     4741 agagctctcc ctatcagtga tagagatctc cctatcagtg atagagatcg tcgacgagct
     4801 cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga
     4861 agacaccggg accgatccag cctccggact ctagcgttta aacttaagct tggtaccgag
     4921 ctcggatcca ctagtccagt gtggtggaat tctgcagata tcaacaagtt tgtacaaaaa
     4981 agcaggctcc accatgaagt ttctcgtgtt cttgggcatt atcaccacag tcgcggcttt
     5041 tcatcaagaa tgctctttgc agtcttgtac acagcatcaa ccttatgtcg tcgatgatcc
     5101 ttgcccgatt cacttttaca gtaaatggta tatcagggta ggggccataa aaagcgcacc
     5161 attgattgaa ttgtgtgtgg acgaggcagg gagcaaaagt ccgattcaat acattgacat
     5221 cggtaattat actgtcagtt gttccccctt tacgatcaac tgccaagaac ccaaactggg
     5281 gagcttggtg gtccgatgtt ccttctatga agattttctt gagtatcacg atgtcagggt
     5341 ggtccttgac tttatagacc cagctttctt gtacaaagtg gttgatatca aggccgccga
     5401 catcaccagc ctgtacaaga aagtggggtg gagccatccc cagttcgaga agggcggagg
     5461 atctggcggc ggaagcggcg gaggaagctg gtcccaccct cagtttgaga aaggcaccga
     5521 gctgggcagc accatggcca gctaccccta cgacgtgccc gactacgcca aggacaacac
     5581 cgtgcccctg aagctgatcg ccctgctggc caacggcgag ttccactctg gcgagcagct
     5641 gggcgagaca ctgggcatga gcagagccgc catcaacaag cacatccaga ccctgcggga
     5701 ctggggcgtg gacgtgttca cagtgcccgg caagggctac agcctgcccg agcccatcca
     5761 gctgctgaac gccaagcaga tcctgggcca gctggatggc ggatctgtgg ccgtgctgcc
     5821 cgtgatcgac agcaccaacc agtatctgct cgaccggatc ggcgagctga agtccggcga
     5881 cgcctgtatc gccgagtacc agcaggccgg cagaggcggc aggggcagaa agtggttcag
     5941 ccccttcggc gccaacctgt acctgagcat gttctggaga ctggaacagg gcccagccgc
     6001 cgctattggc ctgagcctgg tcatcggcat cgtgatggcc gaggtgctgc ggaagctggg
     6061 cgccgacaaa gtgcgcgtga agtggcccaa cgacctgtat ctgcaggacc ggaagctggc
     6121 cggcatcctg gtggaactga ccggcaaaac aggcgacgcc gcccagatcg tgatcggcgc
     6181 tggcatcaac atggccatgc ggcgggtgga agagagcgtg gtcaaccagg gctggatcac
     6241 cctgcaggaa gccggcatca acctggaccg gaacaccctg gccgccatgc tgatcagaga
     6301 gctgagagcc gccctggaac tgttcgagca ggaaggcctg gccccctacc tgagcagatg
     6361 ggagaagctg gacaacttca tcaaccggcc cgtgaaactg atcatcggcg acaaagagat
     6421 cttcggcatc agccggggca tcgacaagca gggggccctg ctcctggaac aggacggcat
     6481 catcaagccc tggatgggcg gcgagatcag cctgcggagc gccgagaagt gatgactcga
     6541 gtctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     6601 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     6661 cctttcctaa taaaatgagg aaattgcatc gca
//
