LOCUS       sequence-309543-        6513 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MAC-tagged gene expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6513)
  AUTHORS   Liu X, Salokas K, Weldatsadik RG, Gawriyski L, Varjosalo M
  TITLE     Combined proximity labeling and affinity purification-mass 
            spectrometry workflow for mapping and visualizing protein 
            interaction networks.
  JOURNAL   Nat Protoc. 2020 Aug 10. pii: 10.1038/s41596-020-0365-x. doi: 
            10.1038/s41596-020-0365-x.
  PUBMED    32778839
REFERENCE   2  (bases 1 to 6513)
  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: "Nat Protoc.
            2020 Aug 10. pii: 10.1038/s41596-020-0365-x. doi: 
            10.1038/s41596-020-0365-x."
COMMENT     Sequence Label: MAC-SARS-CoV-2 Orf6
FEATURES             Location/Qualifiers
     source          1..6513
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    206..224
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    227..245
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     255..279
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    422..446
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(636..660)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             707..730
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             767..790
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             821..847
                     /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             821..847
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(848..1195,1196..1198,1199..1807)
                     /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(1851..1868)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1857..2081
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2214..2233)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2365..2412
                     /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             2420..3442
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2420..3442
                     /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    3572..3693
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3609..3628)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3663..3682
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3742..3758)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3742..3758)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3755..3777)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3766..3782
                     /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(3790..3796,3797..3814,3815..3820))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3835..3856
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3973..3990)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4144..4732)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4224..4243)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4903..5694,5695..5763))
                     /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     5526..5545
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5764..5868)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5943..5962)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6134..6513
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctctccct atcagtgata gagatctccc tatcagtgat
      241 agagatcgtc gacgagctcg tttagtgaac cgtcagatcg cctggagacg ccatccacgc
      301 tgttttgacc tccatagaag acaccgggac cgatccagcc tccggactct agcgtttaaa
      361 cttaagcttg gtaccgagct cggatccact agtccagtgt ggtggaattc tgcagatatc
      421 aacaagtttg tacaaaaaag caggctccac catgtttcat cttgtcgatt ttcaagttac
      481 gattgcggag atactcctca taatcatgcg gacatttaag gtttccattt ggaaccttga
      541 ttacattatt aacctgatta tcaagaatct ctccaagagc ctgaccgaga ataagtattc
      601 tcagttggac gaggagcagc ctatggaaat agacgaccca gctttcttgt acaaagtggt
      661 tgatatcaag gccgccgaca tcaccagcct gtacaagaaa gtggggtgga gccatcccca
      721 gttcgagaag ggcggaggat ctggcggcgg aagcggcgga ggaagctggt cccaccctca
      781 gtttgagaaa ggcaccgagc tgggcagcac catggccagc tacccctacg acgtgcccga
      841 ctacgccaag gacaacaccg tgcccctgaa gctgatcgcc ctgctggcca acggcgagtt
      901 ccactctggc gagcagctgg gcgagacact gggcatgagc agagccgcca tcaacaagca
      961 catccagacc ctgcgggact ggggcgtgga cgtgttcaca gtgcccggca agggctacag
     1021 cctgcccgag cccatccagc tgctgaacgc caagcagatc ctgggccagc tggatggcgg
     1081 atctgtggcc gtgctgcccg tgatcgacag caccaaccag tatctgctcg accggatcgg
     1141 cgagctgaag tccggcgacg cctgtatcgc cgagtaccag caggccggca gaggcggcag
     1201 gggcagaaag tggttcagcc ccttcggcgc caacctgtac ctgagcatgt tctggagact
     1261 ggaacagggc ccagccgccg ctattggcct gagcctggtc atcggcatcg tgatggccga
     1321 ggtgctgcgg aagctgggcg ccgacaaagt gcgcgtgaag tggcccaacg acctgtatct
     1381 gcaggaccgg aagctggccg gcatcctggt ggaactgacc ggcaaaacag gcgacgccgc
     1441 ccagatcgtg atcggcgctg gcatcaacat ggccatgcgg cgggtggaag agagcgtggt
     1501 caaccagggc tggatcaccc tgcaggaagc cggcatcaac ctggaccgga acaccctggc
     1561 cgccatgctg atcagagagc tgagagccgc cctggaactg ttcgagcagg aaggcctggc
     1621 cccctacctg agcagatggg agaagctgga caacttcatc aaccggcccg tgaaactgat
     1681 catcggcgac aaagagatct tcggcatcag ccggggcatc gacaagcagg gggccctgct
     1741 cctggaacag gacggcatca tcaagccctg gatgggcggc gagatcagcc tgcggagcgc
     1801 cgagaagtga tgactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     1861 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     1921 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     1981 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2041 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2101 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2161 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2221 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2281 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2341 ttagggtgat ggttcacgta cctagaagtt cctattccga agttcctatt ctctagaaag
     2401 tataggaact tccttggcca aaaagcctga actcaccgcg acgtctgtcg agaagtttct
     2461 gatcgaaaag ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg
     2521 tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga
     2581 tggtttctac aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc
     2641 ggaagtgctt gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc
     2701 acagggtgtc acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt
     2761 cgcggaggcc atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc
     2821 attcggaccg caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc
     2881 tgatccccat gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc
     2941 gcaggctctc gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt
     3001 gcacgcggat ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat
     3061 tgactggagc gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg
     3121 gaggccgtgg ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga
     3181 gcttgcagga tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta
     3241 tcagagcttg gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc
     3301 aatcgtccga tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc
     3361 cgtctggacc gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac
     3421 tcgtccgagg gcaaaggaat agcacgtact acgagatttc gattccaccg ccgccttcta
     3481 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     3541 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     3601 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     3661 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     3721 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     3781 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     3841 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     3901 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     3961 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4021 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4081 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4141 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4201 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4261 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4321 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4381 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4441 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4501 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4561 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4621 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4681 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4741 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4801 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4861 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4921 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     4981 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5041 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5101 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5161 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5221 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5281 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5341 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5401 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5461 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5521 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5581 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5641 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     5701 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     5761 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     5821 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     5881 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc
     5941 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     6001 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     6061 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
     6121 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
     6181 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     6241 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     6301 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     6361 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     6421 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     6481 tacatctacg tattagtcat cgctattacc atg
//
