LOCUS       sequence-309533-        6444 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 6444)
  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 6444)
  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 Orf10
FEATURES             Location/Qualifiers
     source          1..6444
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(23..40)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(194..782)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(274..293)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(953..1744,1745..1813))
                     /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     1576..1595
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1814..1918)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1993..2012)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2184..2563
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2564..2767
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2718..2738
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2769..2787
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2790..2808
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2818..2842
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    2985..3009
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3130..3154)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3201..3224
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3261..3284
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3315..3341
                     /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             3315..3341
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(3342..3689,3690..3692,3693..4301)
                     /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(4345..4362)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4351..4575
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4708..4727)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4859..4906
                     /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             4914..5936
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4914..5936
                     /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    6066..6187
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6103..6122)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6157..6176
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6236..6252)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6236..6252)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6249..6271)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6260..6276
                     /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(6284..6290,6291..6308,6309..6314))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6329..6350
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
       61 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      121 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      181 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      241 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      301 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      361 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      421 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
      481 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      541 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      601 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
      661 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      721 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
      781 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
      841 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
      901 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
      961 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1021 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     1081 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     1141 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1201 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1261 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1321 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1381 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1441 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1501 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1561 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1621 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1681 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1741 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1801 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1861 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     1921 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     1981 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     2041 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
     2101 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
     2161 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
     2221 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     2281 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     2341 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     2401 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     2461 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     2521 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
     2581 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     2641 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     2701 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     2761 cagagctctc cctatcagtg atagagatct ccctatcagt gatagagatc gtcgacgagc
     2821 tcgtttagtg aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag
     2881 aagacaccgg gaccgatcca gcctccggac tctagcgttt aaacttaagc ttggtaccga
     2941 gctcggatcc actagtccag tgtggtggaa ttctgcagat atcaacaagt ttgtacaaaa
     3001 aagcaggctc caccatgggc tacatcaatg tcttcgcctt tcctttcacc atctattctc
     3061 tcttgttgtg tcgcatgaac tctaggaatt acatagctca ggtcgatgtt gtaaacttca
     3121 atctcacaga cccagctttc ttgtacaaag tggttgatat caaggccgcc gacatcacca
     3181 gcctgtacaa gaaagtgggg tggagccatc cccagttcga gaagggcgga ggatctggcg
     3241 gcggaagcgg cggaggaagc tggtcccacc ctcagtttga gaaaggcacc gagctgggca
     3301 gcaccatggc cagctacccc tacgacgtgc ccgactacgc caaggacaac accgtgcccc
     3361 tgaagctgat cgccctgctg gccaacggcg agttccactc tggcgagcag ctgggcgaga
     3421 cactgggcat gagcagagcc gccatcaaca agcacatcca gaccctgcgg gactggggcg
     3481 tggacgtgtt cacagtgccc ggcaagggct acagcctgcc cgagcccatc cagctgctga
     3541 acgccaagca gatcctgggc cagctggatg gcggatctgt ggccgtgctg cccgtgatcg
     3601 acagcaccaa ccagtatctg ctcgaccgga tcggcgagct gaagtccggc gacgcctgta
     3661 tcgccgagta ccagcaggcc ggcagaggcg gcaggggcag aaagtggttc agccccttcg
     3721 gcgccaacct gtacctgagc atgttctgga gactggaaca gggcccagcc gccgctattg
     3781 gcctgagcct ggtcatcggc atcgtgatgg ccgaggtgct gcggaagctg ggcgccgaca
     3841 aagtgcgcgt gaagtggccc aacgacctgt atctgcagga ccggaagctg gccggcatcc
     3901 tggtggaact gaccggcaaa acaggcgacg ccgcccagat cgtgatcggc gctggcatca
     3961 acatggccat gcggcgggtg gaagagagcg tggtcaacca gggctggatc accctgcagg
     4021 aagccggcat caacctggac cggaacaccc tggccgccat gctgatcaga gagctgagag
     4081 ccgccctgga actgttcgag caggaaggcc tggcccccta cctgagcaga tgggagaagc
     4141 tggacaactt catcaaccgg cccgtgaaac tgatcatcgg cgacaaagag atcttcggca
     4201 tcagccgggg catcgacaag cagggggccc tgctcctgga acaggacggc atcatcaagc
     4261 cctggatggg cggcgagatc agcctgcgga gcgccgagaa gtgatgactc gagtctagag
     4321 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     4381 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     4441 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     4501 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     4561 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     4621 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     4681 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     4741 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     4801 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtacctaga
     4861 agttcctatt ccgaagttcc tattctctag aaagtatagg aacttccttg gccaaaaagc
     4921 ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg
     4981 acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc
     5041 gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt
     5101 atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt ggggaattca
     5161 gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc
     5221 ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat gcgatcgctg
     5281 cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat
     5341 acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa
     5401 ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt
     5461 gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg
     5521 tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg
     5581 attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc
     5641 agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg cggctccggg
     5701 cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg
     5761 atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg
     5821 tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag
     5881 tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaatagcacg
     5941 tactacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     6001 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     6061 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     6121 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     6181 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     6241 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     6301 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     6361 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     6421 aacgcgcggg gagaggcggt ttgc
//
