LOCUS       sequence-309553-        6549 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 6549)
  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 6549)
  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 Orf9c
FEATURES             Location/Qualifiers
     source          1..6549
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(16..33)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(187..775)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(267..286)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(946..1737,1738..1806))
                     /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     1569..1588
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1807..1911)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1986..2005)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2177..2556
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2557..2760
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2711..2731
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2762..2780
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2783..2801
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2811..2835
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    2978..3002
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3228..3252)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3299..3322
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3359..3382
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3413..3439
                     /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             3413..3439
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(3440..3787,3788..3790,3791..4399)
                     /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(4443..4460)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4449..4673
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4806..4825)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4957..5004
                     /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             5012..6034
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             5012..6034
                     /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    6164..6285
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6201..6220)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6255..6274
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6334..6350)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6334..6350)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6347..6369)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6358..6374
                     /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(6382..6388,6389..6406,6407..6412))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6427..6448
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
       61 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
      121 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
      181 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
      241 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
      301 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
      361 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
      421 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
      481 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
      541 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
      601 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
      661 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
      721 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
      781 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
      841 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
      901 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
      961 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     1021 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     1081 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     1141 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     1201 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     1261 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     1321 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     1381 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     1441 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     1501 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     1561 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     1621 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     1681 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     1741 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     1801 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     1861 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     1921 ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact
     1981 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg
     2041 ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc
     2101 gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg
     2161 gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg
     2221 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
     2281 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
     2341 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
     2401 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
     2461 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
     2521 cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc
     2581 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
     2641 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
     2701 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct
     2761 ctccctatca gtgatagaga tctccctatc agtgatagag atcgtcgacg agctcgttta
     2821 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
     2881 cgggaccgat ccagcctccg gactctagcg tttaaactta agcttggtac cgagctcgga
     2941 tccactagtc cagtgtggtg gaattctgca gatatcaaca agtttgtaca aaaaagcagg
     3001 ctccaccatg cttcaatcct gctataactt cttgaaagag caacactgcc aaaaggcgag
     3061 cacacagaaa ggagccgagg cagccgtaaa gccgcttctt gtccctcatc acgttgtcgc
     3121 cacagtccaa gaaatacagc ttcaagcggc cgttggcgag cttctgctcc tcgaatggct
     3181 cgcaatggcg gtgatgttgt tgctgttgtg ctgctgtctt acggatgacc cagctttctt
     3241 gtacaaagtg gttgatatca aggccgccga catcaccagc ctgtacaaga aagtggggtg
     3301 gagccatccc cagttcgaga agggcggagg atctggcggc ggaagcggcg gaggaagctg
     3361 gtcccaccct cagtttgaga aaggcaccga gctgggcagc accatggcca gctaccccta
     3421 cgacgtgccc gactacgcca aggacaacac cgtgcccctg aagctgatcg ccctgctggc
     3481 caacggcgag ttccactctg gcgagcagct gggcgagaca ctgggcatga gcagagccgc
     3541 catcaacaag cacatccaga ccctgcggga ctggggcgtg gacgtgttca cagtgcccgg
     3601 caagggctac agcctgcccg agcccatcca gctgctgaac gccaagcaga tcctgggcca
     3661 gctggatggc ggatctgtgg ccgtgctgcc cgtgatcgac agcaccaacc agtatctgct
     3721 cgaccggatc ggcgagctga agtccggcga cgcctgtatc gccgagtacc agcaggccgg
     3781 cagaggcggc aggggcagaa agtggttcag ccccttcggc gccaacctgt acctgagcat
     3841 gttctggaga ctggaacagg gcccagccgc cgctattggc ctgagcctgg tcatcggcat
     3901 cgtgatggcc gaggtgctgc ggaagctggg cgccgacaaa gtgcgcgtga agtggcccaa
     3961 cgacctgtat ctgcaggacc ggaagctggc cggcatcctg gtggaactga ccggcaaaac
     4021 aggcgacgcc gcccagatcg tgatcggcgc tggcatcaac atggccatgc ggcgggtgga
     4081 agagagcgtg gtcaaccagg gctggatcac cctgcaggaa gccggcatca acctggaccg
     4141 gaacaccctg gccgccatgc tgatcagaga gctgagagcc gccctggaac tgttcgagca
     4201 ggaaggcctg gccccctacc tgagcagatg ggagaagctg gacaacttca tcaaccggcc
     4261 cgtgaaactg atcatcggcg acaaagagat cttcggcatc agccggggca tcgacaagca
     4321 gggggccctg ctcctggaac aggacggcat catcaagccc tggatgggcg gcgagatcag
     4381 cctgcggagc gccgagaagt gatgactcga gtctagaggg cccgtttaaa cccgctgatc
     4441 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     4501 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     4561 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     4621 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     4681 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     4741 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     4801 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     4861 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     4921 caaaaaactt gattagggtg atggttcacg tacctagaag ttcctattcc gaagttccta
     4981 ttctctagaa agtataggaa cttccttggc caaaaagcct gaactcaccg cgacgtctgt
     5041 cgagaagttt ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg
     5101 cgaagaatct cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa
     5161 tagctgcgcc gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc
     5221 gctcccgatt ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat
     5281 ctcccgccgt gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt
     5341 tctgcagccg gtcgcggagg ccatggatgc gatcgctgcg gccgatctta gccagacgag
     5401 cgggttcggc ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat
     5461 atgcgcgatt gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag
     5521 tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt
     5581 ccggcacctc gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat
     5641 aacagcggtc attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa
     5701 catcttcttc tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg
     5761 gaggcatccg gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct
     5821 tgaccaactc tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg
     5881 tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg
     5941 cagaagcgcg gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg
     6001 acgccccagc actcgtccga gggcaaagga atagcacgta ctacgagatt tcgattccac
     6061 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     6121 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     6181 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     6241 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     6301 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     6361 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     6421 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     6481 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     6541 gcgtattgg
//
