LOCUS       sequence-309559-        6509 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 6509)
  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 6509)
  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 Orf7b
FEATURES             Location/Qualifiers
     source          1..6509
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        204..407
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     358..378
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    409..427
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    430..448
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     458..482
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    625..649
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(835..859)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             906..929
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             966..989
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1020..1046
                     /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             1020..1046
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(1047..1394,1395..1397,1398..2006)
                     /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(2050..2067)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2056..2280
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2413..2432)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2564..2611
                     /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             2619..3641
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2619..3641
                     /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    3771..3892
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3808..3827)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3862..3881
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3941..3957)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3941..3957)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3954..3976)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3965..3981
                     /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(3989..3995,3996..4013,4014..4019))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4034..4055
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4172..4189)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4343..4931)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4423..4442)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5102..5893,5894..5962))
                     /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     5725..5744
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5963..6067)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6142..6161)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(6333..6509,1..203)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
       61 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      121 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      181 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
      241 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
      301 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
      361 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc cctatcagtg
      421 atagagatct ccctatcagt gatagagatc gtcgacgagc tcgtttagtg aaccgtcaga
      481 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg gaccgatcca
      541 gcctccggac tctagcgttt aaacttaagc ttggtaccga gctcggatcc actagtccag
      601 tgtggtggaa ttctgcagat atcaacaagt ttgtacaaaa aagcaggctc caccatgatt
      661 gagctgtctc tcatcgattt ctatctctgt ttcctcgcct tccttctttt cctggtcttg
      721 attatgctga taattttctg gttcagtctc gaattgcaag accataacga gacatgccac
      781 gccgacccag ctttcgacat gccacgccga cccagctttc gacatgccac gccgacccag
      841 ctttcttgta caaagtggtt gatatcaagg ccgccgacat caccagcctg tacaagaaag
      901 tggggtggag ccatccccag ttcgagaagg gcggaggatc tggcggcgga agcggcggag
      961 gaagctggtc ccaccctcag tttgagaaag gcaccgagct gggcagcacc atggccagct
     1021 acccctacga cgtgcccgac tacgccaagg acaacaccgt gcccctgaag ctgatcgccc
     1081 tgctggccaa cggcgagttc cactctggcg agcagctggg cgagacactg ggcatgagca
     1141 gagccgccat caacaagcac atccagaccc tgcgggactg gggcgtggac gtgttcacag
     1201 tgcccggcaa gggctacagc ctgcccgagc ccatccagct gctgaacgcc aagcagatcc
     1261 tgggccagct ggatggcgga tctgtggccg tgctgcccgt gatcgacagc accaaccagt
     1321 atctgctcga ccggatcggc gagctgaagt ccggcgacgc ctgtatcgcc gagtaccagc
     1381 aggccggcag aggcggcagg ggcagaaagt ggttcagccc cttcggcgcc aacctgtacc
     1441 tgagcatgtt ctggagactg gaacagggcc cagccgccgc tattggcctg agcctggtca
     1501 tcggcatcgt gatggccgag gtgctgcgga agctgggcgc cgacaaagtg cgcgtgaagt
     1561 ggcccaacga cctgtatctg caggaccgga agctggccgg catcctggtg gaactgaccg
     1621 gcaaaacagg cgacgccgcc cagatcgtga tcggcgctgg catcaacatg gccatgcggc
     1681 gggtggaaga gagcgtggtc aaccagggct ggatcaccct gcaggaagcc ggcatcaacc
     1741 tggaccggaa caccctggcc gccatgctga tcagagagct gagagccgcc ctggaactgt
     1801 tcgagcagga aggcctggcc ccctacctga gcagatggga gaagctggac aacttcatca
     1861 accggcccgt gaaactgatc atcggcgaca aagagatctt cggcatcagc cggggcatcg
     1921 acaagcaggg ggccctgctc ctggaacagg acggcatcat caagccctgg atgggcggcg
     1981 agatcagcct gcggagcgcc gagaagtgat gactcgagtc tagagggccc gtttaaaccc
     2041 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     2101 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     2161 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     2221 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     2281 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     2341 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2401 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2461 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     2521 tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa
     2581 gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga
     2641 cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct
     2701 cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc
     2761 gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat
     2821 cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct
     2881 attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc
     2941 ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc
     3001 agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg
     3061 atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca
     3121 ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc
     3181 ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg
     3241 gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg
     3301 tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact
     3361 tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca
     3421 ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg
     3481 cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa
     3541 tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg
     3601 gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg
     3661 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     3721 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     3781 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     3841 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     3901 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     3961 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4021 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4081 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4141 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4201 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4261 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4321 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4381 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4441 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4501 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     4561 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     4621 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     4681 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4741 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     4801 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4861 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     4921 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     4981 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5041 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5101 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5161 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5221 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5281 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5341 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5401 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5461 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     5521 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     5581 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     5641 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     5701 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     5761 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     5821 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     5881 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     5941 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6001 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6061 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
     6121 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     6181 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     6241 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
     6301 gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata
     6361 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     6421 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     6481 gacgtatgtt cccatagtaa cgccaatag
//
