LOCUS       Exported                7578 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  MAC-tagged gene expression.
ACCESSION   .
VERSION     .
KEYWORDS    MAC_N_GSK3B
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7578)
  AUTHORS   Liu X, Salokas K, Tamene F, Jiu Y, Weldatsadik RG, Ohman T, 
            Varjosalo M
  TITLE     An AP-MS- and BioID-compatible MAC-tag enables comprehensive mapping
            of protein interactions and subcellular localizations.
  JOURNAL   Nat Commun. 2018 Mar 22;9(1):1188. doi: 10.1038/s41467-018-03523-2.
  PUBMED    29568061
REFERENCE   2  (bases 1 to 7578)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7578
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        30..409
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        410..613
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     564..584
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    615..633
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    636..654
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     664..688
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             856..1818
                     /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="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             1819..1845
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1876..1899
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1936..1959
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     protein_bind    1972..1996
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2819..2830
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     protein_bind    complement(3263..3287)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3325..3342)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3331..3555
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3688..3707)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3839..3886
                     /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             3894..4916
                     /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    5046..5167
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5083..5102)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5137..5156
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5216..5232)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5216..5232)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5229..5251)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5240..5256
                     /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(5264..5294)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5309..5330
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5447..5464)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5618..6206)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5698..5717)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6377..7237)
                     /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     7000..7019
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7238..7342)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7417..7436)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tcgcgatgta cgggccagat atacgcgttg acattgatta ttgactagtt attaatagta
       61 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
      121 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
      181 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
      241 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
      301 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
      361 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt
      421 ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
      481 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
      541 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
      601 tataagcaga gctctcccta tcagtgatag agatctccct atcagtgata gagatcgtcg
      661 acgagctcgt ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct
      721 ccatagaaga caccgggacc gatccagcct ccggactcta gcgtttaaac ttaagcttgg
      781 taccgagctc ggatccacta gtccagtgtg gtggaattct gcagatatcc agcacagtgg
      841 cggccgctcg agaccatgaa ggacaacacc gtgcccctga agctgatcgc cctgctggcc
      901 aacggcgagt tccactctgg cgagcagctg ggcgagacac tgggcatgag cagagccgcc
      961 atcaacaagc acatccagac cctgcgggac tggggcgtgg acgtgttcac agtgcccggc
     1021 aagggctaca gcctgcccga gcccatccag ctgctgaacg ccaagcagat cctgggccag
     1081 ctggatggcg gatctgtggc cgtgctgccc gtgatcgaca gcaccaacca gtatctgctc
     1141 gaccggatcg gcgagctgaa gtccggcgac gcctgtatcg ccgagtacca gcaggccggc
     1201 agaggcggca ggggcagaaa gtggttcagc cccttcggcg ccaacctgta cctgagcatg
     1261 ttctggagac tggaacaggg cccagccgcc gctattggcc tgagcctggt catcggcatc
     1321 gtgatggccg aggtgctgcg gaagctgggc gccgacaaag tgcgcgtgaa gtggcccaac
     1381 gacctgtatc tgcaggaccg gaagctggcc ggcatcctgg tggaactgac cggcaaaaca
     1441 ggcgacgccg cccagatcgt gatcggcgct ggcatcaaca tggccatgcg gcgggtggaa
     1501 gagagcgtgg tcaaccaggg ctggatcacc ctgcaggaag ccggcatcaa cctggaccgg
     1561 aacaccctgg ccgccatgct gatcagagag ctgagagccg ccctggaact gttcgagcag
     1621 gaaggcctgg ccccctacct gagcagatgg gagaagctgg acaacttcat caaccggccc
     1681 gtgaaactga tcatcggcga caaagagatc ttcggcatca gccggggcat cgacaagcag
     1741 ggggccctgc tcctggaaca ggacggcatc atcaagccct ggatgggcgg cgagatcagc
     1801 ctgcggagcg ccgagaagta cccatacgat gttcctgact atgccggtac cgagctcgga
     1861 tccaccatgg ctagctggag ccacccgcag ttcgagaaag gtggaggttc cggaggtgga
     1921 tcgggaggtg gatcgtggag ccacccgcag ttcgaaaaag cggccgatat cacaagtttg
     1981 tacaaaaaag caggctccac catgtcaggg cggcccagaa ccacctcctt tgcggagagc
     2041 tgcaagccgg tgcagcagcc ttcagctttt ggcagcatga aagttagcag agacaaggac
     2101 ggcagcaagg tgacaacagt ggtggcaact cctgggcagg gtccagacag gccacaagaa
     2161 gtcagctata cagacactaa agtgattgga aatggatcat ttggtgtggt atatcaagcc
     2221 aaactttgtg attcaggaga actggtcgcc atcaagaaag tattgcagga caagagattt
     2281 aagaatcgag agctccagat catgagaaag ctagatcact gtaacatagt ccgattgcgt
     2341 tatttcttct actccagtgg tgagaagaaa gatgaggtct atcttaatct ggtgctggac
     2401 tatgttccgg aaacagtata cagagttgcc agacactata gtcgagccaa acagacgctc
     2461 cctgtgattt atgtcaagtt gtatatgtat cagctgttcc gaagtttagc ctatatccat
     2521 tcctttggaa tctgccatcg ggatattaaa ccgcagaacc tcttgttgga tcctgatact
     2581 gctgtattaa aactctgtga ctttggaagt gcaaagcagc tggtccgagg agaacccaat
     2641 gtttcgtata tctgttctcg gtactatagg gcaccagagt tgatctttgg agccactgat
     2701 tatacctcta gtatagatgt atggtctgct ggctgtgtgt tggctgagct gttactagga
     2761 caaccaatat ttccagggga tagtggtgtg gatcagttgg tagaaataat caaggtcctg
     2821 ggaactccaa caagggagca aatcagagaa atgaacccaa actacacaga atttaaattc
     2881 cctcaaatta aggcacatcc ttggactaag gtcttccgac cccgaactcc accggaggca
     2941 attgcactgt gtagccgtct gctggagtat acaccaactg cccgactaac accactggaa
     3001 gcttgtgcac attcattttt tgatgaatta cgggacccaa atgtcaaact accaaatggg
     3061 cgagacacac ctgcactctt caacttcacc actcaagaac tgtcaagtaa tccacctctg
     3121 gctaccatcc ttattcctcc tcatgctcgg attcaagcag ctgcttcaac ccccacaaat
     3181 gccacagcag cgtcagatgc taatactgga gaccgtggac agaccaataa tgctgcttct
     3241 gcatcagctt ccaactccac cgacccagct ttcttgtaca aagtggtgac gtaagctagg
     3301 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     3361 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     3421 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     3481 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     3541 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     3601 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     3661 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     3721 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3781 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtacctaga
     3841 agttcctatt ccgaagttcc tattctctag aaagtatagg aacttccttg gccaaaaagc
     3901 ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg
     3961 acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc
     4021 gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt
     4081 atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt ggggaattca
     4141 gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc
     4201 ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat gcgatcgctg
     4261 cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat
     4321 acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa
     4381 ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt
     4441 gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg
     4501 tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg
     4561 attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc
     4621 agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg cggctccggg
     4681 cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg
     4741 atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg
     4801 tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag
     4861 tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaatagcacg
     4921 tactacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4981 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     5041 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     5101 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     5161 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     5221 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5281 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5341 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     5401 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     5461 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     5521 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5581 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5641 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5701 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5761 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5821 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5881 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5941 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6001 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     6061 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     6121 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     6181 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     6241 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     6301 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     6361 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6421 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6481 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6541 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6601 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6661 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6721 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6781 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6841 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6901 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6961 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7021 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     7081 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     7141 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     7201 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     7261 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     7321 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     7381 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
     7441 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
     7501 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
     7561 taggcgtttt gcgctgct
//
