LOCUS       Exported                7644 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  MAC-tagged gene expression.
ACCESSION   .
VERSION     .
KEYWORDS    MAC_C_TGON2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7644)
  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 7644)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7644
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        13..392
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        393..596
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     547..567
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    598..616
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    619..637
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     647..671
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    814..838
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2159..2183)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2230..2253
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             2290..2313
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             2344..2370
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2371..3330
                     /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(3374..3391)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3380..3604
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3737..3756)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3888..3935
                     /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             3943..4965
                     /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    5095..5216
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5132..5151)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5186..5205
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5265..5281)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5265..5281)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5278..5300)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5289..5305
                     /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(5313..5343)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5358..5379
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5496..5513)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5667..6255)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5747..5766)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6426..7286)
                     /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     7049..7068
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7287..7391)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7466..7485)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
       61 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      121 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      181 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      241 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      301 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      361 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
      421 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
      481 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
      541 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctcc
      601 ctatcagtga tagagatctc cctatcagtg atagagatcg tcgacgagct cgtttagtga
      661 accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg
      721 accgatccag cctccggact ctagcgttta aacttaagct tggtaccgag ctcggatcca
      781 ctagtccagt gtggtggaat tctgcagata tcaacaagtt tgtacaaaaa agcaggctcc
      841 accatgcggt tcgtagttgc cttggtcctc ctgaacgtcg cagcggcggg agccgtgccg
      901 ctcttggcca ccgaaagcgt caagcaagaa gatgctggag tacggccttc tgcaggaaac
      961 gtctccaccc accccagctt gagccaacgg cctggaggct ctaccaagtc gcatccggag
     1021 ccgcagactc caaaagacag ccctagcaag tcgagtgcgg aggcgcagac cccagaagac
     1081 acccccaaca agtcgggtgc ggaggcaaag acccaaaaag acagctccaa caagtcgggt
     1141 gcggaggcaa agacccaaaa aggcagcact agcaagtcgg gttcggaggc gcagaccaca
     1201 aaagacagca ctagtaagtc gcatccggag ctgcagactc caaaagacag cactggcaaa
     1261 tcgggtgcgg aggcgcagac cccagaagac agccccaaca ggtcgggtgc ggaggcaaag
     1321 acccaaaaag acagccctag caagtcaggt tcggaggcgc agaccacaaa agatgtccct
     1381 aataagtcgg gtgcggacgg ccagacccca aaagacggct ccagcaagtc gggtgcggag
     1441 gatcagaccc caaaagacgt ccctaacaag tcgggtgcgg agaagcagac tccaaaagac
     1501 ggctctaaca agtccggtgc agaggagcag ggcccaatag acgggcccag caagtcgggt
     1561 gcggaggagc agacctcaaa agacagccct aacaaggtgg ttccagagca gccttcccgg
     1621 aaagaccatt ccaagcccat ctccaaccct tctgataaca aggagctccc caaggctgac
     1681 acaaaccagc ttgctgacaa agggaagctt tctcctcatg ctttcaaaac cgaatctggg
     1741 gaggaaactg acctcatttc tcccccgcag gaggaagtta agtcttcaga gcctactgag
     1801 gatgtggagc ccaaagaggc tgaagatgat gatacaggac ccgaggaggg ctcaccgccc
     1861 aaagaagaga aagaaaagat gtccggttct gcctccagtg agaaccgtga agggacactt
     1921 tcggattcca cgggtagcga gaaggatgac ctttatccga acggttctgg aaatggcagc
     1981 gcggagagca gccacttctt tgcatatctg gtgactgcag ccattcttgt ggctgtcctc
     2041 tatatcgctc atcacaacaa gcggaagatc attgcttttg tcctggaagg aaaaagatct
     2101 aaagtcaccc ggcggccaaa ggccagtgac taccaacgtt tggaccagaa gtccttggac
     2161 ccagctttct tgtacaaagt ggttgatatc aaggccgccg acatcaccag cctgtacaag
     2221 aaagtggggt ggagccatcc ccagttcgag aagggcggag gatctggcgg cggaagcggc
     2281 ggaggaagct ggtcccaccc tcagtttgag aaaggcaccg agctgggcag caccatggcc
     2341 agctacccct acgacgtgcc cgactacgcc aaggacaaca ccgtgcccct gaagctgatc
     2401 gccctgctgg ccaacggcga gttccactct ggcgagcagc tgggcgagac actgggcatg
     2461 agcagagccg ccatcaacaa gcacatccag accctgcggg actggggcgt ggacgtgttc
     2521 acagtgcccg gcaagggcta cagcctgccc gagcccatcc agctgctgaa cgccaagcag
     2581 atcctgggcc agctggatgg cggatctgtg gccgtgctgc ccgtgatcga cagcaccaac
     2641 cagtatctgc tcgaccggat cggcgagctg aagtccggcg acgcctgtat cgccgagtac
     2701 cagcaggccg gcagaggcgg caggggcaga aagtggttca gccccttcgg cgccaacctg
     2761 tacctgagca tgttctggag actggaacag ggcccagccg ccgctattgg cctgagcctg
     2821 gtcatcggca tcgtgatggc cgaggtgctg cggaagctgg gcgccgacaa agtgcgcgtg
     2881 aagtggccca acgacctgta tctgcaggac cggaagctgg ccggcatcct ggtggaactg
     2941 accggcaaaa caggcgacgc cgcccagatc gtgatcggcg ctggcatcaa catggccatg
     3001 cggcgggtgg aagagagcgt ggtcaaccag ggctggatca ccctgcagga agccggcatc
     3061 aacctggacc ggaacaccct ggccgccatg ctgatcagag agctgagagc cgccctggaa
     3121 ctgttcgagc aggaaggcct ggccccctac ctgagcagat gggagaagct ggacaacttc
     3181 atcaaccggc ccgtgaaact gatcatcggc gacaaagaga tcttcggcat cagccggggc
     3241 atcgacaagc agggggccct gctcctggaa caggacggca tcatcaagcc ctggatgggc
     3301 ggcgagatca gcctgcggag cgccgagaag tgatgactcg agtctagagg gcccgtttaa
     3361 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     3421 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     3481 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     3541 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     3601 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     3661 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     3721 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3781 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3841 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtacctagaa gttcctattc
     3901 cgaagttcct attctctaga aagtatagga acttccttgg ccaaaaagcc tgaactcacc
     3961 gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag
     4021 ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc
     4081 ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt
     4141 gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg
     4201 acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa
     4261 ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt
     4321 agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg
     4381 cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac
     4441 gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac
     4501 tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac
     4561 aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac
     4621 gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc
     4681 tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc
     4741 cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct
     4801 tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca
     4861 caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat
     4921 agtggaaacc gacgccccag cactcgtccg agggcaaagg aatagcacgt actacgagat
     4981 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     5041 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     5101 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     5161 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5221 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     5281 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5341 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5401 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5461 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5521 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5581 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5641 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5701 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5761 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5821 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5881 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5941 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     6001 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6061 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     6121 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6181 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6241 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6301 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6361 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6421 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6481 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6541 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6601 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6661 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6721 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     6781 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     6841 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6901 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6961 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     7021 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     7081 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     7141 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     7201 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     7261 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7321 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7381 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc
     7441 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     7501 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
     7561 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
     7621 cgctgcttcg cgatgtacgg gcca
//
