LOCUS       sequence-441751-        6900 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MAC-tagged COMMD3 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6900)
  AUTHORS   Laulumaa S, Kumpula EP, Huiskonen JT, Varjosalo M
  TITLE     Structure and interactions of the endogenous human Commander 
            complex.
  JOURNAL   Nat Struct Mol Biol. 2024 Jun;31(6):925-938. doi: 
            10.1038/s41594-024-01246-1. Epub 2024 Mar 8.
  PUBMED    38459129
REFERENCE   2  (bases 1 to 6900)
  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"; doi: 
            "10.1038/s41594-024-01246-1"; journalName: "Nat Struct Mol Biol"; 
            date: "2024-06"; volume: "31"; issue: "6"; pages: "925-938"
COMMENT     Sequence Label: MAC_N_COMMD3
FEATURES             Location/Qualifiers
     source          1..6900
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    586..604
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    607..625
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     635..659
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(827..1177,1178..1180,1181..1789)
                     /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             1790..1816
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1847..1870
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1907..1930
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     protein_bind    1943..1967
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2556..2580)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2618..2635)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2624..2848
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2981..3000)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3132..3179
                     /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             3187..4209
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3187..4209
                     /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    4339..4460
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4376..4395)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4430..4449
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4509..4525)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4509..4525)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4522..4544)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4533..4549
                     /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(4557..4563,4564..4581,4582..4587))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4602..4623
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4740..4757)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4911..5499)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4991..5010)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5670..6461,6462..6530))
                     /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     6293..6312
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6531..6635)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6710..6729)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctccct atcagtgata
      601 gagatctccc tatcagtgat agagatcgtc gacgagctcg tttagtgaac cgtcagatcg
      661 cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc
      721 tccggactct agcgtttaaa cttaagcttg gtaccgagct cggatccact agtccagtgt
      781 ggtggaattc tgcagatatc cagcacagtg gcggccgctc gagaccatga aggacaacac
      841 cgtgcccctg aagctgatcg ccctgctggc caacggcgag ttccactctg gcgagcagct
      901 gggcgagaca ctgggcatga gcagagccgc catcaacaag cacatccaga ccctgcggga
      961 ctggggcgtg gacgtgttca cagtgcccgg caagggctac agcctgcccg agcccatcca
     1021 gctgctgaac gccaagcaga tcctgggcca gctggatggc ggatctgtgg ccgtgctgcc
     1081 cgtgatcgac agcaccaacc agtatctgct cgaccggatc ggcgagctga agtccggcga
     1141 cgcctgtatc gccgagtacc agcaggccgg cagaggcggc aggggcagaa agtggttcag
     1201 ccccttcggc gccaacctgt acctgagcat gttctggaga ctggaacagg gcccagccgc
     1261 cgctattggc ctgagcctgg tcatcggcat cgtgatggcc gaggtgctgc ggaagctggg
     1321 cgccgacaaa gtgcgcgtga agtggcccaa cgacctgtat ctgcaggacc ggaagctggc
     1381 cggcatcctg gtggaactga ccggcaaaac aggcgacgcc gcccagatcg tgatcggcgc
     1441 tggcatcaac atggccatgc ggcgggtgga agagagcgtg gtcaaccagg gctggatcac
     1501 cctgcaggaa gccggcatca acctggaccg gaacaccctg gccgccatgc tgatcagaga
     1561 gctgagagcc gccctggaac tgttcgagca ggaaggcctg gccccctacc tgagcagatg
     1621 ggagaagctg gacaacttca tcaaccggcc cgtgaaactg atcatcggcg acaaagagat
     1681 cttcggcatc agccggggca tcgacaagca gggggccctg ctcctggaac aggacggcat
     1741 catcaagccc tggatgggcg gcgagatcag cctgcggagc gccgagaagt acccatacga
     1801 tgttcctgac tatgccggta ccgagctcgg atccaccatg gctagctgga gccacccgca
     1861 gttcgagaaa ggtggaggtt ccggaggtgg atcgggaggt ggatcgtgga gccacccgca
     1921 gttcgaaaaa gcggccgata tcacaagttt gtacaaaaaa gttggcacca tggagctctc
     1981 ggagtctgtg cagaaaggct tccagatgct ggcggatccc cgctccttcg actccaacgc
     2041 cttcacgctt ctcctccggg cggcattcca gagtctgctg gacgcccagg cggacgaggc
     2101 cgtgttagat catccagact tgaaacatat cgacccagtg gttttaaaac attgtcatgc
     2161 agcagctgca acttacatac tagaggcagg aaagcaccga gctgacaagt caactctaag
     2221 cacttatcta gaagactgta aatttgacag agagcgaata gaactgtttt gcacggaata
     2281 tcagaataat aagaattccc tagaaatcct actgggaagt ataggcagat ctctccctca
     2341 tataacggat gtttcttggc gcttggaata tcagataaag accaatcaac ttcataggat
     2401 gtacagacct gcatatttgg tgaccttaag tgtacagaac actgattccc catcctatcc
     2461 agagattagt tttagttgca gcatggaaca attacaggac ttggtgggga aacttaaaga
     2521 tgcttcgaaa agcctggaaa gagcaactca gttgttgcca actttcttgt acaaagtggt
     2581 gacgtaagct aggggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     2641 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     2701 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     2761 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     2821 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     2881 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     2941 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     3001 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     3061 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     3121 tcacgtacct agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc
     3181 ttggccaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc
     3241 gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc
     3301 gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa
     3361 gatcgttatg tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac
     3421 attggggaat tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg
     3481 ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg
     3541 gatgcgatcg ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa
     3601 ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg
     3661 tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat
     3721 gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc
     3781 ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag
     3841 gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg
     3901 gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg
     3961 ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt
     4021 gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc
     4081 ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat
     4141 ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca
     4201 aaggaatagc acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     4261 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     4321 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     4381 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     4441 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     4501 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4561 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4621 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4681 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4741 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4801 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4861 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4921 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4981 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5041 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5101 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5161 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5221 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5281 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5341 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5401 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5461 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5521 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5581 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5641 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5701 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5761 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5821 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5881 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5941 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     6001 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6061 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6121 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6181 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6241 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6301 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6361 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6421 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6481 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6541 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6601 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6661 gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct
     6721 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt
     6781 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
     6841 atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
//
