LOCUS       sequence-441730-        6861 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MAC-tagged COMMD8 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6861)
  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 6861)
  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_COMMD8
FEATURES             Location/Qualifiers
     source          1..6861
                     /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(2517..2541)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2579..2596)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2585..2809
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2942..2961)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3093..3140
                     /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             3148..4170
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3148..4170
                     /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    4300..4421
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4337..4356)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4391..4410
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4470..4486)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4470..4486)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4483..4505)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4494..4510
                     /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(4518..4524,4525..4542,4543..4548))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4563..4584
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4701..4718)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4872..5460)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4952..4971)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5631..6422,6423..6491))
                     /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     6254..6273
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6492..6596)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6671..6690)
                     /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 gttggcatgg agccggaaga
     1981 ggggacgccc ttgtggcggc tgcagaagct gccggccgag ctgggcccgc agcttcttca
     2041 caaaataatt gatggcattt gtggtcgagc ttatcctgtg taccaagatt atcacactgt
     2101 ttgggaatca gaagaatgga tgcacgtttt agaagatatt gccaaatttt tcaaagccat
     2161 agttggtaaa aacttacctg atgaagagat atttcagcag ttgaatcagt tgaattcact
     2221 tcatcaagaa actatcatga aatgcgtgaa aagtaggaaa gatgaaatca aacaggctct
     2281 gtcaagagaa atagttgcta tttcctctgc acagctacag gattttgatt ggcaggtaaa
     2341 gcttgcactt tccagtgaca agattgctgc attacgaatg ccacttttaa gcctgcatct
     2401 agatgtaaaa gaaaatggtg aagtaaaacc ttattctatt gaaatgagta gagaggagct
     2461 gcagaatcta atacagtcct tggaagcagc gaataaggtg gtcctgcagt tgaaataccc
     2521 aactttcttg tacaaagtgg tgacgtaagc taggggcccg tttaaacccg ctgatcagcc
     2581 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     2641 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     2701 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     2761 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     2821 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     2881 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     2941 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3001 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3061 aaacttgatt agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct
     3121 ctagaaagta taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag
     3181 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa
     3241 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc
     3301 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc
     3361 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc
     3421 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg
     3481 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg
     3541 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc
     3601 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg
     3661 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg
     3721 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca
     3781 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc
     3841 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg
     3901 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac
     3961 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga
     4021 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga
     4081 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc
     4141 cccagcactc gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc
     4201 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4261 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     4321 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4381 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     4441 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4501 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4561 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4621 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4681 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4741 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4801 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4861 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4921 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4981 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     5041 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5101 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     5161 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     5221 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     5281 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     5341 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     5401 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     5461 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5521 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5581 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     5641 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     5701 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     5761 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     5821 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     5881 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5941 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     6001 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     6061 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     6121 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     6181 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     6241 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     6301 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     6361 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     6421 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     6481 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     6541 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     6601 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     6661 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     6721 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
     6781 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
     6841 tacgggccag atatacgcgt t
//
