LOCUS       sequence-441677-        6912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MAC-tagged COMMD4 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6912)
  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 6912)
  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_COMMD4
FEATURES             Location/Qualifiers
     source          1..6912
                     /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(2568..2592)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2630..2647)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2636..2860
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2993..3012)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3144..3191
                     /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             3199..4221
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3199..4221
                     /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    4351..4472
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4388..4407)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4442..4461
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4521..4537)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4521..4537)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4534..4556)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4545..4561
                     /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(4569..4575,4576..4593,4594..4599))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4614..4635
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4752..4769)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4923..5511)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5003..5022)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5682..6473,6474..6542))
                     /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     6305..6324
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6543..6647)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6722..6741)
                     /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 tgaggttccg
     1981 gttctgtggt gatctggact gtcccgactg ggtcctggca gaaatcagca cgctggccaa
     2041 gatgtcctct gtgaagttgc ggctgctctg cagccaggta ctaaaggagc tgctgggaca
     2101 ggggattgat tatgagaaga tcctgaagct cacggctgac gccaagtttg agtcaggcga
     2161 tgtgaaggcc acagtggcag tgctgagttt catcctctcc agtgcggcca agcacagtgt
     2221 cgatggcgaa tccttgtcca gtgaactgca gcagctgggg ctgcccaaag agcacgcggc
     2281 cagcctgtgc cgctgttatg aggagaagca aagccccttg cagaagcact tgcgggtctg
     2341 cagcctacgc atgaataggt tggcaggtgt gggctggcgg gtggactaca ccctgagctc
     2401 cagcctgctg caatccgtgg aagagcccat ggtgcacctg cggctggagg tggcagctgc
     2461 cccagggacc ccagcccagc ctgttgccat gtccctctca gcagacaagt tccaggtcct
     2521 cctggcagaa ctgaagcagg cccagaccct gatgagctcc ctgggcttgc caactttctt
     2581 gtacaaagtg gtgacgtaag ctaggggccc gtttaaaccc gctgatcagc ctcgactgtg
     2641 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     2701 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     2761 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     2821 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     2881 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     2941 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3001 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3061 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3121 tagggtgatg gttcacgtac ctagaagttc ctattccgaa gttcctattc tctagaaagt
     3181 ataggaactt ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg
     3241 atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt
     3301 gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat
     3361 ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg
     3421 gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca
     3481 cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc
     3541 gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca
     3601 ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct
     3661 gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg
     3721 caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg
     3781 cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt
     3841 gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg
     3901 aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag
     3961 cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat
     4021 cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca
     4081 atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc
     4141 gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact
     4201 cgtccgaggg caaaggaata gcacgtacta cgagatttcg attccaccgc cgccttctat
     4261 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     4321 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     4381 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     4441 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     4501 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     4561 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     4621 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     4681 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4741 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4801 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4861 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4921 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4981 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     5041 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     5101 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     5161 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     5221 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     5281 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5341 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     5401 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5461 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5521 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5581 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5641 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5701 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5761 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5821 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5881 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5941 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     6001 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     6061 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     6121 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     6181 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     6241 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     6301 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6361 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6421 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     6481 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6541 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     6601 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6661 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
     6721 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
     6781 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
     6841 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
     6901 gatatacgcg tt
//
