LOCUS       sequence-441676-        7206 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MAC-tagged VPS26C in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7206)
  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 7206)
  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_VPS26C
FEATURES             Location/Qualifiers
     source          1..7206
                     /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(2862..2886)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2924..2941)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2930..3154
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3287..3306)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3438..3485
                     /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             3493..4515
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3493..4515
                     /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    4645..4766
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4682..4701)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4736..4755
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4815..4831)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4815..4831)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4828..4850)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4839..4855
                     /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(4863..4869,4870..4887,4888..4893))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4908..4929
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5046..5063)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5217..5805)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5297..5316)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5976..6767,6768..6836))
                     /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     6599..6618
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6837..6941)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7016..7035)
                     /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 tggggaccgc
     1981 cctggacatc aagattaaaa gagcgaataa agtttatcac gccggggaag tgctctctgg
     2041 cgtggtggtc atatcgagta aggattcagt ccaacaccag ggagtgtctt tgaccatgga
     2101 aggaactgta aacctccagc tcagtgccaa aagtgtgggt gtgtttgaag ctttttataa
     2161 ttctgttaag cctatccaga ttatcaacag caccatagaa atggtgaagc cggggaaatt
     2221 tcccagcggc aaaacagaaa tcccttttga atttcctctg cacttgaagg gtaacaaagt
     2281 tctgtatgag acgtatcatg gcgtgtttgt caacattcag tatacactgc gctgtgacat
     2341 gaagcggtct ctgttggcca aggacttgac aaagacctgt gaatttatcg ttcactccgc
     2401 tcctcagaag gggaagttta ctcccagtcc cgtggacttc acgattacac ctgaaacctt
     2461 acagaacgtc aaagagagag ctttgcttcc caaatttctc cttcgaggac atctcaactc
     2521 aacaaactgt gtcatcacgc agccactaac gggagagctg gtggtggaga gctcggaagc
     2581 cgccatcaga agcgtggagc tgcagctggt gcgcgtggag acgtgcgggt gtgcagaagg
     2641 ctatgcccgc gacgccacgg agattcagaa cattcagatc gccgacgggg atgtgtgcag
     2701 gggcctctct gtccccatct acatggtctt ccctaggctg ttcacctgcc ctacactgga
     2761 gaccaccaac ttcaaagtgg aatttgaggt taacatcgtg gtgctgcttc accctgacca
     2821 cctcatcacg gagaacttcc cgctgaagct ctgcaggata ttgccaactt tcttgtacaa
     2881 agtggtgacg taagctaggg gcccgtttaa acccgctgat cagcctcgac tgtgccttct
     2941 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     3001 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     3061 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     3121 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     3181 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     3241 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     3301 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     3361 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     3421 gatggttcac gtacctagaa gttcctattc cgaagttcct attctctaga aagtatagga
     3481 acttccttgg ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa
     3541 aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc
     3601 agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc
     3661 tacaaagatc gttatgttta tcggcacttt gcatcggccg cgctcccgat tccggaagtg
     3721 cttgacattg gggaattcag cgagagcctg acctattgca tctcccgccg tgcacagggt
     3781 gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag
     3841 gccatggatg cgatcgctgc ggccgatctt agccagacga gcgggttcgg cccattcgga
     3901 ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat tgctgatccc
     3961 catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct
     4021 ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct cgtgcacgcg
     4081 gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt cattgactgg
     4141 agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt ctggaggccg
     4201 tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca
     4261 ggatcgccgc ggctccgggc gtatatgctc cgcattggtc ttgaccaact ctatcagagc
     4321 ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc
     4381 cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg
     4441 accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag cactcgtccg
     4501 agggcaaagg aatagcacgt actacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4561 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4621 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     4681 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     4741 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     4801 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     4861 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     4921 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4981 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     5041 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5101 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5161 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5221 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5281 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5341 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5401 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5461 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5521 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5581 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5641 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5701 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5761 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5821 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     5881 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     5941 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     6001 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6061 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6121 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6181 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6241 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6301 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6361 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6421 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6481 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6541 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6601 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6661 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6721 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6781 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6841 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     6901 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     6961 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
     7021 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
     7081 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
     7141 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
     7201 cgcgtt
//
