LOCUS       sequence-365404-        7190 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express yeast MCP1 in Expi293F cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7190)
  AUTHORS   Adlakha J, Hong Z, Li P, Reinisch KM
  TITLE     Structural and biochemical insights into lipid transport by VPS13 
            proteins.
  JOURNAL   J Cell Biol. 2022 Jun 6;221(5):e202202030. doi: 
            10.1083/jcb.202202030. Epub 2022 Mar 31.
  PUBMED    35357422
REFERENCE   2  (bases 1 to 7190)
  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"; journalName: "J Cell 
            Biol."; date: "2022-06-6"; volume: "221(5):e202202030. doi"; pages: 
            " 10.1083/jcb.202202030. Epub 2022 Mar 31"
COMMENT     Sequence Label: pCMV10-3xFLAG-prs-MCP1
FEATURES             Location/Qualifiers
     source          1..7190
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..148
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     140..157
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     141..157
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        318..697
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        698..901
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     852..872
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             931..996
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             997..1020
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    1949..2571
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2148..2167)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2595..2615)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2613..2910
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2964..3758
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3018..3037)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3628..3647
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4410..4544
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4447..4466)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4501..4520
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4549..4568)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4550..4568)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4582..4598)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4582..4598)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4595..4617)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4606..4622
                     /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(4630..4636,4637..4654,4655..4660))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4675..4696
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4813..4830)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4984..5572)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5064..5083)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5743..6534,6535..6603))
                     /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     6366..6385
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6604..6708)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6735..7163
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6822..6841)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7032..7053
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
       61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
      121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
      181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
      241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
      361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      901 tcgtttagtg aaccgtcaga attaaccatg gactacaaag accatgacgg tgattataaa
      961 gatcatgaca tcgattacaa ggatgacgat gacaagctgg aagttctgtt ccaggggccc
     1021 cttgcggccg ccatgataaa gttgcatgaa gtgcctccag aaccagtaga ccctgctagt
     1081 ctccctcatg atgttaatgc acatagcccc gaaggagatg gtaatcctga taagaggaaa
     1141 aaaatatttg gcattccgta tcctttttcc aggagttcat gtcggagatt cctttggaac
     1201 tgtcaaaaga tttcggtact gcccatggca ttgtatttcc cactacatgc agcaaataca
     1261 ttaatcacgc cggcagtatc acctgattct gctcctgacg acgttttaat gatggtgaga
     1321 gagattttac cgtccataac taccaagctg ttagttgcgg gtattactct tcatgtttct
     1381 gctggtgtct tgctccgaat cgtgaacaac tggaacaagc caagaagaaa tagacatcga
     1441 catttaaaga tatcggccga acaagatcta tcccaagatt cgattggttt aactggtgga
     1501 atttctggct atttatttgg attgtataag accttccgca ttcctccaca agtgattagt
     1561 ggatacatcc ttgttcccgt cctgatatac cacttattaa ttatgaaatg ggtgccgaat
     1621 tccatctcta ctgaggtgga tttcgcctct ataaagcaat tactatctag caagaacagg
     1681 tggtggaaat ggcttggagg cctggtgccg ttagctattc tcctcgagtc aggtgtatat
     1741 catataggtt cggggctctg ccgttatttc ggtgttagaa aaatgactag caggaaaaaa
     1801 tggtccacag ctatcaatct attaacactc gttgggttcg tttctctgat tagattgatg
     1861 aaagaggatt ctacgaaact gggtcccaac caattcgaaa gcattttcaa aaagatccgg
     1921 ttgctgttgc acgtgaatta gggatcccgg gtggcatccc tgtgacccct ccccagtgcc
     1981 tctcctggcc ctggaagttg ccactccagt gcccaccagc cttgtcctaa taaaattaag
     2041 ttgcatcatt ttgtctgact aggtgtcctt ctataatatt atggggtgga ggggggtggt
     2101 atggagcaag gggcaagttg ggaagacaac ctgtagggcc tgcggggtct attgggaacc
     2161 aagctggagt gcagtggcac aatcttggct cactgcaatc tccgcctcct gggttcaagc
     2221 gattctcctg cctcagcctc ccgagttgtt gggattccag gcatgcatga ccaggctcag
     2281 ctaatttttg tttttttggt agagacgggg tttcaccata ttggccaggc tggtctccaa
     2341 ctcctaatct caggtgatct acccaccttg gcctcccaaa ttgctgggat tacaggcgtg
     2401 aaccactgct cccttccctg tccttctgat tttaaaataa ctataccagc aggaggacgt
     2461 ccagacacag cataggctac ctggccatgc ccaaccggtg ggacatttga gttgcttgct
     2521 tggcactgtc ctctcatgcg ttgggtccac tcagtagatg cctgttgaat tgggtacgcg
     2581 gccagcttgg ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     2641 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     2701 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     2761 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     2821 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     2881 attccagaag tagtgaggag gcttttttgg aggaattgat cagcttggga tctgatcaag
     2941 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     3001 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     3061 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     3121 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     3181 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     3241 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     3301 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     3361 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     3421 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     3481 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     3541 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     3601 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     3661 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     3721 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     3781 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     3841 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     3901 ggatctcatg ctggagttct tcgcccaccc cgggctcgat cccctcgcga gttggttcag
     3961 ctgctgcctg aggctggacg acctcgcgga gttctaccgg cagtgcaaat ccgtcggcat
     4021 ccaggaaacc agcagcggct atccgcgcat ccatgccccc gaactgcagg agtggggagg
     4081 cacgatggcc gctttggtcg acccggacgg gacgctcctg cgcctgatac agaacgaatt
     4141 gcttgcaggc atctcatgag tgtgtcttcc cgttttccgc ctgaggtcac tgcgtggatg
     4201 gagcgctggc gcctgctgcg cgacggcgag ctgctcacca cccactcgcc aagctggaac
     4261 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gccgatcata atcagccata
     4321 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     4381 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     4441 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     4501 gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatcaattcc ctatagtgag
     4561 tcgtattaaa ttcgtaatca tgtcatagct gtttcctgtg tgaaattgtt atccgctcac
     4621 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     4681 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     4741 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4801 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4861 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4921 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4981 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5041 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5101 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5161 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5221 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5281 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5341 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5401 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5461 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5521 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     5581 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     5641 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     5701 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     5761 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5821 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5881 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5941 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     6001 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     6061 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     6121 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     6181 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     6241 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     6301 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     6361 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     6421 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6481 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6541 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     6601 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     6661 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6721 aaaagtgcca cctgacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     6781 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     6841 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     6901 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     6961 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     7021 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     7081 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     7141 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
//
