LOCUS       sequence-235292-        7392 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses myc tagged BioID and human Rab7a in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7392)
  AUTHORS   Hoyer MJ, Chitwood PJ, Ebmeier CC, Striepen JF, Qi RZ, Old WM, 
            Voeltz GK
  TITLE     A Novel Class of ER Membrane Proteins Regulates ER-Associated 
            Endosome Fission.
  JOURNAL   Cell. 2018 Sep 20;175(1):254-265.e14. doi: 
            10.1016/j.cell.2018.08.030. Epub 2018 Sep 13.
  PUBMED    30220460
REFERENCE   2  (bases 1 to 7392)
  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: 
            "10.1016/j.cell.2018.08.030"; journalName: "Cell"; date: 
            "2018-09-20- 20"; volume: "175"; issue: "1"; pages: "254-265.e14"
COMMENT     Sequence Label: myc-BioID-Rab7
FEATURES             Location/Qualifiers
     source          1..7392
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     249..268
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        249..267
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      285..294
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             294..323
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             join(327..674,675..677,678..1286)
                     /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="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             complement(1324..1341)
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     CDS             1359..1679
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     primer_bind     complement(2369..2386)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2375..2599
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2645..3073
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2732..2751)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2942..2963
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3082..3102)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3087..3416
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3267..3402
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3329..3348
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3483..4277
                     /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(3537..3556)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4147..4166
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4451..4572
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4488..4507)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4542..4561
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4621..4637)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4621..4637)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4634..4656)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4645..4661
                     /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(4669..4675,4676..4693,4694..4699))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4714..4735
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4852..4869)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5023..5611)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5103..5122)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5782..6573,6574..6642))
                     /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     6405..6424
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6643..6747)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6822..6841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        7013..7392
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc ttactggctt
      241 atcgaaatta atacgactca ctatagggag acccaagctg gctagccacc atggaacaaa
      301 aactcatctc agaagaggat ctcgacaagg acaacaccgt gcccctgaag ctgatcgccc
      361 tgctggccaa cggcgagttc cactctggcg agcagctggg agagaccctg ggaatgagca
      421 gagccgccat caacaagcac atccagacac tgagagactg gggagtggac gtgttcaccg
      481 tgcctggcaa gggctacagc ctgcctgagc ctatccagct gctgaacgcc aagcagatcc
      541 tgggacagct ggatggcgga agcgtggccg tgctgcctgt gatcgactcc accaatcagt
      601 acctgctgga cagaatcgga gagctgaagt ccggcgacgc ctgcatcgcc gagtaccagc
      661 aggctggcag aggaggcaga ggacggaagt ggttcagccc attcggagcc aacctgtacc
      721 tgtccatgtt ctggagactg gagcagggac ctgctgctgc catcggactg agtctggtga
      781 tcggaatcgt gatggccgag gtgctgagaa agctgggagc cgacaaggtg agagtgaagt
      841 ggcctaatga cctgtacctc caggaccgca agctggctgg catcctggtg gagctgacag
      901 gcaagacagg cgatgccgct cagatcgtga tcggagccgg aatcaacatg gccatgagaa
      961 gagtggagga gagcgtggtg aaccagggct ggatcaccct gcaggaggct ggcatcaacc
     1021 tggaccggaa caccctggcc gccatgctga tcagagagct gagagccgct ctggagctgt
     1081 tcgagcagga gggactggct ccttacctga gcagatggga gaagctggac aacttcatca
     1141 acagacctgt gaagctgatc atcggcgaca aggaaatctt cggcatctcc agaggaatcg
     1201 acaagcaggg agctctgctg ctggagcagg acggaatcat caagccctgg atgggcggag
     1261 aaatctccct gagaagcgca gagaagctcg aggctgatcc caccagatct gcgaattcag
     1321 gtgctggtgc tggtgctggt gctatcctat ctaaactggg agtgcaggtg gaaaccatct
     1381 ccccaggaga cgggcgcacc ttccccaagc gcggccagac ctgcgtggtg cactacaccg
     1441 ggatgcttga agatggaaag aaatttgatt cctcccggga cagaaacaag ccctttaagt
     1501 ttatgctagg caagcaggag gtgatccgag gctgggaaga aggggttgcc cagatgagtg
     1561 tgggtcagag agccaaactg actatatctc cagattatgc ctatggtgcc actgggcacc
     1621 caggcatcat cccaccacat gccactctcg tcttcgatgt ggagcttcta aaactggaag
     1681 aagcttcgaa ttctgcagtc gacacctcta ggaagaaagt gttgctgaag gttatcatcc
     1741 tgggagattc tggagtcggg aagacatcac tcatgaacca gtatgtgaat aagaaattca
     1801 gcaatcagta caaagccaca ataggagctg actttctgac caaggaggtg atggtggatg
     1861 acaggctagt cacaatgcag atatgggaca cagcaggaca ggaacggttc cagtctctcg
     1921 gtgtggcctt ctacagaggt gcagactgct gcgttctggt atttgatgtg actgccccca
     1981 acacattcaa aaccctagat agctggagag atgagtttct catccaggcc agtccccgag
     2041 atcctgaaaa cttcccattt gttgtgttgg gaaacaagat tgacctcgaa aacagacaag
     2101 tggccacaaa gcgggcacag gcctggtgct acagcaaaaa caacattccc tactttgaga
     2161 ccagtgccaa ggaggccatc aacgtggagc aggcgttcca gacgattgca cggaatgcac
     2221 ttaagcagga aacggaggtg gagctgtaca acgaatttcc tgaacctatc aaactggaca
     2281 agaatgaccg ggccaaggcc tcggcagaaa gctgcagttg ctgaggatcc gagctcggta
     2341 ccaagcttaa gtttaaaccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     2401 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     2461 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     2521 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     2581 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     2641 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     2701 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     2761 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     2821 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     2881 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     2941 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3001 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     3061 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     3121 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     3181 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3241 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     3301 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     3361 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     3421 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     3481 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     3541 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     3601 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     3661 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     3721 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     3781 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     3841 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     3901 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     3961 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     4021 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     4081 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     4141 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     4201 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     4261 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     4321 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     4381 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     4441 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     4501 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     4561 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     4621 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4681 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4741 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4801 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4861 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4921 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4981 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     5041 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     5101 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     5161 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     5221 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     5281 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     5341 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     5401 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5461 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5521 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5581 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5641 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5701 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5761 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5821 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5881 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5941 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     6001 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     6061 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     6121 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     6181 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     6241 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     6301 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     6361 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6421 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6481 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6541 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6601 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6661 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6721 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
     6781 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
     6841 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
     6901 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
     6961 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga
     7021 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     7081 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     7141 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     7201 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     7261 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     7321 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     7381 gctattacca tg
//
