C1(1-29)-TurboID-V5_pCDNA3

LOCUS       C1(1-29)-TurboID-V5_pCDNA3                6564 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  expresses V5-tagged TurboID on the mammalian ER membrane.
ACCESSION   .
VERSION     .
KEYWORDS    C1(1-29)-TurboID-V5_pCDNA3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6564)
  AUTHORS   Branon TC, Bosch JA, Sanchez AD, Udeshi ND, Svinkina T, Carr SA, 
            Feldman JL, Perrimon N, Ting AY
  TITLE     Efficient proximity labeling in living cells and organisms with 
            TurboID.
  JOURNAL   Nat Biotechnol. 2018 Aug 20. pii: nbt.4201. doi: 10.1038/nbt.4201.
  PUBMED    30125270
REFERENCE   2  (bases 1 to 6564)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6564
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        115..494
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        495..698
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     649..669
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     743..762
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        743..761
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      780..789
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             945..1901
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKLG
                     ADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGWI
                     TLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGDK
                     EIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     CDS             1908..1949
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     promoter        complement(1997..2015)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1998..2015)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2035..2052)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2041..2265
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2311..2739
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2398..2417)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2608..2629
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2748..2768)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2753..3082
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2933..3068
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2995..3014
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3149..3943
                     /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(3203..3222)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3813..3832
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4117..4238
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4154..4173)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4208..4227
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4287..4303)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4287..4303)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4300..4322)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4311..4327
                     /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(4335..4365)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4380..4401
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4518..4535)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4689..5277)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4769..4788)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5448..6308)
                     /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     6071..6090
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6309..6413)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6488..6507)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
       61 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      121 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      181 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      241 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      301 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      361 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      421 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      481 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      541 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      601 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      661 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      721 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgggccg
      781 ccaccatgga ccctgtggtc gtgctggggc tgtgcctgag ctgtctgctg ctgctgagcc
      841 tgtggaagca gtcctacgga ggcgggggat caggaagcgg cggccgcgga tcaggaagcg
      901 gatcaggaag cggatcagga agcggatcag gaagcggatc aggaaaagac aatactgtgc
      961 ctctgaagct gatcgctctc ctggctaatg gcgagttcca tagtggcgaa cagctgggag
     1021 aaaccctggg catgtccagg gccgctatca acaagcacat tcagactctg cgcgactggg
     1081 gcgtggacgt gttcaccgtg cccggaaagg gctactctct gcccgagcct atcccgctgc
     1141 tgaacgctaa acagattctg ggacagctgg acggcgggag cgtggcagtc ctgcctgtgg
     1201 tcgactccac caatcagtac ctgctggatc gaatcggcga gctgaagagt ggggatgctt
     1261 gcattgcaga atatcagcag gcagggagag gaagcagagg gaggaaatgg ttctctcctt
     1321 ttggagctaa cctgtacctg agtatgtttt ggcgcctgaa gcggggacca gcagcaatcg
     1381 gcctgggccc ggtcatcgga attgtcatgg cagaagcgct gcgaaagctg ggagcagaca
     1441 aggtgcgagt caaatggccc aatgacctgt atctgcagga tagaaagctg gcaggcatcc
     1501 tggtggagct ggccggaata acaggcgatg ctgcacagat cgtcattggc gccgggatta
     1561 acgtggctat gaggcgcgtg gaggaaagcg tggtcaatca gggctggatc acactgcagg
     1621 aagcagggat taacctggac aggaatactc tggccgctac gctgatccga gagctgcggg
     1681 cagccctgga actgttcgag caggaaggcc tggctccata tctgccacgg tgggagaagc
     1741 tggataactt catcaataga cccgtgaagc tgatcattgg ggacaaagag attttcggga
     1801 ttagccgggg gattgataaa cagggagccc tgctgctgga acaggacgga gttatcaaac
     1861 cctggatggg cggagaaatc agtctgcggt ctgccgaaaa ggctagcggc aagcccatcc
     1921 ccaaccccct gctgggcctg gacagcacct aatgaggatc cggcaagccc atccccaacc
     1981 ccctgctggg ccctattcta tagtgtcacc taaatgctag agctcgctga tcagcctcga
     2041 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     2101 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2161 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2221 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2281 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     2341 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2401 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2461 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2521 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2581 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2641 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2701 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     2761 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2821 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     2881 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     2941 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3001 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     3061 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3121 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3181 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     3241 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     3301 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     3361 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     3421 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     3481 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     3541 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     3601 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     3661 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     3721 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     3781 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     3841 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     3901 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     3961 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4021 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4081 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     4141 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4201 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     4261 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     4321 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     4381 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     4441 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     4501 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     4561 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4621 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4681 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     4741 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4801 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     4861 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     4921 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     4981 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5041 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5101 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     5161 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5221 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     5281 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     5341 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     5401 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     5461 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     5521 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     5581 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     5641 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     5701 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     5761 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     5821 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     5881 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     5941 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6001 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6061 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     6121 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     6181 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     6241 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     6301 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     6361 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     6421 ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca
     6481 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg
     6541 tgttggaggt cgctgagtag tgcg
//