LOCUS       Exported                6386 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  To fuse your protein of interest to the C-terminus of BioID2 and use
            in proximity-dependent biotin identification (BioID); MCS and a 25nm
            GS linker are present downstream of BioID2 and myc tags..
ACCESSION   .
VERSION     .
KEYWORDS    mycBioID2-13X Linker-MCS
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6386)
  AUTHORS   Kim DI, Jensen SC, Noble KA, Kc B, Roux KH, Motamedchaboki K, Roux 
            KJ
  TITLE     An improved smaller biotin ligase for BioID proximity labeling.
  JOURNAL   Mol Biol Cell. 2016 Feb 24. pii: mbc.E15-12-0844.
  PUBMED    26912792
REFERENCE   2  (bases 1 to 6386)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Biol 
            Cell. 2016 Feb 24. pii: mbc.E15-12-0844."
FEATURES             Location/Qualifiers
     source          1..6386
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     polyA_signal    6..230
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      276..704
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(363..382)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     573..594
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(713..733)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        718..1047
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      898..1033
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     960..979
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1114..1908
                     /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(1168..1187)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1778..1797
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2082..2203
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2119..2138)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2173..2192
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2252..2268)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2252..2268)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2265..2287)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2276..2292
                     /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(2300..2330)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2345..2366
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2483..2500)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2654..3242)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2734..2753)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3413..4273)
                     /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     4036..4055
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4274..4378)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4453..4472)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4644..5023
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5024..5227
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5178..5198
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5272..5291
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5272..5290
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      5308..5317
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             5317..5346
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             5350..6045
                     /codon_start=1
                     /product="promiscuous R40G mutant of a biotin protein 
                     ligase from Aquifex aeolicus (Kim et al., 2016)"
                     /label=BioID2
                     /translation="FKNLIWLKEVDSTQERLKEWNVSYGTALVADRQTKGRGGLGRKWL
                     SQEGGLYFSFLLNPKEFENLLQLPLVLGLSVSEALEEITEIPFSLKWPNDVYFQEKKVS
                     GVLCELSKDKLIVGIGINVNQREIPEEIKDRATTLYEITGKDWDRKEVLLKVLKRISEN
                     LKKFKEKSFKEFKGKIESKMLYLGEEVKLLGEGKITGKLVGLSEKGGALILTEEGIKEI
                     LSGEFSLRRS"
     primer_bind     complement(join(6386,1..17))
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"