pcDNA3.1 MCS-BirA(R118G)-HAP

LOCUS       pcDNA3.1 MCS-BirA(R118G)-HAP                6380 bp ds-DNA     circular SYN 05-JAN-2023
DEFINITION  To fuse your protein of interest to the N-terminus of BirA(R118G) 
            and use in proximity-dependent biotin identification (BioID); HA 
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1 MCS-BirA(R118G)-HA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6380)
  AUTHORS   Roux KJ, Kim DI, Raida M, Burke B
  TITLE     A promiscuous biotin ligase fusion protein identifies proximal and 
            interacting proteins in mammalian cells.
  JOURNAL   J Cell Biol. 2012 Mar 12.
  PUBMED    22412018
REFERENCE   2  (bases 1 to 6380)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 5, 2023 from SnapGene Server 1.1.58
            http://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..6380
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(90..109)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        281..660
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        661..864
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     815..835
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     909..928
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        909..927
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             997..1956
                     /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             1960..1986
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2017..2034)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2023..2247
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2293..2721
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2380..2399)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2590..2611
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2730..2750)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2735..3064
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2915..3050
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2977..2996
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3131..3925
                     /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(3185..3204)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3795..3814
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4099..4220
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4136..4155)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4190..4209
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4269..4285)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4269..4285)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4282..4304)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4293..4309
                     /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(4317..4347)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4362..4383
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4500..4517)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4671..5259)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4751..4770)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5430..6290)
                     /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     6053..6072
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6291..15)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga
       61 tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
      121 cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa
      181 gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt
      241 tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt
      301 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      361 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      421 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      481 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      541 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
      601 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
      661 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
      721 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      781 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      841 tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc ttactggctt
      901 atcgaaatta atacgactca ctatagggag acccaagctg gctagcgctt aaggcctgtt
      961 aaccggtcgt acgtctccgg attcgaattc ggatccaagg acaacaccgt gcccctgaag
     1021 ctgatcgccc tgctggccaa cggcgagttc cactctggcg agcagctggg agagaccctg
     1081 ggaatgagca gagccgccat caacaagcac atccagacac tgagagactg gggagtggac
     1141 gtgttcaccg tgcctggcaa gggctacagc ctgcctgagc ctatccagct gctgaacgcc
     1201 aagcagatcc tgggacagct ggatggcgga agcgtggccg tgctgcctgt gatcgactcc
     1261 accaatcagt acctgctgga cagaatcgga gagctgaagt ccggcgacgc ctgcatcgcc
     1321 gagtaccagc aggctggcag aggaggcaga ggacggaagt ggttcagccc attcggagcc
     1381 aacctgtacc tgtccatgtt ctggagactg gagcagggac ctgctgctgc catcggactg
     1441 agtctggtga tcggaatcgt gatggccgag gtgctgagaa agctgggagc cgacaaggtg
     1501 agagtgaagt ggcctaatga cctgtacctc caggaccgca agctggctgg catcctggtg
     1561 gagctgacag gcaagacagg cgatgccgct cagatcgtga tcggagccgg aatcaacatg
     1621 gccatgagaa gagtggagga gagcgtggtg aaccagggct ggatcaccct gcaggaggct
     1681 ggcatcaacc tggaccggaa caccctggcc gccatgctga tcagagagct gagagccgct
     1741 ctggagctgt tcgagcagga gggactggct ccttacctga gcagatggga gaagctggac
     1801 aacttcatca acagacctgt gaagctgatc atcggcgaca aggaaatctt cggcatctcc
     1861 agaggaatcg acaagcaggg agctctgctg ctggagcagg acggaatcat caagccctgg
     1921 atgggcggag aaatctccct gagaagcgca gagaaggctt acccttacga tgtaccggat
     1981 tacgcatagg cggcctaagt ttaaaccgct gatcagcctc gactgtgcct tctagttgcc
     2041 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     2101 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     2161 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     2221 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     2281 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     2341 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     2401 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     2461 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     2521 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     2581 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     2641 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     2701 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     2761 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     2821 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2881 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     2941 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     3001 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     3061 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg
     3121 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     3181 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     3241 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     3301 gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     3361 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     3421 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     3481 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     3541 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     3601 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg
     3661 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3721 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3781 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3841 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     3901 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3961 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     4021 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     4081 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     4141 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     4201 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     4261 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4321 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4381 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4441 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4501 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4561 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4621 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4681 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4741 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4801 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4861 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4921 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4981 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5041 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5101 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5161 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5221 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5281 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5341 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5401 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5461 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5521 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5581 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5641 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5701 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5761 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5821 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5881 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5941 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6001 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6061 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6121 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6181 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6241 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6301 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6361 tgtatttaga aaaataaaca
//