LOCUS       sequence-308667-        7603 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For doxycycline-inducible expression of 3xHA-TurboID on N-terminus 
            of bait protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7603)
  AUTHORS   May DG, Scott KL, Campos AR, Roux KJ
  TITLE     Comparative Application of BioID and TurboID for Protein-Proximity 
            Biotinylation.
  JOURNAL   Cells. 2020 Apr 25;9(5). pii: cells9051070. doi: 
            10.3390/cells9051070.
  PUBMED    32344865
REFERENCE   2  (bases 1 to 7603)
  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: "Cells."; 
            date: "2020-04-25"; volume: "9(5). pii: cells9051070. doi"; pages: "
            10.3390/cells9051070"
COMMENT     Sequence Label: 3xHA-TurboID pRetrox
FEATURES             Location/Qualifiers
     source          1..7603
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             38..454
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     384..406
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     422..438
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        490..804
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    498..516
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    534..552
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    569..587
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    605..623
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    641..659
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    676..694
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    712..730
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     777..801
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             827..853
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             857..883
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             887..913
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             920..1876
                     /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"
     promoter        1896..2395
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1931..1954)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2328..2347
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2416..3015
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2416..2435)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2912..2932
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3196..3621
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(3724..3746)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3882..3902)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3887..4216
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4067..4202
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4238..4257
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4338..4355)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4509..5097)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4589..4608)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5268..6059,6060..6128))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6129..6233)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6301..6319
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        6419..6798
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6799..7002
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             7003..7178
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    7241..7440
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
ORIGIN
        1 ctgtctgact gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt
       61 aagtttgacc ttaggtcact ggaaagatgt cgagcggatc gctcacaacc agtcggtaga
      121 tgtcaagaag agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg
      181 atggccgcga gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt
      241 ttcacctggc ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt
      301 ggcttttgac ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct
      361 tcctccatcc gccccgtctc tcccccttga acctcctcgt tcgaccccgc ctcgatcctc
      421 cctttatcca gccctcactc cttctctagg cgccggaatt gaagatctga ggccctttcg
      481 tcttcactcg agtttactcc ctatcagtga tagagaacgt atgtcgagtt tactccctat
      541 cagtgataga gaacgatgtc gagtttactc cctatcagtg atagagaacg tatgtcgagt
      601 ttactcccta tcagtgatag agaacgtatg tcgagtttac tccctatcag tgatagagaa
      661 cgtatgtcga gtttatccct atcagtgata gagaacgtat gtcgagttta ctccctatca
      721 gtgatagaga acgtatgtcg aggtaggcgt gtacggtggg aggcctatat aagcagagct
      781 cgtttagtga accgtcagat cgcctggaga aggatccgcc accatgtacc cgtatgatgt
      841 tccggattac gctggctatc cctacgacgt gcccgactat gccgggtacc cctatgacgt
      901 cccagactac gcagctagca aagacaatac tgtgcctctg aagctgatcg ctctcctggc
      961 taatggcgag ttccatagtg gcgaacagct gggagaaacc ctgggcatgt ccagggccgc
     1021 tatcaacaag cacattcaga ctctgcgcga ctggggcgtg gacgtgttca ccgtgcccgg
     1081 aaagggctac tctctgcccg agcctatccc gctgctgaac gctaaacaga ttctgggaca
     1141 gctggacggc gggagcgtgg cagtcctgcc tgtggtcgac tccaccaatc agtacctgct
     1201 ggatcgaatc ggcgagctga agagtgggga tgcttgcatt gcagaatatc agcaggcagg
     1261 gagaggaagc agagggagga aatggttctc tccttttgga gctaacctgt acctgagtat
     1321 gttttggcgc ctgaagcggg gaccagcagc aatcggcctg ggcccggtca tcggaattgt
     1381 catggcagaa gcgctgcgaa agctgggagc agacaaggtg cgagtcaaat ggcccaatga
     1441 cctgtatctg caggatagaa agctggcagg catcctggtg gagctggccg gaataacagg
     1501 cgatgctgca cagatcgtca ttggcgccgg gattaacgtg gctatgaggc gcgtggagga
     1561 aagcgtggtc aatcagggct ggatcacact gcaggaagca gggattaacc tggacaggaa
     1621 tactctggcc gctacgctga tccgagagct gcgggcagcc ctggaactgt tcgagcagga
     1681 aggcctggct ccatatctgc cacggtggga gaagctggat aacttcatca atagacccgt
     1741 gaagctgatc attggggaca aagagatttt cgggattagc cgggggattg ataaacaggg
     1801 agccctgctg ctggaacagg acggagttat caaaccctgg atgggcggag aaatcagtct
     1861 gcggtctgcc gaaaaggccg gctaggaatt ctaccgggta ggggaggcgc ttttcccaag
     1921 gcagtctgga gcatgcgctt tagcagcccc gctgggcact tggcgctaca caagtggcct
     1981 ctggcctcgc acacattcca catccaccgg taggcgccaa ccggctccgt tctttggtgg
     2041 ccccttcgcg ccaccttcta ctcctcccct agtcaggaag ttcccccccg ccccgcagct
     2101 cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc actagtctcg tgcagatgga
     2161 cagcaccgct gagcaatgga agcgggtagg cctttggggc agcggccaat agcagctttg
     2221 ctccttcgct ttctgggctc agaggctggg aaggggtggg tccgggggcg ggctcagggg
     2281 cgggctcagg ggcggggcgg gcgcccgaag gtcctccgga ggcccggcat tctgcacgct
     2341 tcaaaagcgc acgtctgccg cgctgttctc ctcttcctca tctccgggcc tttcgacctg
     2401 cagcccaagc ttaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     2461 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     2521 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     2581 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     2641 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     2701 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     2761 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     2821 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     2881 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     2941 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     3001 aagcccggtg cctgacgccc gccccacgac ccgcagcgcc cgaccgaaag gagcgcacga
     3061 ccccatgcat cggcgtctcg agatatcagt ggtccaggct ctagttttga ctcaacaata
     3121 tcaccagctg aagcctatag agtacgagcc atagataaaa taaaagattt tatttagtct
     3181 ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag cttaagtaac
     3241 gccattttgc aaggcatgga aaaatacata actgagaata gagaagttca gatcaaggtc
     3301 aggaacagat ggaacagggt cgaccctaga gaaccatcag atgtttccag ggtgccccaa
     3361 ggacctgaaa tgaccctgtg ccttatttga actaaccaat cagttcgctt ctcgcttctg
     3421 ttcgcgcgct tctgctcccc gagctcaata aaagagccca caacccctca ctcggggcgc
     3481 cagtcctccg attgactgag tcgcccgggt acccgtgtat ccaataaacc ctcttgcagt
     3541 tgcatccgac ttgtggtctc gctgttcctt gggagggtct cctctgagtg attgactacc
     3601 cgtcagcggg ggtctttcat ttgggggctc gtccgggatc gggagacccc tgcccaggga
     3661 ccaccgaccc accaccggga ggtaagctgg ctgcctcgcg cgtttcggtg atgacggtga
     3721 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     3781 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     3841 gacccagtca cgtagcgata gcggagtgta gatccggctg tggaatgtgt gtcagttagg
     3901 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     3961 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     4021 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     4081 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     4141 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     4201 cctaggcttt tgcaaaaagc ttactggctt aactatgcgg catcagagca gattgtactg
     4261 agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc
     4321 aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     4381 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4441 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4501 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     4561 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4621 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     4681 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     4741 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     4801 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     4861 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     4921 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     4981 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5041 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     5101 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     5161 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     5221 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     5281 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     5341 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     5401 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     5461 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     5521 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     5581 gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     5641 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     5701 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     5761 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     5821 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     5881 tcaacacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     5941 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     6001 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     6061 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     6121 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     6181 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     6241 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
     6301 aataggcgta tcacgaggcc ctttcgtctt caagaattag cttggccatt gcatacgttg
     6361 tatccatatc ataatatgta catttatatt ggctcatgtc caacattacc gccatgttga
     6421 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
     6481 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
     6541 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
     6601 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
     6661 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
     6721 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
     6781 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
     6841 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
     6901 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
     6961 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcaataaaa gagcccacaa
     7021 cccctcactc ggcgcgccag tcttccgata gactgcgtcg cccgggtacc cgtattccca
     7081 ataaagcctc ttgctgtttg catccgaatc gtggtctcgc tgttccttgg gagggtctcc
     7141 tctgagtgat tgactaccca cgacgggggt ctttcatttg ggggctcgtc cgggatttgg
     7201 agacccctgc ccagggacca ccgacccacc accgggaggt aagctggcca gcaacttatc
     7261 tgtgtctgtc cgattgtcta gtgtctatgt ttgatgttat gcgcctgcgt ctgtactagt
     7321 tagctaacta gctctgtatc tggcggaccc gtggtggaac tgacgagttc tgaacacccg
     7381 gccgcaaccc tgggagacgt cccagggact ttgggggccg tttttgtggc ccgacctgag
     7441 gaagggagtc gatgtggaat ccgaccccgt caggatatgt ggttctggta ggagacgaga
     7501 acctaaaaca gttcccgcct ccgtctgaat ttttgctttc ggtttggaac cgaagccgcg
     7561 cgtcttgtct gctgcagcgc tgcagcatcg ttctgtgttg tct
//
