LOCUS       sequence-300204-        8325 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses high-affinity split-TurboID N-terminal fragment in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8325)
  AUTHORS   Cho KF, Branon TC, Rajeev S, Svinkina T, Udeshi ND, Thoudam T, Kwak 
            C, Rhee HW, Lee IK, Carr SA, Ting AY
  TITLE     Split-TurboID enables contact-dependent proximity labeling in cells.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 May 18. pii: 1919528117. doi: 
            10.1073/pnas.1919528117.
  PUBMED    32424107
REFERENCE   2  (bases 1 to 8325)
  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: "Proc Natl 
            Acad Sci U S A. 2020 May 18. pii: 1919528117. doi: 
            10.1073/pnas.1919528117."
COMMENT     Sequence Label: pLX304 CMV FKBP-V5-sTurboID-high affinity (N)
FEATURES             Location/Qualifiers
     source          1..8325
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        65..368
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        369..572
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     523..543
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    595..619
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             985..1026
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     misc_feature    1332..1920
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1385..1405)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1803..1814)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             1992..2225
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2303..2424
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2340..2359)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2394..2413
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2464..2599
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2619..2638)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2620..2638)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2648..2665)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2648..2664)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2657..2679)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2806..3261
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2893..2912)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3103..3124
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3287..3391
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3392..3460,3461..4252)
                     /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     complement(3610..3629)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4423..5011
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4912..4931
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5165..5182
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5299..5320
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5335..5340,5341..5358,5359..5365)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5373..5389
                     /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)."
     primer_bind     5378..5400
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5397..5413
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5397..5413
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5432..5452
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5434..5452
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5480..5706
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5707..5887
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5934..6059
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6552..6785
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6970..7014
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        7189..7689
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     7608..7626
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             7701..8096
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    8156..8273
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 cccggggtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
       61 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
      121 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      181 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      241 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
      301 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      361 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
      421 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
      481 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
      541 gtgtacggtg ggaggtctat ataagcagag ctctctggct aactgtcggg atcaacaagt
      601 ttgtacaaaa aagcaggctt cgaaggagat agaaccatgc aggtggaaac catctcccca
      661 ggagacgggc gcaccttccc caagcgcggc cagacctgcg tggtgcacta caccgggatg
      721 cttgaagatg gaaagaaatt tgattcctcc cgggacagaa acaagccctt taagtttatg
      781 ctaggcaagc aggaggtgat ccgaggctgg gaagaagggg ttgcccagat gagtgtgggt
      841 cagagagcca aactgactat atctccagat tatgcctatg gtgccactgg gcacccaggc
      901 atcatcccac cacatgccac tctcgtcttc gatgtggagc ttctaaaact ggaagggagt
      961 ggcagcggag gtagtgggag cggtggcaag cccatcccca accccctgct gggcctggac
     1021 agcaccaagg gctcgggctc gacctcgggc tcgggcaccg gtaaagacaa tactgtgcct
     1081 ctgaagctga tcgctctcct ggctaatggc gagttccata gtggcgaaca gctgggagaa
     1141 accctgggca tgtccagggc cgctatcaac aagcacattc agactctgcg cgactggggc
     1201 gtggacgtgt tcaccgtgcc cggaaagggc tactctctgc ccgagcctat cccgctgctg
     1261 aacgctaaac agattctggg acagctggac ggctaataag ctagcgctaa ccggtggcgc
     1321 gttaagtcga caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     1381 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     1441 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     1501 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     1561 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     1621 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     1681 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     1741 cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     1801 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     1861 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     1921 gtcgacttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
     1981 aaagggggga ctggaagggc taattcactc ccaacgaaga caagatctgc tttttgcttg
     2041 tactgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     2101 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     2161 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     2221 tagcagtacg tatagtagtt catgtcatct tattattcag tatttataac ttgcaaagaa
     2281 atgaatatca gagagtgaga ggaacttgtt tattgcagct tataatggtt acaaataaag
     2341 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     2401 gtccaaactc atcaatgtat cttatcatgt ctggctctag ctatcccgcc cctaactccg
     2461 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     2521 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     2581 ggaggctttt ttggaggcct agggacgtac ccaattcgcc ctatagtgag tcgtattacg
     2641 cgcgctcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     2701 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     2761 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg
     2821 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2881 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2941 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     3001 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     3061 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     3121 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     3181 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     3241 aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3301 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3361 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     3421 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     3481 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3541 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     3601 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     3661 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     3721 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     3781 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     3841 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     3901 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     3961 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     4021 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     4081 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     4141 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     4201 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     4261 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4321 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4381 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4441 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4501 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4561 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4621 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4681 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4741 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4801 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4861 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4921 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4981 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5041 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     5101 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     5161 gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc
     5221 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
     5281 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
     5341 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
     5401 aaacagctat gaccatgatt acgccaagcg cgcaattaac cctcactaaa gggaacaaaa
     5461 gctggagctg caagcttaat gtagtcttat gcaatactct tgtagtcttg caacatggta
     5521 acgatgagtt agcaacatgc cttacaagga gagaaaaagc accgtgcatg ccgattggtg
     5581 gaagtaaggt ggtacgatcg tgccttatta ggaaggcaac agacgggtct gacatggatt
     5641 ggacgaacca ctgaattgcc gcattgcaga gatattgtat ttaagtgcct agctcgatac
     5701 ataaacgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
     5761 aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt
     5821 ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc
     5881 tctagcagtg gcgcccgaac agggacttga aagcgaaagg gaaaccagag gagctctctc
     5941 gacgcaggac tcggcttgct gaagcgcgca cggcaagagg cgaggggcgg cgactggtga
     6001 gtacgccaaa aattttgact agcggaggct agaaggagag agatgggtgc gagagcgtca
     6061 gtattaagcg ggggagaatt agatcgcgat gggaaaaaat tcggttaagg ccagggggaa
     6121 agaaaaaata taaattaaaa catatagtat gggcaagcag ggagctagaa cgattcgcag
     6181 ttaatcctgg cctgttagaa acatcagaag gctgtagaca aatactggga cagctacaac
     6241 catcccttca gacaggatca gaagaactta gatcattata taatacagta gcaaccctct
     6301 attgtgtgca tcaaaggata gagataaaag acaccaagga agctttagac aagatagagg
     6361 aagagcaaaa caaaagtaag accaccgcac agcaagcggc cgctgatctt cagacctgga
     6421 ggaggagata tgagggacaa ttggagaagt gaattatata aatataaagt agtaaaaatt
     6481 gaaccattag gagtagcacc caccaaggca aagagaagag tggtgcagag agaaaaaaga
     6541 gcagtgggaa taggagcttt gttccttggg ttcttgggag cagcaggaag cactatgggc
     6601 gcagcgtcaa tgacgctgac ggtacaggcc agacaattat tgtctggtat agtgcagcag
     6661 cagaacaatt tgctgagggc tattgaggcg caacagcatc tgttgcaact cacagtctgg
     6721 ggcatcaagc agctccaggc aagaatcctg gctgtggaaa gatacctaaa ggatcaacag
     6781 ctcctgggga tttggggttg ctctggaaaa ctcatttgca ccactgctgt gccttggaat
     6841 gctagttgga gtaataaatc tctggaacag atttggaatc acacgacctg gatggagtgg
     6901 gacagagaaa ttaacaatta cacaagctta atacactcct taattgaaga atcgcaaaac
     6961 cagcaagaaa agaatgaaca agaattattg gaattagata aatgggcaag tttgtggaat
     7021 tggtttaaca taacaaattg gctgtggtat ataaaattat tcataatgat agtaggaggc
     7081 ttggtaggtt taagaatagt ttttgctgta ctttctatag tgaatagagt taggcaggga
     7141 tattcaccat tatcgtttca gacccacctc ccaaccccga ggggaccctt gcgccttttc
     7201 caaggcagcc ctgggtttgc gcagggacgc ggctgctctg ggcgtggttc cgggaaacgc
     7261 agcggcgccg accctgggtc tcgcacattc ttcacgtccg ttcgcagcgt cacccggatc
     7321 ttcgccgcta cccttgtggg ccccccggcg acgcttcctg ctccgcccct aagtcgggaa
     7381 ggttccttgc ggttcgcggc gtgccggacg tgacaaacgg aagccgcacg tctcactagt
     7441 accctcgcag acggacagcg ccagggagca atggcagcgc gccgaccgcg atgggctgtg
     7501 gccaatagcg gctgctcagc agggcgcgcc gagagcagcg gccgggaagg ggcggtgcgg
     7561 gaggcggggt gtggggcggt agtgtgggcc ctgttcctgc ccgcgcggtg ttccgcattc
     7621 tgcaagcctc cggagcgcac gtcggcagtc ggctccctcg ttgaccgaat caccgacctc
     7681 tctccccagg gggtaccacc atggccaagc ctttgtctca agaagaatcc accctcattg
     7741 aaagagcaac ggctacaatc aacagcatcc ccatctctga agactacagc gtcgccagcg
     7801 cagctctctc tagcgacggc cgcatcttca ctggtgtcaa tgtatatcat tttactgggg
     7861 gaccttgtgc agaactcgtg gtgctgggca ctgctgctgc tgcggcagct ggcaacctga
     7921 cttgtatcgt cgcgatcgga aatgagaaca ggggcatctt gagcccctgc ggacggtgcc
     7981 gacaggtgct tctcgatctg catcctggga tcaaagccat agtgaaggac agtgatggac
     8041 agccgacggc agttgggatt cgtgaattgc tgccctctgg ttatgtgtgg gagggcctgc
     8101 agctgcagta gtaagaattc tagatcttga gacaaatggc agtattcatc cacaatttta
     8161 aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa
     8221 cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt
     8281 attacaggga cagcagagat ccactttggc gccggctcga ggggg
//
