LOCUS       sequence-456960-        7101 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology repair plasmid for endogenous tagging of AP1muA at the 
            C-terminus with TurboID and a V5 epitope tag. Contains a G418 
            resistance cassette for selection of edited cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7101)
  AUTHORS   Stockhammer A, Spalt C, Klemt A, Benz LS, Harel S, Natalia V, Wiench
            L, Freund C, Kuropka B, Bottanelli F
  TITLE     When less is more - a fast TurboID knock-in approach for 
            high-sensitivity endogenous interactome mapping.
  JOURNAL   J Cell Sci. 2024 Aug 15;137(16):jcs261952. doi: 10.1242/jcs.261952. 
            Epub 2024 Aug 28.
  PUBMED    39056144
REFERENCE   2  (bases 1 to 7101)
  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: "J Cell 
            Sci."; date: "2024-08-15"; volume: "137(16):jcs261952. doi"; pages: 
            " 10.1242/jcs.261952. Epub 2024 Aug 28"
COMMENT     Sequence Label: AP1muA-TurboID-polyA-G418 HR
FEATURES             Location/Qualifiers
     source          1..7101
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    71..192
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(108..127)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     162..181
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        218..575
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        218..409
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(241..261)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      426..561
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             610..1404
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             610..1404
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(664..683)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1274..1293
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1592..1611)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     primer_bind     complement(2738..2754)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2738..2754)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2751..2773)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(2926..3514)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3006..3025)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(3631..3655)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(3755..4546,4547..4615))
                     /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"
     promoter        complement(4616..4720)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4819..4841
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4833..4850
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4834..4850
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             6044..7003
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     CDS             7004..7045
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
ORIGIN
        1 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat
       61 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
      121 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
      181 caatgtatct taaggcgtaa attgtaaggc ggccgccctg aggcggaaag aaccagctgt
      241 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
      301 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
      361 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
      421 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
      481 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
      541 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
      601 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
      661 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
      721 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
      781 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
      841 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
      901 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
      961 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     1021 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     1081 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     1141 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     1201 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     1261 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     1321 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     1381 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     1441 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     1501 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     1561 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     1621 gaacccgcgc tatgacggca ataaaaagac agaataaaga attcggggct gtcgcagcca
     1681 acaccccagc ctcggagctc ctggtggcag caccagggga cacacctgcc aaacccacca
     1741 gatggagggg ccctccctgg tctctggcca ccctcccagc ctctgcccag ggacccctgc
     1801 cttccccagg ccatctgctc tgccgtcgac actcgtctca gaagcccctt tcccagaaga
     1861 ggctggtctt caagaagtct cgtttctttg cccctgaagt cagtttcagg ggaaggatgt
     1921 gaaatttttc cgtgtagagg ttacagcctt ttatgctgtt gagctcccag gtaccaaaaa
     1981 gcttggccaa cgcttgccag ccagccagct gcaggtggca tctgccacga aggaagcgcc
     2041 agcctcgcca ggccagcagg ggcgtcgttt tgttgccatt ttgttgaacg ttatgggttt
     2101 atgggtgttc ctggaacttg tctttgtgca ttcgttgctg tttgtgttac cctcactgtc
     2161 cccatgtccc acccacgtcc tacggcactc aggaagcact tggtgaggac gagccctcac
     2221 ccttcttgtc ttccttcccg gcagcgcccg cagcgggcca tttacacgtc gaggctggca
     2281 cctggcgcgc tcgggggcca ctgtagcgtc tgcctgctcc ctggactcgc aggcctcgcc
     2341 tgtggcgcct tcccagggcc agcctgggtc acgagatgct gtcactcagc cagatcagta
     2401 ttgacccacc aggggaggtg gggtttggtg agagacgcca gcctcagact ttttcccact
     2461 gagggtccag agagcggggc cacgtgtcac ccacgtctgc gcttggtcac ccgtcctccc
     2521 caccctgtgt gtgtttatgt catagttaca ttaaattcca ttcattgaat actccacagc
     2581 cttcctctga tgcagcgtgg tcccaactcc tcctgcgtga ccacagggct tgcctggttt
     2641 tgtcgccgac cttccctgtg gccccacgag gtgtcaccag caggctaggt ggaggctcag
     2701 tgatgataag tctgcgatgg tggatgcatg tgtcatggtc atagctgttt cctgtgtgaa
     2761 attgttatcc gctcagaggg cacaatccta ttccgcgcta tccgacaatc tccaagacat
     2821 taggtggagt tcagttcggc gtatggcata tgtcgctgga aagaacatgt gagcaaaagg
     2881 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     2941 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     3001 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3061 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3121 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3181 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3241 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     3301 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     3361 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     3421 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     3481 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     3541 gtctgacgct ctattcaaca aagccgccgt cccgtcaagt cagcgtaaat gggtaggggg
     3601 cttcaaatcg tcctcgtgat accaattcgg agcctgcttt tttgtacaaa cttgttgata
     3661 atggcaattc aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     3721 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     3781 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     3841 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagagc
     3901 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     3961 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     4021 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     4081 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     4141 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     4201 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     4261 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     4321 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     4381 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     4441 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     4501 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     4561 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     4621 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     4681 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     4741 cagatacctg aaacaaaacc catcgtacgg ccaaggaagt ctccaataac tgtgatccac
     4801 cacaagcgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt catgcataat
     4861 ccgcacgcat ctggaataag gaagtgccat tccgcctgac cttggggcag cacagggccg
     4921 agctttggct ccccagcaca ggggctcatg ctcccctccc tggactgggg tgagtggtca
     4981 gtctctgccc atcgccacta gggccagagc tgttggctaa gctggggcag ggcctaggcc
     5041 tgagcgcctc ccccgtctgc tccccagggc ggcaaggagt acctgatgcg ggcccacttc
     5101 ggcctgccta gtgtggaggc cgaagacaag gagggcaagc ccccgatcag tgtcaagttc
     5161 gagatccctt acttcactac ctccggcatc caggtacgta aggcccaggc ggccggggcc
     5221 cagaacaggg acagaggccg caggtgacag atatcaatgc agccagcttg ggaaagggtg
     5281 tcagtcagct gcaatcagga ccctgctgtg ctgtgggcct ctgcctcccc acagacaggg
     5341 tgaggccagg ctgcctctgc tcagccctga cccctgagct caaagctcac tcattctgcc
     5401 ccaaactagc atttgaggct ctgccgtcca ctcatgcccc atgggacctc ttttccctgg
     5461 caccccctca aaagcccaca gagggaggag aggctcccag agagccaggg gccttctggc
     5521 cgggcccccc aagcaggctg ctgaggccgt gtctgcaagc cacatcctgg ctgctcacca
     5581 cttccagagg cggctctgcc ctcccacaca tttgcatggt ccagacaaat ccctttcacc
     5641 cccctgaggc ctgtgctcat cctgtcgtca tggcctcccc tgccagcccc aggctgcccc
     5701 agcaggaaag attaaggggg gaccaacagg gcgggagcct gcggtgctca gggccctgct
     5761 acctgtgccc caggtgcgct acctgaagat cattgagaag agtgggtacc aggccctgcc
     5821 ctgggtgcgt tatatcacgc agaatggagg tgagtgaggc cctacccgtg gggtggggtt
     5881 gtactgaggg gtcctctgtg gctgccccca gggtggagcc atcgggtcgg gtcccgaaag
     5941 cagggagggt gcagagccca gcatgacgcc tccctcctgt ctcctagatt accagctccg
     6001 gacccaggga tcagggtcag gctctggttc cggctcagga tccatgaaag acaatactgt
     6061 gcctctgaag ctgatcgctc tcctggctaa tggcgagttc catagtggcg aacagctggg
     6121 agaaaccctg ggcatgtcca gggccgctat caacaagcac attcagactc tgcgcgactg
     6181 gggcgtggac gtgttcaccg tgcccggaaa gggctactct ctgcccgagc ctatcccgct
     6241 gctgaacgct aaacagattc tgggacagct ggacggcggg agcgtggcag tcctgcctgt
     6301 ggtcgactcc accaatcagt acctgctgga tcgaatcggc gagctgaaga gtggggatgc
     6361 ttgcattgca gaatatcagc aggcagggag aggaagcaga gggaggaaat ggttctctcc
     6421 ttttggagct aacctgtacc tgagtatgtt ttggcgcctg aagcggggac cagcagcaat
     6481 cggcctgggc ccggtcatcg gaattgtcat ggcagaagcg ctgcgaaagc tgggagcaga
     6541 caaggtgcga gtcaaatggc ccaatgacct gtatctgcag gatagaaagc tggcaggcat
     6601 cctggtggag ctggccggaa taacaggcga tgctgcacag atcgtcattg gcgccgggat
     6661 taacgtggct atgaggcgcg tggaggaaag cgtggtcaat cagggctgga tcacactgca
     6721 ggaagcaggg attaacctgg acaggaatac tctggccgct acgctgatcc gagagctgcg
     6781 ggcagccctg gaactgttcg agcaggaagg cctggctcca tatctgccac ggtgggagaa
     6841 gctggataac ttcatcaata gacccgtgaa gctgatcatt ggggacaaag agattttcgg
     6901 gattagccgg gggattgata aacagggagc cctgctgctg gaacaggacg gagttatcaa
     6961 accctggatg ggcggagaaa tcagtctgcg gtctgccgaa aagggcaagc ccatccccaa
     7021 ccccctgctg ggcctggaca gcacctgagc tagcccgcga ctctagatca taatcagcca
     7081 taccacattt gtagaggttt t
//
