LOCUS       sequence-458079-        6589 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing TurboID on the cell surface of mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6589)
  AUTHORS   Wu L, Pandey V, Casha VH, Qu Z, Jami-Alahmadi Y, Gradinaru V, 
            Wohlschlegel JA, Khakh BS
  TITLE     The cell-surface shared proteome of astrocytes and neurons and the 
            molecular foundations of their multicellular interactions.
  JOURNAL   Neuron. 2025 Jun 5:S0896-6273(25)00366-6. doi: 
            10.1016/j.neuron.2025.05.019.
  PUBMED    40499536
REFERENCE   2  (bases 1 to 6589)
  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: "Neuron. 
            2025 Jun 5:S0896-6273(25)00366-6. doi: 
            10.1016/j.neuron.2025.05.019."
COMMENT     Sequence Label: pcDNA3.1-TurboID-TM
FEATURES             Location/Qualifiers
     source          1..6589
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     895..954
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             958..984
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             985..1941
                     /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"
     CDS             1942..1971
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1975..2121
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     promoter        complement(2143..2161)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2144..2161)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2181..2198)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2187..2411
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2457..2885
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2544..2563)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2754..2775
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2894..2914)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2899..3228
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3079..3214
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3141..3160
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3294..4088
                     /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(3348..3367)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3958..3977
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4262..4383
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4299..4318)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4353..4372
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4432..4448)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4432..4448)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4445..4467)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4456..4472
                     /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(join(4480..4486,4487..4504,4505..4510))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4525..4546
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4663..4680)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4834..5422)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4914..4933)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5593..6384,6385..6453))
                     /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(6454..6558)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttatggag
      901 acagacacac tcctgctatg ggtactgctg ctctgggttc caggttccac tggtgactat
      961 ccatatgatg ttccagatta tgctaaagac aatactgtgc ctctgaagct gatcgctctc
     1021 ctggctaatg gcgagttcca tagtggcgaa cagctgggag aaaccctggg catgtccagg
     1081 gccgctatca acaagcacat tcagactctg cgcgactggg gcgtggacgt gttcaccgtg
     1141 cccggaaagg gctactctct gcccgagcct atcccgctgc tgaacgctaa acagattctg
     1201 ggacagctgg acggcgggag cgtggcagtc ctgcctgtgg tcgactccac caatcagtac
     1261 ctgctggatc gaatcggcga gctgaagagt ggggatgctt gcattgcaga atatcagcag
     1321 gcagggagag gaagcagagg gaggaaatgg ttctctcctt ttggagctaa cctgtacctg
     1381 agtatgtttt ggcgcctgaa gcggggacca gcagcaatcg gcctgggccc ggtcatcgga
     1441 attgtcatgg cagaagcgct gcgaaagctg ggagcagaca aggtgcgagt caaatggccc
     1501 aatgacctgt atctgcagga tagaaagctg gcaggcatcc tggtggagct ggccggaata
     1561 acaggcgatg ctgcacagat cgtcattggc gccgggatta acgtggctat gaggcgcgtg
     1621 gaggaaagcg tggtcaatca gggctggatc acactgcagg aagcagggat taacctggac
     1681 aggaatactc tggccgctac gctgatccga gagctgcggg cagccctgga actgttcgag
     1741 caggaaggcc tggctccata tctgccacgg tgggagaagc tggataactt catcaataga
     1801 cccgtgaagc tgatcattgg ggacaaagag attttcggga ttagccgggg gattgataaa
     1861 cagggagccc tgctgctgga acaggacgga gttatcaaac cctggatggg cggagaaatc
     1921 agtctgcggt ctgccgaaaa ggaacaaaaa ctcatctcag aagaggatct gaatgctgtg
     1981 ggccaggaca cgcaggaggt catcgtggtg ccacactcct tgccctttaa ggtggtggtg
     2041 atctcagcca tcctggccct ggtggtgctc accatcatct cccttatcat cctcatcatg
     2101 ctttggcaga agaagccacg ttagtagtct agagggccct attctatagt gtcacctaaa
     2161 tgctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     2221 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     2281 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     2341 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     2401 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     2461 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2521 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2581 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2641 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2701 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2761 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2821 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2881 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     2941 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3001 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3061 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3121 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     3181 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggag
     3241 cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg
     3301 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     3361 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     3421 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg
     3481 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     3541 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     3601 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     3661 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     3721 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     3781 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg
     3841 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     3901 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     3961 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     4021 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     4081 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     4141 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     4201 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     4261 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4321 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4381 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     4441 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4501 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4561 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4621 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4681 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4741 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4801 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4861 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4921 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4981 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5041 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5101 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5161 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5221 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5281 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5341 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5401 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5461 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5521 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5581 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5641 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5701 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5761 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5821 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5881 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5941 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6001 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6061 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6121 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6181 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6241 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6301 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6361 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6421 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6481 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6541 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
