LOCUS       sequence-457348-        7086 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre recombinase-dependent expression of ER TurboID, N-term HA tag, 
            C-term V5 tag, in AAV expression plasmid. Amp selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7086)
  AUTHORS   Wei W, Riley NM, Yang AC, Kim JT, Terrell SM, Li VL, 
            Garcia-Contreras M, Bertozzi CR, Long JZ
  TITLE     Cell type-selective secretome profiling in vivo.
  JOURNAL   Nat Chem Biol. 2021 Mar;17(3):326-334. doi: 
            10.1038/s41589-020-00698-y. Epub 2020 Nov 16.
  PUBMED    33199915
REFERENCE   2  (bases 1 to 7086)
  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"; doi: 
            "10.1038/s41589-020-00698-y"; journalName: "Nat Chem Biol"; date: 
            "2021-03"; volume: "17"; issue: "3"; pages: "326-334"
COMMENT     Sequence Label: pAAV-FLEx-ER-TurboID
FEATURES             Location/Qualifiers
     source          1..7086
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     enhancer        204..583
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          863..1879
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1803..1825
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1887..1906
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1934..1967)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(2018..2051)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2072..2113)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(2120..3076)
                     /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             complement(3083..3109)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     sig_peptide     complement(3113..3172)
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     regulatory      3169..3178
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3179..3212
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3263..3296
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3309..3897
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3362..3382)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3780..3791)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3923..3942)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3988..4043
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     repeat_region   4136..4265
                     /label=AAV2 ITR
     primer_bind     complement(4284..4301)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4284..4300)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4293..4315)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4442..4897
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4529..4548)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4739..4760
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4923..5027
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5028..5096,5097..5888)
                     /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"
     rep_origin      6059..6647
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6548..6567
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6935..6956
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6971..6976,6977..6994,6995..7001)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7009..7025
                     /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     7014..7036
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7033..7049
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7033..7049
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctaccag ggtaatgggg
      181 atcctctaga actatagcta gtcgacattg attattgact agttattaat agtaatcaat
      241 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      301 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      361 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      421 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      481 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      541 tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac
      601 gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat
      661 tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg cgccrggsgg
      721 ggsggggsgg ggsgrggggs ggggsggggs grggsggrgr ggkgcggcgg cagccaatca
      781 gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc ggccctataa
      841 aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc
      901 cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc cacaggtgag
      961 cgggcgggac ggcccttctc ctccgggctg taattagcgc ttggtttaat gacggcttgt
     1021 ttcttttctg tggctgcgtg aaagccttga ggggctccgg gagggccctt tgtgcggggg
     1081 gagcggctcg gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc
     1141 gctgcccggc ggctgtgagc gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt
     1201 gcgcgagggg agcgcggccg ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa
     1261 caaaggctgc gtgcggggtg tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt
     1321 cgggctgcaa ccccccctgc acccccctcc ccgagttgct gagcacggcc cggcttcggg
     1381 tgcggggctc cgtacggggc gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca
     1441 ggkgggggtg ccgggcgggg cggggccgcc ycgggccggg grgggctcgg gggrggggcg
     1501 cggcggcccc cggagcgccg gcggctgtcg aggcgcggcg agccgcagcc attgcctttt
     1561 atggtaatcg tgcgagaggg cgcagggact tcctttgtcc caaatctgtg cggagccgaa
     1621 atctgggagg cgccgccgca ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg
     1681 caggaaggaa atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc cccttctccc
     1741 tctccagcct cggggctgtc cgcgggggga cggctgcctt cgggggggac ggggcagggc
     1801 ggggttcggc ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc atgttcatgc
     1861 cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt
     1921 tggcaaagaa ttcataactt cgtataggat actttatacg aagttatgca gaatggtagc
     1981 tggattgtag ctgctattag caatatgaaa cctcttaata acttcgtata gcatacatta
     2041 tacgaagtta ttgcagttag agttcatctt tggtgctgtc caggcccagc agggggttgg
     2101 ggatgggctt gccctgcagc ttttcggcag accgcagact gatttctccg cccatccagg
     2161 gtttgataac tccgtcctgt tccagcagca gggctccctg tttatcaatc ccccggctaa
     2221 tcccgaaaat ctctttgtcc ccaatgatca gcttcacggg tctattgatg aagttatcca
     2281 gcttctccca ccgtggcaga tatggagcca ggccttcctg ctcgaacagt tccagggctg
     2341 cccgcagctc tcggatcagc gtagcggcca gagtattcct gtccaggtta atccctgctt
     2401 cctgcagtgt gatccagccc tgattgacca cgctttcctc cacgcgcctc atagccacgt
     2461 taatcccggc gccaatgacg atctgtgcag catcgcctgt tattccggcc agctccacca
     2521 ggatgcctgc cagctttcta tcctgcagat acaggtcatt gggccatttg actcgcacct
     2581 tgtctgctcc cagctttcgc agcgcttctg ccatgacaat tccgatgacc gggcccaggc
     2641 cgattgctgc tggtccccgc ttcaggcgcc aaaacatact caggtacagg ttagctccaa
     2701 aaggagagaa ccatttcctc cctctgcttc ctctccctgc ctgctgatat tctgcaatgc
     2761 aagcatcccc actcttcagc tcgccgattc gatccagcag gtactgattg gtggagtcga
     2821 ccacaggcag gactgccacg ctcccgccgt ccagctgtcc cagaatctgt ttagcgttca
     2881 gcagcgggat aggctcgggc agagagtagc cctttccggg cacggtgaac acgtccacgc
     2941 cccagtcgcg cagagtctga atgtgcttgt tgatagcggc cctggacatg cccagggttt
     3001 ctcccagctg ttcgccacta tggaactcgc cattagccag gagagcgatc agcttcagag
     3061 gcacagtatt gtctttgcta gcagcgtaat ctggaacatc gtatgggtag tcaccagtgg
     3121 aacctggaac ccagagcagc agcacccata gcaggagtgt gtctgtctcc atggtggcat
     3181 aacttcgtat aaagtatcct atacgaagtt atttgcctta acccagaaat tatcactgtt
     3241 attctttaga atggtgcaaa gaataacttc gtataatgta tgctatacga agttataagc
     3301 ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     3361 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     3421 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     3481 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     3541 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     3601 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     3661 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
     3721 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     3781 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     3841 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
     3901 gataccgtcg atgaggatcc gatctttttc cctctgccaa aaattatggg gacatcatga
     3961 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
     4021 gttggaattt tttgtgtctc tcactcggaa gcaattcgtt gatctgaatt tcgaccaccc
     4081 ataataccca ttaccctggt agataagtag catggcgggt taatcattaa ctacaaggaa
     4141 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     4201 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     4261 cgcagcctta attaacctaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     4321 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     4381 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     4441 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4501 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4561 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4621 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     4681 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     4741 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     4801 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     4861 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     4921 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     4981 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5041 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5101 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     5161 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     5221 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     5281 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     5341 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     5401 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     5461 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     5521 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     5581 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     5641 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     5701 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     5761 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     5821 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     5881 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     5941 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     6001 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6061 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6121 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     6181 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     6241 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     6301 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     6361 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     6421 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     6481 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     6541 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6601 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6661 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     6721 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     6781 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac
     6841 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
     6901 ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg
     6961 caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat
     7021 aacaatttca cacaggaaac agctatgacc atgattacgc cagatttaat taaggcctta
     7081 attagg
//
