LOCUS       sequence-357575-        7635 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Ezrin fused with BioID2 at the C-terminus in astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7635)
  AUTHORS   Soto JS, Jami-Alahmadi Y, Chacon J, Moye SL, Diaz-Castro B, 
            Wohlschlegel JA, Khakh BS
  TITLE     Astrocyte-neuron subproteomes and obsessive-compulsive disorder 
            mechanisms.
  JOURNAL   Nature. 2023 Apr;616(7958):764-773. doi: 10.1038/s41586-023-05927-7.
            Epub 2023 Apr 12.
  PUBMED    37046092
REFERENCE   2  (bases 1 to 7635)
  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/s41586-023-05927-7"; journalName: "Nature"; date: 
            "2023-04"; volume: "616"; issue: "7958"; pages: "764-773"
COMMENT     Sequence Label: pZac2.1 gfaABC1D-Ezr-BioID2-HA
FEATURES             Location/Qualifiers
     source          1..7635
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   259..388
                     /label=AAV2 ITR
     intron          1239..1371
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1354..1373
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1416..1434
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3262..3960
                     /codon_start=1
                     /product="promiscuous R40G mutant of a biotin protein 
                     ligase from Aquifex aeolicus (Kim et al., 2016)"
                     /label=BioID2
                     /translation="MFKNLIWLKEVDSTQERLKEWNVSYGTALVADRQTKGRGGLGRKW
                     LSQEGGLYFSFLLNPKEFENLLQLPLVLGLSVSEALEEITEIPFSLKWPNDVYFQEKKV
                     SGVLCELSKDKLIVGIGINVNQREIPEEIKDRATTLYEITGKDWDRKEVLLKVLKRISE
                     NLKKFKEKSFKEFKGKIESKMLYLGEEVKLLGEGKITGKLVGLSEKGGALILTEEGIKE
                     ILSGEFSLRRS"
     CDS             3970..3996
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    4023..4611
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4076..4096)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4494..4505)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4660..4781
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4671..4690)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4725..4744
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4943..5072
                     /label=AAV2 ITR
     primer_bind     complement(5091..5108)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5091..5107)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5100..5122)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5249..5704
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5336..5355)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5546..5567
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5730..5834
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5835..5903,5904..6695)
                     /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      6866..7454
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7355..7374
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccagattta
      241 attaaggcct taattaggct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc
      301 gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga
      361 gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta
      421 tctacgtagc catgctctag gaagatctaa catatcctgg tgtggagtag gggacgctgc
      481 tctgacagag gctcgggggc ctgagctggc tctgtgagct ggggaggagg cagacagcca
      541 ggccttgtct gcaagcagac ctggcagcat tgggctggcc gccccccagg gcctcctctt
      601 catgcccagt gaatgactca ccttggcaca gacacaatgt tcggggtggg cacagtgcct
      661 gcttcccgcc gcaccccagc ccccctcaaa tgccttccga gaagcccatt gagcaggggg
      721 cttgcattgc accccagcct gacagcctgg catcttggga taaaagcagc acagccccct
      781 aggggctgcc cttgctgtgt ggcgccaccg gcggtggaga acaaggctct attcagcctg
      841 tgcccaggaa aggggatcag gggatgccca ggcatggaca gtgggtggca gggggggaga
      901 ggagggctgt ctgcttccca gaagtccaag gacacaaatg ggtgagggga gagctctccc
      961 catagctggg ctgcggccca accccacccc ctcaggctat gccagggggt gttgccaggg
     1021 gcacccgggc atcgccagtc tagcccactc cttcataaag ccctcgcatc ccaggagcga
     1081 gcagagccag agcaggttgg agaggagacg catcacctcc gctgctcgca agctttattg
     1141 cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca acagtctcga
     1201 acttaagctg cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag gttacaagac
     1261 aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct
     1321 gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca ggtgtccact
     1381 cccagttcaa ttacagctct taaggctaga gtacttaata cgactcacta taggctagcc
     1441 tcgagctcaa gcttcgaatt caccaccatg ccgaaaccaa tcaatgtccg agttaccacc
     1501 atggatgcag agctggagtt tgcaatccag ccaaatacaa ctggaaaaca gctttttgat
     1561 caggtggtaa agactatcgg cctccgggaa gtgtggtact ttggcctcca ctatgtggat
     1621 aataaaggat ttcctacctg gctgaagctg gataagaagg tgtctgccca ggaggtcagg
     1681 aaggagaatc ccctccagtt caagttccgg gccaagttct accctgaaga tgtggctgag
     1741 gagctcatcc aggacatcac ccagaaactt ttcttcctcc aagtgaagga aggaatcctt
     1801 agcgatgaga tctactgccc ccctgagact gccgtgctct tggggtccta cgctgtgcag
     1861 gccaagtttg gggactacaa caaagaagtg cacaagtctg ggtacctcag ctctgagcgg
     1921 ctgatccctc aaagagtgat ggaccagcac aaacttacca gggaccagtg ggaggaccgg
     1981 atccaggtgt ggcatgcgga acaccgtggg atgctcaaag ataatgctat gttggaatac
     2041 ctgaagattg ctcaggacct ggaaatgtat ggaatcaact atttcgagat aaaaaacaag
     2101 aaaggaacag acctttggct tggagttgat gcccttggac tgaatattta tgagaaagat
     2161 gataagttaa ccccaaagat tggctttcct tggagtgaaa tcaggaacat ctctttcaat
     2221 gacaaaaagt ttgtcattaa acccatcgac aagaaggcac ctgactttgt gttttatgcc
     2281 ccacgtctga gaatcaacaa gcggatcctg cagctctgca tgggcaacca tgagttgtat
     2341 atgcgccgca ggaagcctga caccatcgag gtgcagcaga tgaaggccca ggcccgggag
     2401 gagaagcatc agaagcagct ggagcggcaa cagctggaaa cagagaagaa aaggagagaa
     2461 accgtggaga gagagaaaga gcagatgatg cgcgagaagg aggagttgat gctgcggctg
     2521 caggactatg aggagaagac aaagaaggca gagagagagc tctcggagca gattcagagg
     2581 gccctgcagc tggaggagga gaggaagcgg gcacaggagg aggccgagcg cctagaggct
     2641 gaccgtatgg ctgcactgcg ggctaaggag gagctggaga gacaggcggt ggatcagata
     2701 aagagccagg agcagctggc tgcggagctt gcagaataca ctgccaagat tgccctcctg
     2761 gaagaggcgc ggaggcgcaa ggaggatgaa gttgaagagt ggcagcacag ggccaaagaa
     2821 gcccaggatg acctggtgaa gaccaaggag gagctgcacc tggtgatgac agcacccccg
     2881 cccccaccac cccccgtgta cgagccggtg agctaccatg tccaggagag cttgcaggat
     2941 gagggcgcag agcccacggg ctacagcgcg gagctgtcta gtgagggcat ccgggatgac
     3001 cgcaatgagg agaagcgcat cactgaggca gagaagaacg agcgtgtgca gcggcagctg
     3061 ctgacgctga gcagcgagct gtcccaggcc cgagatgaga ataagaggac ccacaatgac
     3121 atcatccaca acgagaacat gaggcaaggc cgggacaagt acaagacgct gcggcagatc
     3181 cggcagggca acaccaagca gcgcatcgac gagttcgagg ccctgtcgtc gacggtaccg
     3241 cgggcccggg atccaccgga aatgttcaaa aatcttattt ggctgaagga agttgatagc
     3301 acccaggagc ggcttaagga atggaatgtc tcctatggca ctgcactcgt cgccgataga
     3361 cagaccaaag gtagaggcgg tctcggacgg aagtggttga gtcaggaggg cggtctttac
     3421 ttctctttct tgttgaatcc aaaagagttt gaaaatttgt tgcaacttcc actggtcttg
     3481 ggactctccg tgagcgaggc actcgaagag attacagaga taccatttag tctgaagtgg
     3541 cccaacgacg tttattttca ggagaaaaag gtttcagggg tcctttgtga gctcagtaag
     3601 gacaagctga tagtgggaat cggtataaac gtgaatcaaa gagagatccc tgaagagatc
     3661 aaggatcggg ccacaacact ttatgaaatc actggaaagg attgggacag aaaggaggtc
     3721 ttgctgaagg ttctcaagcg aatttccgaa aatctcaaga aatttaagga aaaatccttt
     3781 aaagagttca aaggaaaaat tgagtcaaag atgttgtacc ttggagagga ggttaaactc
     3841 ttgggagaag gcaagattac cgggaaactt gtgggcctga gtgaaaaggg cggtgctttg
     3901 atacttacag aggaaggcat aaaggagata ctgtccggcg aatttagctt gcgacggtca
     3961 gctagcgctt acccttacga tgtaccggat tacgcatagc cggtatcatc aagcttatcg
     4021 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     4081 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     4141 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     4201 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     4261 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     4321 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     4381 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     4441 tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     4501 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     4561 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     4621 tctagagtcg acccgggcgg ccgcttcgag cagacatgat aagatacatt gatgagtttg
     4681 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     4741 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     4801 attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcaag taaaacctct
     4861 acaaatgtgg taaaatcgat aaggatcttc ctagagcatg gctacgtaga taagtagcat
     4921 ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac tccctctctg
     4981 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     5041 cgggcggcct cagtgagcga gcgagcgcgc agccttaatt aacctaattc actggccgtc
     5101 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     5161 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     5221 cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg
     5281 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     5341 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     5401 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     5461 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     5521 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     5581 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     5641 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca
     5701 atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5761 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5821 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5881 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5941 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     6001 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     6061 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     6121 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     6181 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     6241 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     6301 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     6361 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6421 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6481 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6541 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6601 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6661 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     6721 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6781 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6841 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6901 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6961 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     7021 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     7081 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     7141 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     7201 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     7261 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     7321 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     7381 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     7441 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     7501 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     7561 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     7621 gcgaggaagc ggaag
//
