LOCUS       sequence-350566-        6641 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses membrane bound biotin ligase, BioID2, in neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6641)
  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 6641)
  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 hSyn1-Lck-13xLinker-BioID2-BioID2-HA
FEATURES             Location/Qualifiers
     source          1..6641
                     /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
     promoter        465..912
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     intron          959..1091
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1074..1093
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1136..1154
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1472..2170
                     /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             2255..2953
                     /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             2963..2989
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    3014..3602
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3067..3087)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3485..3496)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3666..3787
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3677..3696)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3731..3750
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3949..4078
                     /label=AAV2 ITR
     primer_bind     complement(4097..4114)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4097..4113)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4106..4128)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4255..4710
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4342..4361)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4552..4573
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4736..4840
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4841..4909,4910..5701)
                     /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(5059..5078)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5872..6460
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6361..6380
                     /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 gaagatctga gggccctgcg tatgagtgca agtgggtttt
      481 aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga cccactggac
      541 aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg gaggggaaac
      601 aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt gccttcgccc
      661 ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac gtcactcgcc
      721 ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc cgccggccca
      781 gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca tctgcgctgc
      841 ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt cgtgtcgtgc
      901 ctgagagcgc agtcgagaga acttaagctg cagaagttgg tcgtgaggca ctgggcaggt
      961 aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct tgtcgagaca
     1021 gagaaaactc ttgcgtttct gataggcacc tattggtctt actgacatcc actttgcctt
     1081 tctctccaca ggtgtccact cccagttcaa ttacagctct taaggctaga gtacttaata
     1141 cgactcacta taggctagcc tcgagctcgg atccaccacc atgggctgtg gctgcagctc
     1201 aaaccctgaa gatgactgga tggagaacat tgacgtgtgt gagaactgcc attatccccc
     1261 gccgggtgga ggcgggtctg gaggcggggg tagtggcggg ggtggaagcg ggggtggagg
     1321 cgggtcgggt ggcggaggta gcggaggcgg tggaagtgga ggcggtggaa gtggtggagg
     1381 aggttctggt ggcggaggta gcggtggcgg aggtagcgga ggcggtggaa gtggtggcgg
     1441 aggtagcgga ggcggtggat cggtcgacac gatgttcaaa aatcttattt ggctgaagga
     1501 agttgatagc acccaggagc ggcttaagga atggaatgtc tcctatggca ctgcactcgt
     1561 cgccgataga cagaccaaag gtagaggcgg tctcggacgg aagtggttga gtcaggaggg
     1621 cggtctttac ttctctttct tgttgaatcc aaaagagttt gaaaatttgt tgcaacttcc
     1681 actggtcttg ggactctccg tgagcgaggc actcgaagag attacagaga taccatttag
     1741 tctgaagtgg cccaacgacg tttattttca ggagaaaaag gtttcagggg tcctttgtga
     1801 gctcagtaag gacaagctga tagtgggaat cggtataaac gtgaatcaaa gagagatccc
     1861 tgaagagatc aaggatcggg ccacaacact ttatgaaatc actggaaagg attgggacag
     1921 aaaggaggtc ttgctgaagg ttctcaagcg aatttccgaa aatctcaaga aatttaagga
     1981 aaaatccttt aaagagttca aaggaaaaat tgagtcaaag atgttgtacc ttggagagga
     2041 ggttaaactc ttgggagaag gcaagattac cgggaaactt gtgggcctga gtgaaaaggg
     2101 cggtgctttg atacttacag aggaaggcat aaaggagata ctgtccggcg aatttagctt
     2161 gcgacggtca agcttaggtg gaggaggttc tggaggcggt ggaagtggtg gcggaggtag
     2221 cggtggagga ggttctatgg tgtctaagga attcatgttc aaaaatctta tttggctgaa
     2281 ggaagttgat agcacccagg agcggcttaa ggaatggaat gtctcctatg gcactgcact
     2341 cgtcgccgat agacagacca aaggtagagg cggtctcgga cggaagtggt tgagtcagga
     2401 gggcggtctt tacttctctt tcttgttgaa tccaaaagag tttgaaaatt tgttgcaact
     2461 tccactggtc ttgggactct ccgtgagcga ggcactcgaa gagattacag agataccatt
     2521 tagtctgaag tggcccaacg acgtttattt tcaggagaaa aaggtttcag gggtcctttg
     2581 tgagctcagt aaggacaagc tgatagtggg aatcggtata aacgtgaatc aaagagagat
     2641 ccctgaagag atcaaggatc gggccacaac actttatgaa atcactggaa aggattggga
     2701 cagaaaggag gtcttgctga aggttctcaa gcgaatttcc gaaaatctca agaaatttaa
     2761 ggaaaaatcc tttaaagagt tcaaaggaaa aattgagtca aagatgttgt accttggaga
     2821 ggaggttaaa ctcttgggag aaggcaagat taccgggaaa cttgtgggcc tgagtgaaaa
     2881 gggcggtgct ttgatactta cagaggaagg cataaaggag atactgtccg gcgaatttag
     2941 cttgcgacgg tcagctagcg cttaccctta cgatgtaccg gattacgcat agggatccaa
     3001 ttcgcttatc gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     3061 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     3121 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     3181 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     3241 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     3301 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     3361 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt
     3421 tccttggctg ctcgcctatg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     3481 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     3541 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     3601 gcatcgatac cggaattctg cagatatcta gagtcgaccc gggcggccgc ttcgagcaga
     3661 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     3721 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     3781 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg agatgtggga
     3841 ggttttttaa agcaagtaaa acctctacaa atgtggtaaa atcgataagg atcttcctag
     3901 agcatggcta cgtagataag tagcatggcg ggttaatcat taactacaag gaacccctag
     3961 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     4021 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagcc
     4081 ttaattaacc taattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     4141 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     4201 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc
     4261 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     4321 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     4381 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     4441 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     4501 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     4561 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     4621 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     4681 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     4741 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     4801 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     4861 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4921 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     4981 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     5041 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     5101 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     5161 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     5221 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     5281 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     5341 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     5401 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     5461 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     5521 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5581 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5641 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5701 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     5761 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     5821 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5881 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5941 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     6001 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     6061 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     6121 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     6181 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     6241 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6301 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     6361 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     6421 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     6481 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     6541 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     6601 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa g
//
