LOCUS       sequence-432989-        8538 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of human AP3D1 (1-617) and AP3S1 AP-3 core 
            hemicomplex.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8538)
  AUTHORS   Begley M, Aragon M, Baker RW
  TITLE     A structure-based mechanism for initiation of AP-3 coated vesicle 
            formation.
  JOURNAL   Proc Natl Acad Sci U S A. 2024 Dec 24;121(52):e2411974121. doi: 
            10.1073/pnas.2411974121. Epub 2024 Dec 20.
  PUBMED    39705307
REFERENCE   2  (bases 1 to 8538)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2024-12-24"; volume: "121(52):e2411974121. 
            doi"; pages: " 10.1073/pnas.2411974121. Epub 2024 Dec 20"
COMMENT     Sequence Label: MZB035
FEATURES             Location/Qualifiers
     source          1..8538
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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)."
     RBS             42..64
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1925..1948
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1958..2611
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2569..2591
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     promoter        2744..2762
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2763..2787
                     /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)."
     CDS             3484..3528
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(3566..3584)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3580..3627
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3664..4119
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3751..3770)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3961..3982
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(4234..5025,5026..5094))
                     /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     4857..4876
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5095..5187)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5268..5856
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5757..5776
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6010..6027
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6042..6184
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6110..6129)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6270..6292
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6286..6477)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(7049..8131)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     8093..8112
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(8132..8209)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     8415..8434
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(8522..8538,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
       61 gagatatacc atggctttga aaatggttaa aggctctatt gatcgcatgt tcgataagaa
      121 cttacaggac ttagtccgcg gcattcgtaa ccataaggag gatgaggcga aatacatttc
      181 gcaatgtatc gatgagatta aacaggaact taaacaagac aatattgcag tgaaagcgaa
      241 cgcggtttgt aaactgactt acctgcaaat gctgggttac gatatatcgt gggcggcttt
      301 caatattatt gaggtcatgt cggcatccaa atttacattt aagcgcatcg gttatctggc
      361 agcgtctcag tcatttcatg aggggacaga tgttataatg ctgactacaa atcaaatccg
      421 taaggattta tcttctccca gccagtacga cacgggggtg gcgctgacgg gacttagctg
      481 ttttgttacg cctgatttag ctagagattt ggccaatgac attatgacct tgatgagcca
      541 tacaaaaccg tacatccgca aaaaggcagt ccttattatg tataaggtct ttttgaaata
      601 cccggaatct cttcgtcctg catttcctag attgaaggag aagctggagg acccggaccc
      661 aggtgttcaa tcagccgcag tcaatgtgat atgcgagctg gcccgccgta atcctaagaa
      721 ttatctgtca ctggctccgc ttttctttaa gcttatgacc tcatccacga ataactgggt
      781 tcttataaag atcattaaac tttttggcgc tcttacacct ttggagccca gacttggaaa
      841 aaaattgatt gagccgttaa cgaaccttat ccattctacg agtgccatgt ctctgctgta
      901 cgagtgtgtc aataccgtta ttgccgtatt gattagtctg tcatccggga tgcctaatca
      961 ctctgcttcc atacaattat gcgtacagaa attgagaatc ttgatcgaag attccgacca
     1021 gaatcttaag tacttaggac tgcttgctat gtcaaagata ttaaaaactc acccaaagtc
     1081 cgtccagtcg cacaaagatt tgattttgca atgcctggac gataaggacg aatcgatacg
     1141 tttaagagct ttggacttgt tgtatggcat ggtgtccaaa aagaatttga tggaaatagt
     1201 aaagaagctt atgacgcatg ttgacaaggc tgagggaacg acatatcggg atgaattgct
     1261 gacaaagata attgatatat gttcgcaatc caactaccag tacatcacta atttcgagtg
     1321 gtacattagt atcctggtcg agcttacaag acttgagggc acacggcacg ggcacttgat
     1381 agcagcacaa atgttggacg tggcaataag agtgaaggca atccggaaat ttgcagtatc
     1441 ccagatgagt gctctgctgg actccgccca ccttcttgct agctctacac agagaaacgg
     1501 aatttgcgag gtgttatatg cggctgcttg gatatgcgga gaattctccg agcacttgca
     1561 agaaccacac catacacttg aggcaatgct gcggcctcgc gtgactacct tgcctggaca
     1621 catccaagct gtatacgtcc aaaacgttgt gaagttatat gctagtatcc tgcaacaaaa
     1681 ggaacaagct ggggaggccg aaggcgccca agctgtgaca caactgatgg tagaccgtct
     1741 tccccagttc gtccagtcag ccgatttgga agttcaggaa agagcgtcgt gcattctgca
     1801 gttagtcaaa catatccaaa agctgcaagc aaaggatgtg ccggtagccg aggaagtctc
     1861 agcattgttc gcaggagagt tgaacccggt ggccccaaag gcgcagaaaa aggtcccggt
     1921 cggcctggaa gttctgttcc aggggcccgg gtccggcatg tcccctatac taggttattg
     1981 gaaaattaag ggccttgtgc aacccactcg acttcttttg gaatatcttg aagaaaaata
     2041 tgaagagcat ttgtatgagc gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt
     2101 gggtttggag tttcccaatc ttccttatta tattgatggt gatgttaaat taacacagtc
     2161 tatggccatc atacgttata tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga
     2221 gcgtgcagag atttcaatgc ttgaaggagc ggttttggat attagatacg gtgtttcgag
     2281 aattgcatat agtaaagact ttgaaactct caaagttgat tttcttagca agctacctga
     2341 aatgctgaaa atgttcgaag atcgtttatg tcataaaaca tatttaaatg gtgatcatgt
     2401 aacccatcct gacttcatgt tgtatgacgc tcttgatgtt gttttataca tggacccaat
     2461 gtgcctggat gcgttcccaa aattagtttg ttttaaaaaa cgtattgaag ctatcccaca
     2521 aattgataag tacttgaaat ccagcaagta tatagcatgg cctttgcagg gctggcaagc
     2581 cacgtttggt ggtggcgacc atcctccaaa atcggatctg gtgccgtaat aatgaggatc
     2641 cgaattcgag ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc
     2701 gaacagaaag taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata
     2761 ggggaattgt gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga
     2821 gatatacata tgattaaggc tattttaatc ttcaacaacc atggcaaacc ccgcctgtcc
     2881 aagttctatc agccatattc tgaggatacg caacaacaaa ttatacggga aacctttcac
     2941 cttgtttcta agagagatga aaacgtttgc aattttttgg aaggcggcct tctgatcgga
     3001 ggcagcgata acaaattgat ataccggcac tacgcgacac tgtatttcgt gttctgtgtc
     3061 gattccagtg agtccgagtt gggcatactt gacctgatac aggtcttcgt tgagactttg
     3121 gataagtgct ttgagaacgt ctgcgaattg gacctgatct tccatgtcga taaggtacat
     3181 aacatcttgg ctgagatggt aatgggagga atggtcctgg agacaaacat gaatgagatt
     3241 gtaacccaga tagacgctca aaacaaattg gaaaagtcgg aagctggatt ggcaggagcc
     3301 ccagcgcgtg cggtgtctgc tgttaaaaac atgaatttgc cagaaatacc tagaaatata
     3361 aacatagggg acatctcgat caaggtccca aaccttccct catttaagta gtgacatatg
     3421 gcagatctca attggatatc ggccggccac gcgatcgctg acgtcggtac cctcgagtct
     3481 ggtaaagaaa ccgctgctgc gaaatttgaa cgccagcaca tggactcgtc tactagcgca
     3541 gcttaattaa cctaggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     3601 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat
     3661 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     3721 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     3781 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     3841 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     3901 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3961 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     4021 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     4081 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttc tggcggcacg atggcatgag
     4141 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     4201 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     4261 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     4321 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     4381 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     4441 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     4501 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     4561 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     4621 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     4681 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     4741 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     4801 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     4861 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     4921 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     4981 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     5041 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     5101 cctttttcaa tcatgattga agcatttatc agggttattg tctcatgagc ggatacatat
     5161 ttgaatgtat ttagaaaaat aaacaaatag gtcatgacca aaatccctta acgtgagttt
     5221 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     5281 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     5341 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     5401 ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     5461 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     5521 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     5581 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     5641 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     5701 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     5761 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     5821 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     5881 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     5941 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     6001 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc
     6061 cttacgcatc tgtgcggtat ttcacaccgc atatatggtg cactctcagt acaatctgct
     6121 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     6181 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     6241 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     6301 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     6361 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa
     6421 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     6481 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     6541 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     6601 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     6661 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     6721 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     6781 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     6841 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     6901 tagccgggtc ctcaacgaca ggagcacgat catgctagtc atgccccgcg cccaccggaa
     6961 ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt
     7021 gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     7081 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     7141 ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct
     7201 ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct
     7261 gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca
     7321 ctaccgagat gtccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca
     7381 gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt
     7441 gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct
     7501 gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag
     7561 aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca
     7621 cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag
     7681 agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct
     7741 ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca
     7801 ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac
     7861 ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca
     7921 gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc
     7981 ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag
     8041 aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact
     8101 ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg
     8161 ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga
     8221 cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg
     8281 agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc
     8341 acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc
     8401 cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg
     8461 gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga tcgatctcga tcccgcgaaa
     8521 ttaatacgac tcactata
//
