LOCUS       sequence-353158-        6157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression in bacterial cells. PX domain, S17-M197.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6157)
  AUTHORS   Gileadi, et al
  TITLE     TREAT-AD plasmids 2021
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 6157)
  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: 
            "unpublished"
COMMENT     Sequence Label: SNX32A-c014
FEATURES             Location/Qualifiers
     source          1..6157
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             12..34
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             54..71
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             930..974
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             975..992
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1045..1063)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1059..1106
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1143..1598
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1230..1249)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1440..1461
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1690..2505)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1690..2505)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2413..2432
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2497..2516
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2627..3215
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3116..3135
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3369..3386
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3401..3540
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3466..3485)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3626..3648
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3642..3833)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4642..5724)
                     /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     5686..5705
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5725..5802)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6008..6027
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6111..6130
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6111..6129
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6130..6154
                     /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)."
ORIGIN
        1 ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgtgcagc agccatcatc
       61 atcatcatca cggcagcggc agcggctctg accaggaggc aaaaccttca actgaggact
      121 tgggggataa gaaggaaggt gaatatatta aactcaaagt cattggacag gatagcagtg
      181 agattcactt caaagtgaaa atgacaacac atctcaagaa actcaaagaa tcatactgtc
      241 aaagacaggg tgttccaatg aattcactca ggtttctctt tgagggtcag agaattgctg
      301 ataatcatac tccaaaagaa ctgggaatgg aggaagaaga tgtgattgaa gtttaccagg
      361 agcaaacggg aggttcggtg gatctgcagg gagacagctc cttacaggtg gagatttctg
      421 acgcagtgag tgagcgggac aaggtgaaat tcactgttca aacaaagagc tgcctccctc
      481 acttcgccca gaccgagttc tcagtcgtgc ggcagcacga ggagttcatc tggctgcatg
      541 atgcctacgt ggagaatgag gagtacgccg gcctcatcat ccccccagcc cctccgaggc
      601 cagactttga ggcttcgagg gaaaagctac agaaattggg cgagggggac agctctgtca
      661 ctcgggaaga gtttgccaag atgaagcagg agctggaagc ggagtacctg gccatcttta
      721 agaagacagt tgcgatgcac gaagtctttc tgcagcgcct ggcggcccac cccaccctgc
      781 gtcgagacca caacttcttt gtgtttttgg aatatggaca ggatctgagt gtccggggga
      841 agaacaggaa ggagctcctc ggagggtttc tgaggaatat tgtgaagtcc gcggatgaag
      901 ccctcatcac gggcatgtga agcggtggcg gtctgaacga catcttcgag gctcagaaaa
      961 tcgaatggca cgaacaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa
     1021 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     1081 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg
     1141 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     1201 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     1261 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     1321 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     1381 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     1441 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     1501 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     1561 taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat
     1621 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     1681 aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg
     1741 attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag
     1801 gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc
     1861 aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg
     1921 agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc agacttgttc
     1981 aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat
     2041 tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac
     2101 aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga
     2161 atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa
     2221 ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt
     2281 cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg
     2341 tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga
     2401 ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt
     2461 taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt
     2521 actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa cgtgagtttt
     2581 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     2641 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     2701 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     2761 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     2821 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     2881 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     2941 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     3001 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     3061 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     3121 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     3181 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     3241 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     3301 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     3361 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     3421 ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa tctgctctga
     3481 tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg
     3541 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     3601 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     3661 tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca
     3721 cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc
     3781 tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc
     3841 tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg
     3901 ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa
     3961 caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc
     4021 ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc
     4081 cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa
     4141 ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac
     4201 gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc
     4261 cgggtcctca acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata
     4321 atggcctgct tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg
     4381 gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag
     4441 cggtcctcgc cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata
     4501 aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg
     4561 actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa
     4621 cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4681 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt
     4741 ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg
     4801 agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat
     4861 ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga
     4921 gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat
     4981 ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt
     5041 ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg
     5101 attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa
     5161 tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
     5221 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
     5281 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
     5341 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
     5401 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
     5461 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
     5521 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
     5581 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
     5641 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
     5701 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
     5761 tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc
     5821 ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg
     5881 ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc
     5941 ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt
     6001 ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc
     6061 cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat taatacgact
     6121 cactataggg gaattgtgag cggataacaa ttcccct
//
