LOCUS       sequence-490440-        6087 bp    DNA     circular SYN 23-OCT-2025
DEFINITION  Bacterial expression of SHANK1 PDZ domain (654-762).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6087)
  AUTHORS   Otani Y, Goodall E, Srinivasan V, Ball NJ, Barnett SFH, Zech T, 
            Saarikangas J, Goult BT
  TITLE     Cyclin-dependent kinase-like 5 (CDKL5) binds to talin and is 
            anchored at the postsynaptic density via direct interaction with PDZ
            domains
  JOURNAL   bioRxiv 2025.08.27.672337; doi: 
            https://doi.org/10.1101/2025.08.27.672337
REFERENCE   2  (bases 1 to 6087)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 23, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2025.08.27.672337; doi: https://doi.org/10.1101/2025.08.27.672337"
COMMENT     Sequence Label: mSHANK1_PDZ
FEATURES             Location/Qualifiers
     source          1..6087
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     209..228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        209..227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /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             267..289
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             300..317
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             318..359
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             360..380
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(788..806)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      802..849
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1087..1105)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1173..1277
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1278..1346,1347..2138)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1496..1515)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2309..2897
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2798..2817
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3051..3068
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3083..3222
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3148..3167)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3308..3330
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3324..3515)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4827..5909)
                     /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     5871..5890
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5910..5987)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
       61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
      121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat atacatatgc
      301 atcatcacca tcaccatggt aagcctatcc ctaaccctct cctcggtctc gattctacgg
      361 aaaacctgta ttttcaggga attgatccct tcaccggtag cgactatatc atcaaagaga
      421 aaaccgttct gctgcagaaa aaggatagcg aaggttttgg ttttgttctg cgtggtgcaa
      481 aagcacagac cccgattgaa gaatttaccc cgacaccggc atttccggca ctgcagtatc
      541 tggaaagcgt tgatgaaggt ggtgttgcat ggcgtgcagg tctgcgtatg ggtgattttc
      601 tgattgaagt taatggtcag aacgtggtga aagttggtca tcgtcaggtt gttaatatga
      661 ttcgtcaagg tggtaatacc ctgatggtta aagttgttat ggttacccgt catccggata
      721 tgtaaaaggg cgagctcaga tccggctgct aacaaagccc gaaaggaagc tgagttggct
      781 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
      841 ggttttttgc tgaaaggagg aactatatcc ggatatcccg caagaggccc ggcagtaccg
      901 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
      961 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
     1021 cgcattaaag ctagcttatc gatgataagc tgtcaaacat gagaattaat tcttgaagac
     1081 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     1141 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     1201 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     1261 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     1321 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     1381 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     1441 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     1501 gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc cgcatacact
     1561 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     1621 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     1681 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     1741 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     1801 agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     1861 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     1921 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     1981 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     2041 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     2101 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     2161 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     2221 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     2281 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     2341 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     2401 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     2461 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     2521 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     2581 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     2641 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     2701 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     2761 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     2821 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     2881 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     2941 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     3001 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     3061 tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     3121 tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     3181 gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca
     3241 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     3301 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg
     3361 cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca
     3421 tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg
     3481 gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt
     3541 ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact
     3601 gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg
     3661 cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta
     3721 ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag
     3781 ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt
     3841 gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt
     3901 gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg
     3961 agcacgatca tgcgcacccg tggccaggac ccaacgctgc ccgagatgcg ccgcgtgcgg
     4021 ctgctggaga tggcggacgc gatggatatg ttctgccaag ggttggtttg cgcattcaca
     4081 gttctccgca agaattgatt ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc
     4141 cgccggcttc cattcaggtc gaggtggccc ggctccatgc accgcgacgc aacgcgggga
     4201 ggcagacaag gtatagggcg gcgcctacaa tccatgccaa cccgttccat gtgctcgccg
     4261 aggcggcata aatcgccgtg acgatcagcg gtccaatgat cgaagttagg ctggtaagag
     4321 ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc tacctgcctg gacagcatgg
     4381 cctgcaacgc gggcatcccg atgccgccgg aagcgagaag aatcataatg gggaaggcca
     4441 tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag cgcgtcggcc gccatgccgg
     4501 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     4561 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     4621 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     4681 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     4741 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     4801 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4861 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     4921 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     4981 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     5041 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     5101 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     5161 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     5221 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     5281 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     5341 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     5401 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     5461 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     5521 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     5581 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     5641 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     5701 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     5761 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     5821 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     5881 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     5941 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     6001 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6061 caccgccgcc gcaaggaatg gtgcatg
//
